INFLUENCE OF MI GENES ON GRAFTED SOLANUM LYCOPERSICUM L. CULTIVARS
FOR CONTROL OF TRIALEURODES VAPORARIORUM (INSECTA) IN THE PRESENCE
AND ABSENCE OF MELOIDOGYNE INCOGNITA (NEMATODA).
By
Bahodir Ruzumboevich Eshchanov

A DISSERTATION
Submitted to
Michigan State University
in partial fulfillment of the requirements
for the degree of
Entomology—Doctor of Philosophy
2017

ABSTRACT
INFLUENCE OF MI GENES ON GRAFTED SOLANUM LYCOPERSICUM L. CULTIVARS
FOR CONTROL OF TRIALEURODES VAPORARIORUM (INSECTA) IN THE PRESENCE
AND ABSENCE OF MELOIDOGYNE INCOGNITA (NEMATODA).
By
Bahodir Ruzumboevich Eshchanov
Solanum lycopersicum L (tomato) is native to South and Central America. It is grown worldwide on about ten million acres, yielding circa 185 million tons of fruit on an annual basis. To
make this crop available on a year-round basis, greenhouse tomatoes are commonly grown in
countries such as the Netherlands, Canada and Uzbekistan. Tomato is the second staple
vegetable in Central Asia after potato. About four million tons were produced in Central Asia in
2013. Tomatoes are eaten raw or cooked and are rich in essential vitamins and minerals. Their
color comes from lycopene, which is responsible for the antioxidants reported to alleviate risk to
human health issues such as heart disease, hypertension, congestive heart failure, prostate cancer
and atherosclerosis.
Tomato can be susceptible to numerous insect pests and infectious disease pathogens. Tomato
hornworm (Manduca quinquemaculata), cotton bollworm (Helicoverpa armigera), corn
earworm (Helicoverpa zea), potato aphid (Macrosiphum euphorbiae), green peach aphid (Myzus
persicae), silver leaf whitefly (Bemisia tabaci), and greenhouse whitefly (Trialeurodes
vaporariorum Westwood) are key insect pests of tomato. Whitefly is important because of its
ability to cause direct feeding damage and vector viruses such as tomato yellow leaf curl virus
(TYLCV) of the begomovirus genus.
Numerous tomato cultivars have been developed for desirable, pest, horticultural and food
attributes. Some have been adopted for their pest or pathogen resistance characteristics. For
example, Anahu is a determinant tomato cv that has the Mi gene which confers resistance to

southern root knot nematode (Meloidogyne incognita (Kofoid & White) Chitwood), and crossresistance to potato aphid (Macrosiphum euphorbiae (Thomas)) and sweet potato whitefly
(Bemisia tabaci (Gennadius)). Not all resistant tomato varieties, however, are suitable for
tomato production, or preferred by farmers and consumers. Grafting has the potential to bring all
the essential traits together.
Grafting is a practice used in vegetable production systems to decrease risk to soil-borne
diseases, increase salt tolerance, increase tolerance to other abiotic factors, increase drought
tolerance, increase plant vigor, and increase yield.
The research presented herein was aimed at determining if the grafted susceptible tomato cv
Rutgers on top of resistant tomato cv Anahu will control two important tomato pests. Moreover,
we measured plant biomass (fruit, stem and leaf, and root) of these two tomato cultivara with and
without grafting in order to evaluate grafting impact on plant biomass allocation. Also, we
counted the Type-D trichome density as these structures are thought to be responsible for pest
resistance by acting as mechanical and chemical barriers.
The results show that Anahu is not a good tomato cultivar to control M. incognita and T.
vaporariorum. Anahu was not completely resistant to M. incognita. Moreover, on Anahu,
significant decrease in T. vaporariorum population density occurred only when the M. incognita
was present. Type-D trichome density was not responsible for a reduction of T. vaporariorum
population density on Anahu. Thus, in order to draw a strong conclusion about Mi gene resistant
to T. vaporariorum and resistance translocation from the rootstock to scion in grafted plants,
there is a need for more advanced research, including molecular studies.

This thesis is dedicated to those who supported and believed in me during the course of my
Ph.D. program.
Thank you for always being supportive.

iv

ACKNOWLEDGEMENTS

First and foremost, I would like to thank my Major Professor, Dr. George Bird. Professor.
George always supported me and encouraged my ideas. I would not have been able to finish this
Ph.D. program without him. Moreover, I am grateful to Prof. Bird’s lab for funding my Ph.D.
program. Secondly, I am indebted to Prof. Dr Karim Maridia, Coordinator of the Integrated Pest
Management Collaborative Research Supporting Program for Central Asia (IMP CRSP for
Central Asia). Without Prof. Maredia’s ideas and initiations I would not be here in the USA. I
would like to thank the IPM CRSP team for their activities throughout Central Asia. Especially,
Prof. Dr. Frank Zalom who served as my external committee member and funded a significant
part of my Ph.D. research program. As well as, many thanks go to my Graduate Committee
members: Professor Dr.’s George Bird, Karim Maredia, Larry Gut, William Ravlin, David
Smitley and Frank Zalom. A special thanks to Prof. Dr. Larry Gut for his enormous guidance of
my dissertation writing and additional funding for my Ph.D. research.
Also, superior thank you goes to Heather Lenartson-Kluge, who was very patient, guided me
through all the Entomology Department requirements and paper work during my Ph.D. program.
Furthermore, I am very grateful to good friend and a sports partner, Fred Warner, who was ready
to answer any nematology questions and was helpful during my Ph.D. research program.
Last, but not least, I am obliged for the financial support from Untied States Agency for
International Development (USAID), Integrated Pest Management Collaborative Research
Support Program (IPM CRSP), University of California-Davis and Michigan State University.
Financial support for the first two years of my graduate degree program was provided by the
IPM CRSP Innovation Lab sponsored Central Asia Regional IPM Project under the U.S. Agency

v

for International Development Cooperative Agreement No: EPP-A-00-0400016-00. I am also
grateful to the MSU Asian Studies Center for providing the Delia Koo Fellowship for the Spring
Semester 2017. Also, I am very thankful to Organization of International Student and Scholars
for financial supports for International Family Health Insurance Award and International Student
Emergency Tunisian Findings.
Next, I express my deep appreciation to the greenhouse and growth-chamber facilities
management system at Michigan State University, who provided me generous access and help
with the facilities throughout my Ph.D. research.
Also, I would like to thank my all friends and students in both the Nematology Laboratory and
Dr. McCullough’s Laboratory for their help and support. Especially I would like to expand my
gratuity to Rasulbek Makhmudovich Makhmudov and Anthony Thierry Cognon who assisted me
during this long journey.
Finally, I would like to thank my lovely wife Manzura Atamuratovna Sharipova and my children
Azamjon Eshchanov and Diyora Eshchanov. Thank you very much, my love for your sleepless
nights for nematode and whitefly counting. I love you.

vi

TABLE OF CONTENTS

LIST OF TABLES ......................................................................................................................... xi
LIST OF FIGURES ..................................................................................................................... xiv
KEY TO SYMBOLS AND ABBREVIATIONS ....................................................................... xvii
INTRODUCTION .......................................................................................................................... 1
CHAPTER 1: IMPACT OF GRAFTING AND PRUNING ON RESOURCE PARTITIONING
OF GREENHOUSE SOLANUM LYCOPERSICUM L. CVS RUTGERS AND ANAHU: WITH
SPECIAL REFERENCE TO CENTRAL ASIA ............................................................................ 4
Abstract ............................................................................................................................... 4
Introduction ......................................................................................................................... 5
Materials and Methods........................................................................................................ 8
Plant material .......................................................................................................... 8
Grafting ................................................................................................................... 9
Greenhouse ............................................................................................................. 9
Pruning .................................................................................................................. 10
Data collection ...................................................................................................... 10
Data analysis ......................................................................................................... 11
Results ............................................................................................................................... 11
Tomato Plant Fresh Weight Biomass Partitioning of Rutgers and Anahu and
Impact of Grafting................................................................................................. 11
Lightly Pruning Impact on Plant Fresh Weight Biomass Partitioning ................. 12
Heavy Pruning Impact on Plant Fresh Weight Biomass Partitioning ................... 13
Dry Weights of Rutgers and Anahu Among Grafting Treatments ....................... 14
Discussion ......................................................................................................................... 15
Conclusion ........................................................................................................................ 16
APPENDICES .................................................................................................................. 18
Appendix A. Statistical Tables ............................................................................. 19
Appendix B. Figures ............................................................................................. 23
REFERENCES ................................................................................................................. 29
CHAPTER 2: BIOMASS ALLOCATION ASSOCIATED WITH HOMO-GRAFTED AND
HETERO-GRAFTED SOLANUM LYCOPERSICUM L. CVS RUTGERS AND ANAHU
GROWN UNDER FIELD CONDITIONS. .................................................................................. 32
Abstract ............................................................................................................................. 32
Introduction ....................................................................................................................... 33
Materials and methods ...................................................................................................... 34
Plant material ........................................................................................................ 35

vii

Grafting ................................................................................................................. 35
Field ...................................................................................................................... 36
Data colocation ..................................................................................................... 36
Data analysis ......................................................................................................... 36
Results ............................................................................................................................... 37
Impact of Grafting on Biomass Partitioning After 50 Days Under Field
Conditions ............................................................................................................. 37
Impact of Grafting on Tomato Plant Biomass Partitioning at After 100 Days
Under Field Conditions ......................................................................................... 38
Discussion ......................................................................................................................... 39
Conclusion ........................................................................................................................ 40
APPENDICES .................................................................................................................. 42
Appendix A. Statistical Tables ............................................................................. 43
Appendix B. Figures ............................................................................................. 45
REFERENCES ................................................................................................................. 46
CHAPTER 3: INFLUENCE OF GRAFTING SOLANUM LYCOPERSICUM L. CVS ANAHU
AND RUTGERS ON PLANT BIOMASS PARTITIONING IN THE PRESENCE AND
ABSENCE OF MELOIDOGYNE INCOGNITA (NEMATODA): WITH THE SPECIFIC
REFERENCE TO CENTRAL ASIA............................................................................................ 50
Abstract ............................................................................................................................. 50
Introduction ....................................................................................................................... 51
Materials and Methods...................................................................................................... 53
Experimental Design............................................................................................. 54
Plant materials....................................................................................................... 54
Pruning .................................................................................................................. 55
Grafting ................................................................................................................. 55
Meloidogyne. incognita Population Density......................................................... 56
Data Analysis ........................................................................................................ 57
Results ............................................................................................................................... 57
Grafting Impact on Fruit Fresh Tissue Weight ..................................................... 57
Impact of Pruning on Plant Population Density Under Greenhouse Conditions .. 58
Grafting Impact on Stem/Leaf Fresh Weight........................................................ 59
Grafting Impact on Root Fresh Weight ................................................................ 59
Meloidogyne incognita Population Density.......................................................... 59
Biomass Partitioning ............................................................................................. 61
Non-pruned Experiment........................................................................................ 61
Lightly-pruned Experiment ................................................................................... 62
Heavily-pruned Experiment .................................................................................. 64
Discussion ......................................................................................................................... 64
Conclusion ........................................................................................................................ 68
APPENDICES .................................................................................................................. 70
Appendix A. Statistical Tables ......................................................................................... 71

viii

Appendix B. Figures ............................................................................................. 75
REFERENCES ................................................................................................................. 86
CHAPTER 4: IMPACT OF SOLANUM LYCOPERSICUM L. GRAFTING ON
TRIALEURODES VAPORARIORUM WESTWOOD (INSECTA) DEVELOPMENT UNDER
GROWTH CHAMBER CONDITIONS: WITH SPECIAL REFERENCE TO THE MI GENE,
TYPE-D TRICHOMES AND CENTRAL ASIA. ........................................................................ 90
Abstract ............................................................................................................................. 90
Introduction ....................................................................................................................... 90
Materials and Methods...................................................................................................... 92
Experimental Design............................................................................................. 92
Plant materials....................................................................................................... 92
Grafting ................................................................................................................. 93
Growth Chamber ................................................................................................... 93
T. vaporariorum Population Density Bioassay ..................................................... 93
Type-D trichome density ...................................................................................... 94
Data Analysis ........................................................................................................ 94
Results ............................................................................................................................... 95
Nymphs ................................................................................................................. 95
Pre-adults and Hatched Pupa Cases ...................................................................... 95
Total Population Density of T. vaporariorum ....................................................... 95
Type-D Trichome Density after 7 and 25 days of Transplant into Plastic Pots ... 96
Discussion ......................................................................................................................... 97
Conclusion ...................................................................................................................... 100
APPENDICES ................................................................................................................ 101
Appendix A. Statistical Tables ........................................................................... 102
Appendix B. Figures ........................................................................................... 105
REFERENCES ............................................................................................................... 108
CHAPTER 5: JOINT IMPACT OF MELOIDOGYNE INCOGNITA (KOFOID AND
WHITE1919) CHITWOOD 1949 AND WESTWOOD ON GRAFTED SOLANUM
LYCOPERSICUM L. CULTIVARS: WITH SPECIAL REFERENCE TO THE MI GENE. .. 112
Abstract ........................................................................................................................... 112
Introduction ..................................................................................................................... 112
Materials and Methods.................................................................................................... 114
Experimental Design........................................................................................... 114
Plant Materials .................................................................................................... 115
Grafting ............................................................................................................... 115
Growth Chamber ................................................................................................. 116
Trialeurodes vaporariorum Population Density Bioassay .................................. 116
Meloidogyne incognita Population Density........................................................ 116
Trichome Types .................................................................................................. 117
Data Analysis ...................................................................................................... 118

ix

Results ............................................................................................................................. 118
Greenhouse Whitefly .......................................................................................... 118
Type-D Trichome Density in the Presence of M. incognita ............................... 118
Stem/leaf Biomass in the Presence and Absence of Trialeurodes vaporariorum 119
Root Biomass in the Presence and the Absence of the Trialeurodes vaporariorum
............................................................................................................................. 120
Meloidogyne incognita Population Development in the Presence and the Absence
of Trialeurodes vaporariorum ............................................................................. 121
Biomass Partitioning in the Presence and Absence of Trialeurodes vaporariorum
and Meloidogyne incognita................................................................................. 121
Relationship between Solanum lycopersicum L Root Fresh Weight and
Trialeurodes vaporariorum.................................................................................. 122
Discussion ....................................................................................................................... 123
Conclusion ...................................................................................................................... 125
APPENDICES ................................................................................................................ 127
Appendix A. Statistical Tables ........................................................................... 128
Appendix B. Figures ........................................................................................... 134
Appendix C. Specimens ...................................................................................... 140
REFERENCES ............................................................................................................... 141
CONCLUSION ........................................................................................................................... 144

x

LIST OF TABLES

Table 1. Impact of homo-grafting and hetero-grafting on biomass partitioning of tomato
cvs Rutgers and Anahu............................................................................................................... 19
Table 2. Impact of homo-grafting and hetero-grafting on biomass partitioning of tomato
cvs Rutgers and Anahu with light pruning. ............................................................................. 20
Table 3. Impact of homo-grafting and hetero-grafting on biomass partitioning of tomato
cvs Rutgers and Anahu with heavy pruning. ........................................................................... 21
Table 4. Influence of tomato grafting on stem/leaf dry weight biomass partitioning
associated with three pruning regimes, including: non-pruned, lightly pruned, and heavily
pruned. ......................................................................................................................................... 22
Table 5. Influence of tomato grafting on root dry weight biomass partitioning with three
pruning regimes, including: non-pruned, lightly pruned, and heavily pruned. ................... 22
Table 6. Influence of Solanum lycopersicum L. grafting on stem/leaf dry, root fresh, and
root dry tissue biomass partitioning after 50 days under field conditions. ........................... 43
Table 7. Impact of grafting on biomass partitioning of Solanum lycopersicum cvs Rutgers
and Anahu after 100 days under field conditions. ................................................................... 43
Table 8. Influence of Solanum lycopersicum L. grafting on stem/leaf and root dry weight
biomass partitioning after 100 days under field conditions. ................................................... 44
Table 9. Influence of Solanum lycopersicum L. grafting on fruit tissue in the presence and
absence of Meloidogyne incognita.at initial nematode population densities of Pi=0, Pi=1000,
and Pi=5,000 eggs per experimental unit. A. Non-pruned, B. Lightly pruned, C. Heavily
pruned. ......................................................................................................................................... 71
Table 10. Influence of Solanum lycopersicum L. grafting on stem/leaf tissue in the presence
and absence of Meloidogyne incognita at initial nematode population densities of Pi=0,
Pi=1000, and Pi= 5000 eggs per experimental unit. A. Non-pruned, B. Lightly pruned, C.
Heavily pruned. ........................................................................................................................... 72
Table 11. Influence of Solanum lycopersicum L. grafting on root tissue in the presence and
absence of Meloidogyne incognita at initial nematode population densities of Pi=0, Pi=1000,
and Pi= 5000 eggs per experimental unit. A. Non-pruned, B. Lightly pruned, C. Heavily
pruned. ......................................................................................................................................... 73
Table 12. Influence of Solanum lycopersicum L. grafting on final female population
densities of Meloidogyne incognita associated with initial M. incognita egg population

xi

densities of Pi=1,000 and Pi=5,000 per experimental unit. A. Non-pruned, B. Lightly pruned, C.
Heavily pruned. ........................................................................................................................... 74
Table 13. Influence of Solanum lycopersicum L grafting on T. vaporariorum life stage
population densities. ................................................................................................................. 102
Table 14. Solanum lycopersicum L Type-D trichome density at three plant heights seven
days after transplanting into plastic pot. Lower region (leaves from second branch), Midregion (leaves from a mid-region branch), and Upper region (leaves from last branch one
before merry stem). ................................................................................................................... 102
Table 15. Impact of Solanum lycopersicum L grafting on Type-D Trichrome density seven
days after transplanting into plastic pot. Means for six replicates of each grafting
treatments. Three leaf samples were taken from three leaves of the each tomato plant. .. 103
Table 16. Impact of Solanum lycopersicum L grafting on Type-D trichome density at three
plant heights 25 days after transplanting into plastic pot. Lower region (leaves from second
branch), Mid-region (leaves from a mid-region branch), and Upper region (leaves from last
branch one before merry stem). .............................................................................................. 103
Table 17. Impact of Solanum lycopersicum L grafting on Type-D Trichrome density 25
days after transplanting into plastic pot. Means for six replicates of each grafting
treatments. Three leaf samples were taken from three leaves of the each tomato plant. .. 104
Table 18. T. vaporariorum population density on grafted Solanum lycopersicum after 25
days in the presence of Meloidogyne incognita. ..................................................................... 128
Table 19. Grafted Solanum lycopersicum Type-D trichome density after seven days in the
presence of Meloidogyne incognita.......................................................................................... 128
Table 20. Solanum lycopersicum L Type-D Trichome density at three plant heights after
seven days in the presence of M. incognita. ............................................................................ 129
Table 21. Grafted Solanum lycopersicum Type-D trichome density after 25 days in the
presence of Meloidogyne incognita.......................................................................................... 129
Table 22. Solanum lycopersicum L Type-D Trichome density at plant three plant heights
after 25 days in the presence of Meloidogyne. incognita. ...................................................... 129
Table 23. Grafted Solanum lycopersicum fresh weight of stem and leaf tissue after 24 days
in the presence of Meloidogyne incognita. .............................................................................. 130
Table 24. Grafted Solanum lycopersicum dry weights of stem and leaf tissue after 24 days
in the presence of with Meloidogyne incognita. ..................................................................... 130
Table 25. Impact of Trialeurodes vaporariorum to grafted Solanum lycopersicum fresh
weight of stem/leaf after 24 days in the presence of Meloidogyne incognita. ...................... 131

xii

Table 26. Impact of Trialeurodes vaporariorum on grafted Solanum lycopersicum dry
weight of stem and leaf tissue after 24 days in the presence of Meloidogyne incognita. .... 131
Table 27. Grafted Solanum lycopersicum fresh weight of root tissue after 24 days in the
presence of Meloidogyne incognita.......................................................................................... 131
Table 28. Grafted Solanum lycopersicum dry weight of root tissue after 24 days in the
presence of Meloidogyne incognita.......................................................................................... 132
Table 29. Impact of Trialeurodes vaporariorum on grafted Solanum lycopersicum root
fresh weight after 24 days in the presence of Meloidogyne incognita. ................................. 132
Table 30. Impact of Trialeurodes vaporariorum on grafted Solanum lycopersicum root dry
weight of root tissue after 24 days in the presence of Meloidogyne incognita..................... 133
Table 31. Meloidogyne incognita final female population density on six grafted S.
lycopersicum L. treatments. ..................................................................................................... 133
Table 32. Meloidogyne incognita final female population density on six grafted S.
lycopersicum L treatments in the presence of Trialeurodes vaporariorum. ....................... 133
Table 33. List of voucher specimens. ....................................................................................... 140

xiii

LIST OF FIGURES

Figure 1. Young non-pruned tomato plants under greenhouse research conditions. .......... 23
Figure 2. Maturing and lightly pruned tomato plants under greenhouse research
conditions. .................................................................................................................................... 24
Figure 3. Mature fruit bearing and heavily pruned tomato plants under greenhouse
research conditions. .................................................................................................................... 25
Figure 4. Anahu and Rutgers tomato biomass partitioning signatures for non-grafted,
homo-grafted and hetero-grafted plants. .................................................................................. 26
Figure 5. Influence of light pruning on Anahu and Rutgers tomato biomass partitioning
signatures for non-grafted, homo-grafted and hetero-grafted plants. ................................... 27
Figure 6. Influence of heavy pruning on Anahu and Rutgers tomato biomass partitioning
signatures for non-grafted, homo-grafted and hetero-grafted plants. ................................... 28
Figure 7. Aerial view of the 2015 tomato grafting field research site at the Department of
Entomology Research Farm at Collins Road, Michigan State University. ........................... 45
Figure 8. Tomato grafting research site at the Michigan State University, Department of
Entomology Collins Road Research Farm. .............................................................................. 45
Figure 9. Grafted and non-pruned Solanum lycopersicum L. experiment under greenhouse
conditions. .................................................................................................................................... 75
Figure 10. Grafted Solanum lycopersicum L. heavily pruned and lightly pruned
greenhouse trials. Left: heavily pruning experiment, Right: lightly pruned experiment.
There were 15 experiment units per bench. ............................................................................. 75
Figure 11. Solanum lycopersicum L. root systems: Left. Susceptible tomato cv Rutgers
under initial egg population density (Pi) of 1000 per experimental unit. Right. Susceptible
tomato cv Rutgers under initial egg population densities (Pi) of 0 per experimental unit. .. 76
Figure 12. Root staining method with nylon fine mashed cloth to prevent loss of small
pieces of root tissue. .................................................................................................................... 77
Figure 13. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers in the absence of M. incognita (Pi=0). A. Non-grafted, B. Homo-grafted, C. Heterografted .......................................................................................................................................... 78

xiv

Figure 14. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers in the presence of M. incognita (Pi =1000). A. Non-grafted, B. Homo-grafted, C.
Hetero-grafted. ............................................................................................................................ 79
Figure 15. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers in the absence of M. incognita (Pi = 0) and with lightly pruning. A. Non-grafted, B.
Homo-grafted, C. Hetero-grafted. ............................................................................................. 80
Figure 16. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers in the presence of M. incognita (Pi =1000) and with lightly pruning. A. Non-grafted,
B. Homo-grafted, C. Hetero-grafted. ........................................................................................ 81
Figure 17. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers with M. incognita (Pi = 5000) and with lightly pruning. A. Non-grafted, B. Homografted, C. Hetero-grafted. ......................................................................................................... 82
Figure 18. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers with heavily pruning and in the absence of M. incognita (Pi = 0). A. Non-grafted, B.
Homo-grafted, C. Hetero-grafted. ............................................................................................. 83
Figure 19. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers in the presence of M. incognita (Pi =1000) with heavily pruned. A. Non-grafted, B.
Homo-grafted, C. Hetero-grafted. ............................................................................................. 84
Figure 20. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers with heavily pruning and in the absence of M. incognita (Pi = 5000). A. Non-grafted,
B. Homo-grafted, C. Hetero-grafted. ........................................................................................ 85
Figure 21. Mini-insectary for T. vaporariorum population density bioassay. A: Young
grafted tomato seedling in a mini-insectary. B: Experimental units after inoculation with T.
vaporariorum in a completely randomized design under growth chamber conditions. .... 105
Figure 22. Tomato leaf fixed with tape to make the leaf flat to assist counting whiteflies and
tomato trichomes. ...................................................................................................................... 106
Figure 23. Relationship between lower region Type-D trichome density and total T.
vaporariorum population density............................................................................................ 107
Figure 24. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers in the presence of M. incognita (Pi = 5000). ................................................................. 134
Figure 25. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers in the presence of M. incognita (Pi =5000) and T. vaporariorum. ............................. 135
Figure 26. Relationship between S. lycopersicum root fresh weight and T. vaporariorum
population density per plant. ................................................................................................... 136

xv

Figure 27. Relationship between S. lycopersicum L cv Anahu and cv Rutgers root fresh
weight and T. vaporariorum population density per plant. ................................................. 137
Figure 28. Relationship between S. lycopersicum L homo-grafted Anahu and homo-grafted
Rutgers root fresh weight and T. vaporariorum population density per plant. ................. 138
Figure 29. Relationship between S. lycopersicum L hetero-grafted Anahu and heterografted Rutgers root fresh weight and T. vaporariorum population density per plant. .... 139

xvi

KEY TO SYMBOLS AND ABBREVIATIONS

cv(s) - cultivar(s) (variety)
g - grams
Pi - nematode initial population density per plant
Pf - female final population density per plant
RH - relative humidity
Stem/leaf - stem and weight measured together

xvii

INTRODUCTION
The Michigan State University, Central Asia, Integrated Pest Management, Collaborative
Research Support Program (IPM CRSP) was initiated in 2004, with funding from USAID. It
included three nations: Kyrgyzstan, Tajikistan, and Uzbekistan. The project had two phases.
The first phase consisted of conducting landscape ecology research in Tajikistan, BioLab
enhancement in Uzbekistan and farmer training in Kyrgyzstan. The activities were implemented
by local partners and three project managers. The second phase of the project consisted of
development and implementation of IPM programs for three key-staple Central Asia food crops:
potato in Kyrgyzstan, tomato in Uzbekistan and wheat in Tajikistan. Moreover, the Central Asia
IPM project initiated capacity building for Central Asia through bringing young scientists from
Central Asia to the U.S.A. for a graduate-level education. Thus, as one of these individuals, my
Ph.D. dissertation is directly interlinked with the MSU IPM CRSP for Central Asia.
On a global basis, tomato production under both greenhouse and field conditions has numerous
pest problems. One of the most difficult to control in greenhouse production is the greenhouse
whitefly (Trialeurodes vaporariorum Westwood, 1856). Current, T. vaporariorum control in
tomato production is based mainly on the use of chemical pesticides. For example, in
Uzbekistan and elsewhere, tomatoes grown in greenhouses are sprayed with insecticides weekly
in order to maintain the T. vaporariorum population density under the damage threshold level.
These chemicals are not only dangerous to humans, but their use also can cause long-term
environmental damage that is difficult to remediate.
There are several alternates to chemical control, including the use of resistant or tolerant
cultivars and hybrids. Nevertheless, these does not always keep T. vaporariorum populations
under the damage threshold. Moreover, pests can overcome resistance due to their rapid rates of
reproduction. Another obstacle is the time and cost of breading resistant varieties and hybrids.
1

Another option for pest and disease control in tomato production is vegetable grafting. Grafting
can be used to incorporate several desired characteristics of different varieties together into a
single plant. Vegetable grafting is a surgical procedure. It has been used extensively in East and
South Asia for many ears. Currently, vegetable grafting is being introduced into many European
and North American countries. Thus, it should be possible to use grafted tomato plants to
maintain T. vaporariorum population densities below the threshold level. Moreover, chemical
insecticides, resistant varieties and grafting, mentioned above, can be combined together for
control of T. vaporariorum under greenhouse and field tomato production conditions.
My Ph.D. dissertation entitled “Influence of Mi genes on grafted Solanum lycopersicum L.
cultivars for control of Trialeurodes vaporariorum (Insecta) in the presence and absence of
Meloidogyne incognita (Nematoda)” consists of five chapters. Each chapter was developed from
the results of hypotheses tested during my Ph.D. research program. Plans are to have each
chapter published individually in preview science journals. The hypotheses related to the impact
of tomato grafting, T. vaporariorum, M. incognita and trichomes on biomass partitioning. Since
each chapter is self-contained with a comprehensive literature review this information is not
repeated in this general introduction. The five chapters include:
Chapter 1 “Impact of Grafting and Pruning on Resource Partitioning of Greenhouse Solanum
lycopersicum L. cvs Rutgers and Anahu: With Special Reference to Central Asia”
Chapter 2 “Biomass allocations associated with homo-grafted and hetero-grafted Solanum
lycopersicum L. cvs Rutgers and Anahu grown under field conditions.”
Chapter 3 “Influence of Grafting Solanum lycopersicum L. cvs Anahu and Rutgers on Plant
Biomass Partitioning in the Presence and Absence of Meloidogyne incognita (Nematoda): With
the specific reference to Central Asia.”

2

Chapter 4 “Impact of Solanum lycopersicum L. Grafting on Trialeurodes vaporariorum
Westwood (Insecta) Development Under Growth Chamber Conditions: With Special Reference
to the Mi Gene, Type-D Trichomes and Central Asia.”
Chapter 5 “Joint Impact of Meloidogyne incognita (Kofoid and White1919) Chitwood 1949 and
Westwood on Grafted Solanum lycopersicum L. Cultivars: With Special Reference to the Mi
Gene.”

3

CHAPTER 1: IMPACT OF GRAFTING AND PRUNING ON RESOURCE
PARTITIONING OF GREENHOUSE SOLANUM LYCOPERSICUM L. CVS RUTGERS
AND ANAHU: WITH SPECIAL REFERENCE TO CENTRAL ASIA
Abstract
Numerous tomato cultivars have been developed with desirable horticultural and food attributes.
Some have been adopted due to their pest or pathogen resistance characteristics. However, not
all resistant tomato cultivars are favorable for tomato fruit production. Plant grafting allows for
combining two or more desired characteristics into one plant. Tomato cultivars, however, vary
in how they allocate biomass among their shoot, root and fruit tissues. Understanding biomass
partitioning is important for the selection of appropriate cultivars for grafting and greenhouse
production systems. Rutgers and Anahu are two important tomato cultivars that have desirable
characteristics for tomato production in central Asia. The major objective of this research was to
determine the effects of homo-grafting and hetero-grafting of the two cultivars on plant biomass
partitioning among stem/leaf, root and fruit tissue under greenhouse conditions. Rutgers
partitioned the majority of its biomass to fruit tissue and less to stem/leaf tissue and root tissues.
In contrast, Anahu allocated the majority of its biomass to stem/leaf tissue. Homo-grafted
Rutgers shifted its biomass from fruit to stem/leaf tissue, while homo-grafted Anahu shifted its
biomass to fruit tissue. Hetero-grafted Rutgers shifted its biomass even more towards stem/leaf
tissue, whereas hetero-grafted Anahu retained its original biomass partitioning signature. Thus,
Anahu appears to be manipulating the scion’s biomass partitioning when used as a rootstock.
The impact was most pronounced with respect to fruit tissue or stem/leaf tissue. Root tissue
resource partitioning was not impacted by grafting or pruning. An additional objective was to
determine the impact of light (optimal) or heavy pruning (single stem) on the partitioning of
plant biomass of these tomato cultivars under greenhouse conditions. Heavy pruning entailed
removing all suckers and leaves up to the first fruit cluster and resulted in Anahu allocating more
4

of its biomass to fruit tissue compared to stem/leaf tissue. Under heavy pruning, Rutgers’
biomass partitioning signature was not impacted by homo-grafting or hetero-grafting. The
biomass signature for lightly pruned Rutgers and Anahu was similar to its non-pruned signature.
Understanding biomass partitioning at the cultivar level is important before selecting a rootstock
for commercial production under greenhouse conditions. In general, Anahu is not a good
rootstock for tomato fruit production under greenhouse conditions. Future research with Rutgers
and Anahu grafting and pruning needs to be evaluated in regards to biotic and abiotic stress
factors.
Introduction
Solanum lycopersicum L (tomato) is native to South and Central America (Peralta et al. 2006). It
is grown world-wide on about ten million acres, yielding circa 185 million tons of fruit on an
annual basis (FAOSTAT, 2011). To make this crop available on a year-round basis, greenhouse
tomatoes are commonly grown in countries such as the Netherlands, Canada and Uzbekistan.
Tomato is the second staple vegetable in Central Asia after potato. About four million tons were
produced in Central Asia in 2013 (FAOSTAT, 2014). Tomatoes are eaten raw or cooked and are
rich in essential vitamins and minerals. Their color comes from lycopene, which is responsible
for the antioxidants reported to alleviate risk to human health issues such as heart disease,
hypertension, congestive heart failure, prostate cancer and atherosclerosis (Marry, Datta, Bedi, &
Kaur, 2015).
Tomato can be susceptible to numerous insect pests and infectious disease pathogens. Tomato
hornworm (Manduca quinquemaculata), cotton bollworm (Helicoverpa armigera), corn
earworm (Helicoverpa zea), potato aphid (Macrosiphum euphorbiae), green peach aphid (Myzus
persicae), silver leaf whitefly (Bemisia tabaci), and greenhouse whitefly (Trialeurodes
vaporariorum Westwood) are key insect pests of tomato. Whitefly is important because of its
5

ability to cause direct feeding damage and vector viruses such as tomato yellow leaf curl virus
(TYLCV) of the begomovirus genus. TYLCV causes substantial tomato yield loss annually on a
global basis (Zalom, 2012). Another key greenhouse pathogen is the southern root-knot
nematode (Meloidogyne incognita). In addition to being a pathogen, it can also pre-dispose
plants to soil-borne wilt diseases such as those caused by Verticillium dahliae and Fusarium
oxysporium f. sp. Lycopersici (Zalom, 2012).
Numerous tomato cultivars have been developed for desirable horticultural and food attributes.
Some have been adopted for their pest or pathogen resistance characteristics. Another way to
achieve resistance is through grafting. Plant grafting has been used for many years, including as
a means of managing pests and pathogens through resistance (Lee, 1994). Vegetable grafting
was first practiced widely early in the 20th century (Lee, 1994), as described in Grafted of
Herbaceous Vegetable and Ornamental Crops in Scientia Horticulturae Ashita (1927, 1930,
1934). The technique allows combining two or more desired vegetable characteristics into one
plant. In addition, grafting is used to control soil-borne diseases, increase slat tolerance, increase
nutrient uptake, and increase plant yield.
Tomato cultivars vary in how they allocate biomass among their shoot, root and fruit tissues.
Understanding biomass partitioning is important for the selection of appropriate cultivars for
greenhouse production systems. Biomass partitioning has been studied in detail on non-grafted
tomato plants. It has been shown that a biomass increase in one part of plant effects the biomass
allocated to other parts of the plant (Enquist & Niklas, 2002; Heuvelink, 1997; Marcelis,
Heuvelink, & Goudriaan, 1998). This phenomenon has been studied as a means of increasing
fruit yield, but less extensively for impacts on overall plant biomass partitioning (Dimitrios et al.,
2009, Charles and Xin, 2012). The impact of grafting on biomass partitioning has not been

6

studied under greenhouse conditions (Marcelis, 1993). It has also not been studied in regards to
homo-grafted, and hetero-grafted plants. In homo-grafting, the scion and root tissue are from the
same cultivar, but different plants are grafted. In hetero-grafting, the scion and root tissue come
from two different cultivars.
Rutgers and Anahu are important tomato cultivars that have desirable horticultural and food
characteristics. Rutgers is a determinant cultivar that is typically favored by farmers for its good
quality and high productivity. Anahu is a determinant cv that has the Mi gene which confers
resistant to Meloidogyne incognita (Kofoid & White) Chitwood, and cross-resistance to
Macrosiphum euphorbiae (Thomas) (Goggin, Williamson, & Ullman, 2001; Vos et al., 1998)
and sweet potato whitefly, Bemisia tabaci (Gennadius) (Nombela, Williamson, & Muñiz, 2003;
Goggin et al., 2001). There are several Mi genes in cultivated and wild tomatoes, including Mi-1
to Mi-9. Mi-1 has seven homologues, Mi-1.1 through Mi-1.7 (Seah, Telleen, & Williamson,
2007). Among the Mi-1 homologues, Mi-1.2 is the only functional nematode resistant gene for
M. incognita (Milligan et al., 1998). Mi-1.2 is the gene present in cv Anahu. Anahu is one of
the parents for all other commercial cultivars containing the Mi gene. Anahu is resistant to three
different key pests of tomato. It usually grows to a height of 3-4 feet and produces
approximately five kg of tomato fruit under field conditions (Bobisud, Martin, & Sekioka, 1996).
Pruning tomato plants can be used to increase air flow between leaves and decrease relative
humidity, resulting in less favorable conditions for pathogens (Ferrandino, 1999). Moreover,
Kanyomeka and Shivute (2005) (Kanyomeka & Shivute, 2005)(Kanyomeka & Shivute,
2005)(Kanyomeka & Shivute, 2005)(Kanyomeka & Shivute, 2005)(Kanyomeka & Shivute,
2005) suggested that pruning increases fruit quality, plant health and makes plants less prone to
pest attack. There are two common methods of pruning, simple pruning and Missouri pruning.

7

Simple pruning consists of pinching-off suckers when the suckers are young. This does not take
much effort and does not reduce the plant’s photosynthetic area very much. With the Missouri
system, two leaves are left and the apical meristems of suckers are pinched-off (Ferrandino,
1999). Moreover, pruning is used in greenhouse tomato production to increase the number of
plants per unit area. There are several degrees of severity of pruning tomato plants. Pruning
may be light (optimal) or heavy. In light pruning, 50% of the suckers are removed up to the first
fruit cluster. In heavy pruning in a two stem system, all suckers are removed from the ground to
the first fork (Olson, 1989). Pruning effects have been studied in non-grafted as well as in
grafted tomato plants. On non-grafted tomatoes, heavily-pruned single-stem plants resulted in
low total yield and increased fruit size (Olson, 1989). Double stem pruned tomato plants yielded
more than 3 or 4-stem pruned plants (Mourão, Teixeira, Brito, Ferreira, & Moura, 2014).
The aim of this research was to evaluate the potential of using grafted cv Rutgers and cv Anahu
tomato plants for greenhouse tomato production in Central Asia. The specific objectives of the
research were 1) to determine the effects of homo and hetero grafting of tomato cv Rutgers and
cv Anahu on partitioning of plant biomass among stem/leaf, root and fruit tissue under
greenhouse conditions and 2) to determine the effects of light (optimal) and heavy pruning on
tomato plant biomasses partitioning of plant biomass on the tomato cultivars under greenhouse
conditions.
Materials and Methods
Plant material
The tomato biomass resource partitioning study was conducted under greenhouse conditions at
Michigan State University using the cultivars Anahu and Rutgers. It consisted of five
experiments, each with six grafting treatments and five replicates of each treatment. A single
tomato cv Anahu (University of Hawaii Seed Program) or cv Rutgers (Sustainable Seed Co.) was
8

sown in each cell in 50 or 72 cell seedling trays containing plant growth media (SUREMIX
professional growing media, Michigan Grower Products Inc.) and held in a growth chamber
maintained at 26 C for 16 hours and 20 C for eight hours, day and night, respectively. The
seedlings were watered every other day. Anahu seeds were planted two days before Rutgers
seeds to ensure the same seedling diameter for grafting. This time-interval was based on the
results of preliminary research. After 14 days in the growth chamber, the seedlings were
appropriate for grafting.
Grafting
Rootstocks and scions were cut below the cotyledon at a 45-65 degree angle using a Miter-Cut
Grafting Knife (Johnny's Selected Seeds). Scions and rootstocks were clipped together with
silicon tubes to hold the rootstock and scion together until the grafting wound healed. Silicon
clipper (Hydro-Gardens, Colorado Springs, CO) size varied from 1.5-2.5 mm, depending on
seedling diameter. During graft wound healing, a humidifier (Air Innovations Model #
HUMID06 1.37-gal. Ultrasonic Digital Humidifier) was used to control the relative humidity
(RH). The grafted plants were maintained in a growth chamber for seven days at 24-26 C. The
RH was decreased gradually from 90% after three days, to 85% two days later, to 75% one day
later and then to 65% for the final day. After 7 days, the grafted seedlings were moved to the
laboratory for three days of acclimatization. The six grafting treatments were: 1) non-grafted
Anahu, 2) non-grafted Rutgers, 3) homo-grafted Anahu, 4) homo-grafted Rutgers, 5) heterografted Anahu (Anahu scion and Rutgers rootstock) and 6) hetero-grafted Rutgers (Rutgers scion
and Anahu rootstock).
Greenhouse
Following acclimatization, the 21-day-old tomato plants were transplanted into five-gallon
plastic pots filled with sterilized sandy soil and placed in a greenhouse maintained at 26-28 C.
9

They were organized in a randomized block design and watered as necessary for 125 to 135 days
(Figure 1-Figure 3).
Pruning
Experiments were conducted using both pruned and non-pruned tomato plants. The grafted and
non-grafted plants in the first experiment were not pruned. Those in the second experiment were
lightly pruned and those in the third experiment were heavily pruned. The lightly pruned
experiment was conducted three times. Simple pruning methods were used to remove suckers
from the node. Each sucker was pinched of by hand when it was small, or with garden clippers
when suckers were larger. Under the lightly pruned system, all the suckers and leaves were
removed up to first fruit cluster. On heavily pruned plants, all of the suckers were removed from
the entire plant and leaves were removed to a 3-4-foot height. The greenhouse space for the
research accommodated 60 non-pruned plants; whereas, it accommodated 90 lightly and 90
heavily pruned tomato plants.
Data collection
Throughout the experiments, ripe fruit (90% or more of the skin red in color) were harvested
weekly. Each fruit was weighed individually on a 200g scale on the day of harvest. After 125
days, all remaining fruit were harvested and included in the yield as fruit fresh weight. The
stem/leaf tissues also were collected and weighed right after collection on a five kg scale. Fresh
weights were recorded and the tissue placed in a drying oven at 30-35 C the same day for dry
weight determination. The dry mass was weighed with the same scale after seven days of oven
drying. Tomato plant roots were collected, washed and dried in a salad leaf spin dryer for 2-3
minutes. Fresh weight of root tissue was determined on a 200 g scale. The roots were placed in
a drying oven at the same temperature used for the stem/leaf tissue. Root dry weights were
measured after seven days of drying with the same scale.
10

Data analysis
The data were analyzed using SPSS 24 Grad Primmum Pack. Data distribution normality was
checked with a histogram plot for skewness of the distribution. If the data were positively
skewed, they were transformed using a natural Log or Log10 prior to analysis. If the data were
negatively skewed, the Square Root Method was applied. If data were not normalized, the nonparametric Kruskal-Wallis Test was used as the substitute for a One-Way ANOVA. If the OneWay ANOVA showed a significant difference at the α=0.05, means were separated using the
Tukey’s Test. If the non-parametric Kruskal-Wallis Test was significant at the α=0.05, means
were separated using the Dunn’s Test.
Results
Tomato Plant Fresh Weight Biomass Partitioning of Rutgers and Anahu and Impact of
Grafting
There were significant differences among the six grafted treatments for stem/leaf fresh weight in
the non-pruned experiment (Appendix A. Statistical Tables
Table 1). The hetero-grafted Rutgers stem/leaf tissue was significantly greater than that of
hetero-grafted Anahu. Non-grafted Rutgers and Anahu, however, had different biomass
partitioning signatures and allocated their biomass inversely. Rutgers allocated the majority of
its biomass to fruit tissue (Figure 4). On the other hand, Anahu allocated the greatest amount of
biomass to stem/leaf tissue. In both tomato cultivars, root tissue comprised the least amount of
biomass compared to other parts of the plant. Biomass partitioning in Rutgers changed when it
was homo-grafted, with a gain in stem/leaf tissue and a loss of fruit tissue. Stem/leaf tissue
biomass comprised 52% of the total biomass, while 10% of fruit biomass was lost. Root tissue
biomass allocation, however, did not change on homo-grafted Rutgers, compared to non-grafted
Rutgers. Conversely, Anahu shifted its biomass towards fruit after homo-grafting. An average of

11

9% of the biomass shifted to fruit tissue from stem/leaf tissue, compared to non-grafted Anahu.
Root tissue biomass was essentially unchanged, decreasing by only 1%. . Hetero-grafting
Rutgers shifted 13% of its biomass to stem/leaf tissue, compared to non-grafted Rutgers. Root
tissue biomass did not change compared to non-grafted Rutgers. With hetero-grafted Anahu,
biomass partitioning shifted towards stem/leaf tissue compared to non-grafted Anahu. Stem/leaf
tissue comprised 3% more of the biomass compared to non-grafted Anahu. Root biomass
partitioning was not altered by hetero-grafting.
Lightly Pruning Impact on Plant Fresh Weight Biomass Partitioning
There were no significant difference between the six grafted treatments after light pruning on
fruit, stem/leaf, and root tissue (Table 2). Moreover, with light pruning, there was very little
change in biomass partitioning of Rutgers and Anahu, compare to the non-pruned experiment
(Figure 4Figure 5). Rutgers retained it biomass signature after light pruning, allocating the
highest proportion of biomass to fruit tissue. Light pruning, however, decreased fruit biomass
by 3% and 10%, compared to non-pruned and heavy pruned treatments. Root tissue biomass did
not change after light pruning. Anahu also retained its biomass signature after light pruning,
allocating the highest proportion of biomass to stem/leaf tissue (Figure 4Figure 5). However,
there was a slight 4% increase in fruit biomass after lightly pruning. Root tissue biomass
allocation did not change. The biomass of homo-grafted Rutgers that was light pruned consisted
of 55% stem/leaf tissue and 35% fruit tissue (Figure 4Figure 5). This represented a 9% decrease
in fruit biomass compared to non-pruned plants where 3% shifted to stem/leaf tissue and 6%
shifted to root tissue after light pruning. After light pruning, the biomass of homo-grafted Anahu
consisted of 64% stem/leaf tissue and 31% fruit tissue (Figure 4Figure 5). This represented a 9%
shift in biomass partitioning from fruit to stem/leaf tissue after homo-grafting Anahu. In hetero-

12

grafted Rutgers after light pruning, biomass was almost equally allocated between stem/leaf
tissue and fruit tissue, 49% and 47%, respectively (Figure 4Figure 5). This represented a 6%
shift in biomass partioning from leaf/stem tissue to fruits after light pruning. Root tissue biomass
did not change. Light pruning produced a similar change in biomass partitioning of heterografted Anahu, with about a 7% increase in fruit biomass (Figure 4Figure 6). Root tissue biomass
did not change.
Heavy Pruning Impact on Plant Fresh Weight Biomass Partitioning
There were no significant differences among fruit, stem/leaf, and root tissue on the six grafted
treatments after heavy pruning (Table 3). Heavy pruning, however, drastically change biomass
partitioning among fruit, stem/leaf and root tissues. The heavily pruned Rutgers biomass
signature was similar to the non-pruned Rutgers biomass partitioning signature (Figure 4Figure
6). The greatest amount of biomass was allocated to fruit tissue; there was 7% more fruit
biomass following heavy pruning. Stem/leaf tissue biomass was second greatest and root tissue
was the least. Anahu did not retain its fresh biomass partitioning signature following heavy
pruning, losing 15% of stem/leaf tissue and allocating 16% more biomass to fruit (Figure
4Figure 6). Heavy pruning of homo-grafting Rutgers resulted in a substantial shift in biomass
from fruit towards stem/leaf tissue, with 19% more biomass allocated to stem/leaf tissue (Figure
4Figure 6). Root tissue biomass was essentially unchanged, decreasing by only 1%. Heavily
pruned homo-grafted Anahu plants allocated 41% of their biomass to stem/leaf tissue and 55% to
fruit tissue. This represented a 9% increase in allocation of biomass to fruit (Figure 4-Figure 6).
Root tissue of homo-grafting Anahu was not impacted by heavy pruning. Heavy pruning of
hetero-grafted Rutgers plants significantly altered the partitioning of resources, shifting 23% of
their biomass from leaf/stem to fruit (Figure 4Figure 6). Root tissue biomass was essentially

13

unchanged, decreasing by only 1%. Biomass partitioning of hetero-grafted Anahu also was
substantially shifted by heavy pruning, with 24% more biomass allocated to fruit (Figure
4,Figure 6).
Dry Weights of Rutgers and Anahu Among Grafting Treatments
Dry weight among the six grafting treatments was significantly different (F=4.332, P=0.006) in
the non-pruned experiment. The greatest dry weight of stem/leaf tissue occurred in Heterografted Anahu. The least amount of dry weight was recorded on hetero-grafted Rutgers (Table
4). Hetero-grafted Anahu produced about twice as much and significantly more (P=0.015)
stem/leaf tissue than hetero-grafted Rutgers. Root tissue dry weight biomass was not
significantly different among the six grafted treatments (Table 5). The least amount of root
tissue dry weight was observed on hetero-grafted Rutgers. The greatest amount of root tissue dry
weight biomass was observed on non-grafted Anahu.
There were no significant differences in stem/leaf tissue dry biomass among the six treatments
when the plants were lightly pruned (Table 4). The greatest amount of stem/leaf tissue was
observed on homo-grafted Anahu after light pruning. The lowest amount of stem/leaf tissue
biomass occurred on non-grafted Rutgers. Root tissue dry weight biomass following light
pruning also was not significant different among the six treatments (Table 5). The greatest
amount of root tissue dry weight was observed on hetero-grafted Anahu. The lowest amount of
root dry weight was recorded on non-grafted Anahu.
Heavy pruning of Rutgers and Anahu did not significantly impact stem/leaf tissue biomass
among the six treatments (Table 4). The greatest amount of stem/leaf tissue biomass was
observed on non-grated Anahu after heavy pruning. The lowest tissue dry weight was recorded
on homo-grafted Anahu. Root tissue dry weight also was not significantly different among the
six grafted treatments following light pruning. The greatest amount of root dry tissue was
14

associated with homo-grafted Anahu. The lowest root tissue dry weight was recorded on nongrafted Anahu.
Discussion
Rutgers and Anahu are important tomato cultivars that have desirable characteristics for tomato
production in Central Asia. Rutgers is favored by farmers for its good quality and high
productivity, while Anahu has the Mi gene which confers resistant to some important nematode
and insect pests (Nombela et al., 2003; Goggin et al., 2001). Plant grafting allows for combining
the desired characteristics of each into one plant. A crucial first step in the grafting process is to
ensure that fruit production is not negatively impacted when the cultivars are combined.
Understanding the biomass partitioning signature of each cultivar involved should be a
component of plant grafting systems.
Homo-grafting and hetero-grafting of Rutgers and Anahu impacted the partitioning of plant
biomass among stem/leaf, root and fruit tissue under greenhouse conditions. It is well
documented that non-grafted tomato plants vary in how they allocate biomass (Brian and Karl,
2002; Heuvelink, 1996, 1999; Marcelis et al., 1998). Higashide et al. (2014) did detailed
comparison between two scions and two rootstocks after grafting where hetero-grafting and
homo-grafting impacted scion biomass partitioning differently. Comparison and shift of biomass
partitioning between non-grafted tomato cultivars, homo-grafted, and hetero-grafted plants,
however, is a novel approach to understanding the biomass allocation of the grafted plant.
In comparing non-grafted, homo-grafted and hetero-grafted tomato plants, I found that Anahu
and Rutgers allocated their biomass differently among fruit, stem/leaf, and root fresh tissue.
Non-grafted Rutgers partitioned the majority of its biomass to fruit tissue and less to stem/leaf
and root tissues. In contrast, non-grafted Anahu allocated the majority of its biomass to
stem/leaf tissue. I considered these as the biomass partitioning signatures of tomato cultivars
15

Rutgers and Anahu. The effect of grafting and pruning on the two cultivars was compared to
these characteristic biomass partitioning signatures. Homo-grafting and hetero-grafting shifted
the allocation of plant biomass among the various plant parts. In homo-grafted Rutgers, biomass
shifted from fruit to stem/leaf tissue, while in homo-grafted Anahu fruit tissue comprised more
of the biomass. Hetero-grafting of Rutgers shifted biomass even more towards stem/leaf tissue;
whereas, hetero-grafted Anahu retained its biomass partitioning signature. There was little
change in root biomass allocation following grafting. With the exception of root crop
vegetables, the root system is only a small portion of the vegetable plant’s biomass (Marcelis,
1991). Overall, Anahu appears to be manipulating the scion’s biomass partitioning when it is
used as the rootstock.
Pruning also had a substantial impact on biomass partitioning of grafted and non-grafted Anahu
and Rutgers tomato plants. Furthermore, the effects varied depending on the extensiveness of
pruning. This is consistent with previous findings that plant biomass allocation varies depending
on factors such as the age of the plant and environmental conditions (Marcelis et al., 1998).
Heavy pruning generally had a beneficial effect on Anahu by shifting the allocation of biomass
to fruit production. There was a 16-24% gain in fruit biomass for non-grafted Anahu and homografted or hetero-grafted Anahu. Light pruning of hetero-grafted Rutgers also shifted a high
percentage of biomass to fruit. However, 16% more biomass was allocated to stem/leaf tissue
following heavy pruning of homo-grafted Rutgers. Light pruning had less of an effect on
biomass partitioning. In our studies, the greatest increase in fruit biomass was associated after
heavy pruning.
Conclusion
Overall, grafting and pruning impacted the biomass allocation of homo-grafted and heterografted Rutgers and Anahu plants in different ways. Homo-grafted Anahu appears to not be a
16

good option for greenhouse tomato production due to grafting incompatibility and adverse effect
on fruit yield. In contrast, hetero-grafted Rutgers may by a good choice as it improves yield
under greenhouse conditions after heavy pruning. We propose that hetero-grafting can be useful
in developing tomato plants for commercial greenhouse production systems. Light pruning can
also benefit greenhouse tomato production by increasing yield per plant or total production by
allowing space for more plants per unit area. We were able to place three times more plants in
the greenhouse after pruning, resulting in an increase in total crop yield. Additionally, pruning
of tomato plants can provide more favorable growing conditions and possibly make tomato
plants less susceptible to risk of disease and yield loss associated with various pathogens and
pests (Ferrandino, 1999).

17

APPENDICES

18

Appendix A. Statistical Tables
Table 1. Impact of homo-grafting and hetero-grafting on biomass partitioning of tomato
cvs Rutgers and Anahu.
Non-pruned
Treatments
Non-grafting

Fruit (g)

Stem/leaf (g)

Root ()

Mean

SE

Mean

SE

Mean

SE

Rutgers

2161.00

466.85

1685.22 ab1

21.95

159.6

14.82

Anahu

1441.51

340.20

2350.62 ab

186.19

217.6

31.62

Anahu/Anahu

1760.13

463.11

1838.62 ab

216.77

142

31.11

Rutgers/Rutgers

1678.12

603.80

1975.02 ab

194.81

158.4

14.12

Rutgers/Anahu

1007.96

538.93

1629.82 a

154.76

120

17.65

Anahu/Rutgers

1345.29

520.74

2483.42 b

283.29

195.2

18.62

Homo-grafting

Hetero-grafting

1

Column means followed by different letters are significantly (P = 0.05) different.

19

Table 2. Impact of homo-grafting and hetero-grafting on biomass partitioning of tomato
cvs Rutgers and Anahu with light pruning.
Lightly pruning
Treatments
Non-grafting

Fruit

Stem/leaf

Root

Mean (g)

SE

Mean (g)

SE

Mean (g)

SE

Rutgers

1690.82

320.82

1537.82

107.44

151.8

8.9

Anahu

1262.04

305.43

1676.02

188.01

156.75

56.87

Anahu/Anahu

841.85

157.65

1686.22

240.63

128.2

31.74

Rutgers/Rutgers

1076.28

214.68

1660.22

53.77

285.8

89.92

Rutgers/Anahu

1686.38

299.15

1620.42

211.92

136

28.61

Anahu/Rutgers

1219.74

122.51

1621.42

104.67

214

57.55

Homo-grafting

Hetero-grafting

20

Table 3. Impact of homo-grafting and hetero-grafting on biomass partitioning of tomato
cvs Rutgers and Anahu with heavy pruning.
Heavy pruning
Treatments
Non-grafting

Fruit

Stem/leaf

Root

Mean (g)

SE

Mean (g)

SE

Mean (g)

SE

Rutgers

1269.38

222.17

861.82

55.29

82.8

11.43

Anahu

1433.93

214.56

1024.02

129.11

90.8

28.45

Anahu/Anahu

1180.17

271.63

839.42

75.57

88

6.58

Rutgers/Rutgers

1738.76

317.39

923.42

111.87

93.6

22.79

Rutgers/Anahu

1810.15

148.61

873.82

87.75

73

14.49

Anahu/Rutgers

1222.26

240.73

827.62

72.52

76.4

11.17

Homo-grafting

Hetero-grafting

21

Table 4. Influence of tomato grafting on stem/leaf dry weight biomass partitioning
associated with three pruning regimes, including: non-pruned, lightly pruned, and heavily
pruned.
Stem/leaf
Non-pruned
Lightly pruned
Heavy pruned
Grafting
Mean (g)
SE
Mean (g)
SE
Mean (g)
SE
1
Anahu
38.58
33.77
16.61
460.62 ab
213.82
143.02
Anahu/Anahu
54.76
35.60
7.84
317.02 ab
248.62
103.62
Rutgers/Anahu
28.30
32.65
6.62
283.42 a
229.62
121.42
Anahu/Rutgers
60.35
19.52
7.80
508.62 b
227.22
116.82
Rutgers/Rutgers
50.87
15.08
13.84
333.82 ab
209.82
127.62
Rutgers
4.03
10.58
9.78
319.02 ab
202.42
118.22
1

Column means followed by different letters are significantly (P = 0.05) different.
Table 5. Influence of tomato grafting on root dry weight biomass partitioning with three
pruning regimes, including: non-pruned, lightly pruned, and heavily pruned.
Root
Non-pruned
Lightly pruned
Heavy pruned
Grafting
Mean (g)
SE
Mean (g)
SE
Mean (g)
SE
Anahu
3.42
2.40
0.62
25.75
13.38
5.66
Anahu/Anahu
4.09
13.03
1.84
18.31
28.25
10.71
Rutgers/Anahu
1.89
10.96
1.26
16.80
30.30
7.29
Anahu/Rutgers
2.92
20.20
1.00
22.97
50.49
7.39
Rutgers/Rutgers
2.92
16.58
2.10
20.94
49.15
9.67
Rutgers
1.39
5.06
1.57
19.18
27.61
8.15

1

Column means followed by different letters are significantly (P = 0.05) different.

22

Appendix B. Figures
Figure 1. Young non-pruned tomato plants under greenhouse research conditions.

23

Figure 2. Maturing and lightly pruned tomato plants under greenhouse research
conditions.

24

Figure 3. Mature fruit bearing and heavily pruned tomato plants under greenhouse
research conditions.

25

Figure 4. Anahu and Rutgers tomato biomass partitioning signatures for non-grafted,
homo-grafted and hetero-grafted plants.

26

Figure 5. Influence of light pruning on Anahu and Rutgers tomato biomass partitioning
signatures for non-grafted, homo-grafted and hetero-grafted plants.

27

Figure 6. Influence of heavy pruning on Anahu and Rutgers tomato biomass partitioning
signatures for non-grafted, homo-grafted and hetero-grafted plants.

28

REFERENCES

29

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Barrett, C. E., Zhao, X., & Hodges, A. W. (2012). Cost Benefit Analysis of Using Grafted
Transplants for Root-knot Nematode Management in Organic Heirloom Tomato Production.
HortTechnology, 22(2), 252–257.
Bobisud, C. A., Martin, S. P., & Sekioka, T. T. (1996). Field testing bacterial wilt-resistant
tomato somaclones. Journal of the American Society for Horticultural Science, 121(3), 384–387.
Davis, R. M., Pernezny, K., & Broome, J. C. (2012). Tomato Health Management. APS Press.
Enquist, B. J., & Niklas, K. J. (2002). Global allocation rules for patterns of biomass partitioning
in seed plants. Science, 295(5559), 1517–1520.
FAOSTAT. (2011). FAOSTAT. Retrieved May 23, 2017, from
http://www.fao.org/faostat/en/#home
Ferrandino, F. (1999). Pruning tomatoes. The Taunton Press, New Haven. Retrieved from
http://srmtenv.org/web_docs/outreachnews/PruningTomatoes.pdf
Frank Zalom (2012). Tomato Arthropod Pest Management in "Compendium of tomato diseases
and pests." Jones, J. B., Zitter, T. A., Momol, T. M., & Miller, S. A. (2012). Am Phytopath
Society. Retrieved from
Goggin, F. L., Williamson, V. M., & Ullman, D. E. (2001). Variability in the response of
Macrosiphum euphorbiae and Myzus persicae (Hemiptera: Aphididae) to the tomato resistance
gene Mi. Environmental Entomology, 30(1), 101–106.
Heuvelink, E. (1996). Dry Matter Partitioning in Tomato: Validation of a Dynamic Simulation
Model. Annals of Botany, 77(1), 71–80.
Heuvelink, E. (1997). Effect of fruit load on dry matter partitioning in tomato. Scientia
Horticulturae, 69(1), 51–59.
Higashide, T., Nakano, A., & Yasuba, K. (2014). Yield and Dry Matter Production of a Japanese
Tomato “Momotaro York” Are Improved by Grafting onto a Dutch Rootstock “Maxifort.”
Journal of the Japanese Society for Horticultural Science, 83(3), 235–243.
Kanyomeka, L., & Shivute, B. (2005). Influence of pruning on tomato production under
controlled environments. Agricultura Tropicaet Subtropica, 32(2), 79–81.
Lee, J. (1994). Cultivation of grafted vegetables. I. Current status, grafting methods, and
benefits. HortScience, 29(4), 235–239.
Marcelis, L. F. M. (1991). Simulation of biomass allocation in greenhouse crops-a review. In
International Workshop on Greenhouse Crop Models 328 (pp. 49–68).

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Marcelis, L. F. M. (1993). Fruit growth and biomass allocation to the fruits in cucumber. 1.
Effect of fruit load and temperature. Scientia Horticulturae, 54(2), 107–121.
Marcelis, L. F. M., Heuvelink, E., & Goudriaan, J. (1998). Modelling biomass production and
yield of horticultural crops: A review. Scientia Horticulturae, 74(1–2), 83–111.
Marry, A., Datta, S., Bedi, O., & Kaur, M. (2015). Pharmacological and Therapeutic uses of
Lycopene. International Journal of Recent Advances in Pharmaecetical Research, 5(3), 181–191.
Milligan, S. B., Bodeau, J., Yaghoobi, J., Kaloshian, I., Zabel, P., & Williamson, V. M. (1998).
The root knot nematode resistance gene Mi from tomato is a member of the leucine zipper,
nucleotide binding, leucine-rich repeat family of plant genes. The Plant Cell, 10(8), 1307–1319.
Mourão, I., Teixeira, J., Brito, L. M., Ferreira, M. E., & Moura, M. L. (2014). Pruning System
Effect on Greenhouse Grafted Tomato Yield and Quality. In G. Rahmann & U. Aksoy (Eds.),
Building Organic Bridges (Vol. 3, pp. 941–944). Braunschweig, Germany: Johann Heinrich von
Thünen-Institut.
Nombela, G., Williamson, V. M., & Muñiz, M. (2003). The root-knot nematode resistance gene
Mi-1.2 of tomato is responsible for resistance against the whitefly Bemisia tabaci. Molecular
Plant-Microbe Interactions, 16(7), 645–649.
Olson, S. M. (1989). Pruning method effects on yield, fruit size, and percentage of marketable
fruit of “Sunny”and “Solar Set”tomatoes. In Proc. Fla. State Hort. Soc (Vol. 102, pp. 324–326).
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tomatoes. Retrieved from http://tgc.ifas.ufl.edu/vol56/html/vol56featr.htm
Savvas, D., Papastavrou, D., Ntatsi, G., Ropokis, A., Olympios, C., Hartmann, H., & Schwarz,
D. (2009). Interactive effects of grafting and manganese supply on growth, yield, and nutrient
uptake by tomato. HortScience, 44(7), 1978–1982.
Seah, S., Telleen, A. C., & Williamson, V. M. (2007). Introgressed and endogenous Mi-1 gene
clusters in tomato differ by complex rearrangements in flanking sequences and show sequence
exchange and diversifying selection among homologues. Theoretical and Applied Genetics,
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Nature Biotechnology, 16(13), 1365–1369.

31

CHAPTER 2: BIOMASS ALLOCATION ASSOCIATED WITH HOMO-GRAFTED AND
HETERO-GRAFTED SOLANUM LYCOPERSICUM L. CVS RUTGERS AND ANAHU
GROWN UNDER FIELD CONDITIONS.
Abstract
Biomass partitioning of tomato (Solanum lycopersicum L.) was studied under field and
greenhouse conditions. Tomato grafting is of interest because of its potential for disease and pest
control through resistance. Grafted tomato resource partitioning into stem, leaf, root, and fruit
tissue, however, has not been thoroughly investigated. Understanding allocation of biomass in
grafted tomato plants is important for selection of optimal rootstocks for grafting. Two
experiments, each with six treatments with cvs Anahu and Rutgers were used to study the
impacts of homo-grafting and hetero-grafting of tomato biomass partitioning under field
conditions. The research consisted of a partial-season and a full-season experiment. The partialseason experiment was terminated after 50 days. Under these conditions, stem/leaf dry weight,
root fresh weight and root dry weight of homo-grafting and hetero-grafted cv Anahu were
significantly less than those of non-grafted plants. The same was true for root fresh weight, but
not dry weight of cv Rutgers. Homo-grafting, but not hetero-grafting cv Rutgers resulted in less
stem/leaf dry weight compared to non-grafted plants. The full-season experiment was
terminated after 100 days at the end of fruit harvest. Under these conditions, grafting had no
significant impact on the partitioning of stem/leaf dry weight, root fresh weight, root dry weight
or fruit fresh weight. Overall, grafting did not adversely impacted plant development and fruit
yield. In order to evaluate the complete biological impact of homo-grafting and hetero-grafting,
it will be necessary to conduct the research in the presence of key biological (e.g. disease or pest)
or physical stress factors.

32

Introduction
Solanum lycopersicum L. (tomato) and other vegetable crop biomass partitioning has been
studied extensively in regards to shoot system dry matter (stem/leaf and fruit tissue) under
greenhouse and field conditions (Heuvelink 1999; Heuvelink 1997; Marcelis 1991; Marcelis,
Heuvelink, and Goudriaan 1998; Enquist and Niklas 2002). Biomass allocation models are
based on dry or fresh weights of tissue to simulate how plants integrate their biomass to different
parts the plant. Cultivars have specific biomass partitioning signatures (Chapter 1).
Relatively few biomass partitioning studies have been conducted with grafted vegetables.
Rootstocks are usually selected to overcome various biotic and abiotic stress factors. Following
grafting, biomass resources can be directed to parts of a plant other than fruit tissue. For
instance, rootstock can manipulate scion growth (Kudo and Harada 2007). Thus, biomass
partitioning is important for selecting appropriate rootstocks for grafting.
Grafting is a practice used in vegetable production systems to decrease risk to soil-borne
diseases (Na Liu et al. 2009), increase salt tolerance (Huang et al. 2009; He et al. 2009;
Martinez-Rodriguez et al. 2008), increase tolerance to other abiotic factors (Schwarz et al. 2010),
increase drought tolerance (Altunlu and Gul 2011), increase plant vigor (Venema et al. 2008;
Estañ et al. 2005), and increase yield (Turhan et al. 2011; Huang et al. 2009; Khah et al. 2006b).
Grafting has been used for many years in Japan and Korea to overcome soil-borne diseases (Lee
1994). Interest in grafting vegetable crops is not only increasing in Asian countries but in North
America as well. For instance, 40 million seedlings are grafted in North America each year.
These are produced mainly in Ontario and British Columbia, Canada (Kubota et al. 2008).
Moreover, grafting is environmental friendly and interest/demand is increasing for organic food
production. In addition, pesticide regulations have increased in conventional food production
systems, making alternatives more attractive. It has been demonstrated that grafting can
33

decrease the use of chemical pesticides in both organic and conventional crop production (Rivard
and Louws 2008), making grafting a very important technique in both types of vegetable
production systems.
One of the benefits of vegetable grafting is increased yield and overall plant biomass. There are
numerous studies about the impact of grafting on tomato fruit yield and vigor under
environmental stress conditions (López-Pérez et al. 2006; Khah et al. 2006a). There is, however,
very limited information about the impact of homo- grafting and hetero-grafting on fruit yield
and plant vigor in the absence of environmental stress. Despite the benefit of grafting to
overcome diseases or increase yield, it is important to understand biomass partitioning of a
specific plant cultivar before selecting it as a rootstock. The objective of this study is to
determine the impact of homo-grafted and hetero-grafted tomato cultivars Rutgers and Anahu on
the fresh and dry biomass partitioning of under field conditions: with special reference to tomato
fruit tissue fresh weight, stem/leaf tissue fresh and dry weight, root tissue fresh, and dry weight
of both young plants and plants grown through harvest. An additional objective is to evaluate
Anahu as a compatible rootstock for hetero-grafting for use under field conditions.
Materials and methods
This research was conducted under field conditions at the Michigan State University,
Department of Entomology, Collins Road Research Farm in East Lansing, MI, 42.690396, 84.496250 (Figure 7). It consisted of a partial-season and a full-season experiment, each with six
grafting treatments. The partial season experiment had 15 replicates of each treatment and was
terminated after 50 days. The full-season trial had 10 replicates of each treatment and was
terminated 100 days after transplanting. The six grafting treatments were: 1) non-grafted Anahu,
2) non-grafted Rutgers, 3) homo-grafted Anahu, 4) homo-grafted Rutgers, 5) hetero-grafted

34

Anahu (Anahu scion and Rutgers rootstock) and 6) hetero-grafted Rutgers (Rutgers scion and
Anahu rootstock).
Plant material
The production of grafted tomato plants for the research consisted of planting a single tomato cv
Anahu (University of Hawaii Seed Program) or cv Rutgers (Sustainable Seed Co.) seed in each
cell in 50 or 72 cell seedling trays containing plant growth media (SUREMIX professional
growing media, Michigan Grower Products Inc.) and placed in an environmental chamber
maintained at 26 C for 16 hours and 20 C for eight hours, day and night, respectively. The
seedlings were watered every other day. Anahu seeds were planted two days before Rutgers
seeds to ensure the same seedling diameter at the time of grafting. This time-interval was based
on preliminary research results. After 14 days in the environmental chamber, Anahu and Rutgers
seedlings were 1.5-2.5 mm in diameter, an appropriate size for grafting.
Grafting
Rootstocks and scions were cut below the cotyledon at a 45-65 degree angle using a Miter-Cut
Grafting Knife (Johnny's Selected Seeds). Scions and rootstocks were clipped together with
silicon tubes (Silicon clipper, Hydro-Gardens, Colorado Springs, CO) until the grafting wound
healed. Silicon clipper size varied from 1.5-2.5 mm depending on seedling diameter. A
humidifier (Air Innovations Model # HUMID06 1.37-gal) was used to control the relative
humidity during graft wound healing. The grafted plants were held in an environmental chamber
for seven days at 24-26 C temperature. The relative humidity was decreased gradually from 90%
after three days, to 85% two days later, to 75% one day later and then to 65% for the final day.
After seven days, the grafted seedlings were moved to the laboratory for three days of
acclimatization

35

Field
The young tomato plants were transplanted into three raised beds: 60 centimeter wide, 30
centimeter high (Figure 8). The distance from the center of the row was 120 centimeters and
each row was 10 meters in length. There were 40 centimeters between each plant within the
row. Each treatment was replicated fifteen times in the 50-day experiment and ten times in the
100-day. The plants were maintained in a completely randomized design under field conditions.
Data colocation
The plants were watered as necessary using a drip irrigation system and fertilized every other
week. Tomato harvest in the full-season experiment began on September 19th, 2015. Soon after
the first harvest, the plants were infected with late blight and the remaining tomatoes were
harvested on October 8, 2015. The stem/leaf fresh weights were not recorded. Immediately after
collection, stem/leaf tissue was placed in a drying oven at 30-35 C. Tomato plant roots were
washed and fresh weight recorded. The roots were immediately placed in a drying oven at the
same temperature used for the stem/leaf tissue. After seven days, the stem/leaf and root dry
weights were recorded.
Data analysis
The data were analyzed using SPSS 24 Grad Primmum Pack. The data distribution normality
was checked with a histogram plot for skewness. If the data were positively skewed, a natural
Log or Log10 transformation was used. If the data were negatively skewed, the Square Root
Method was applied. If the data were not normalized, the non-parametric Kruskal-Wallis Test
was used as the substitute for One-Way ANOVA. If the One-Way ANOVA was significant at
α=0.05, it was followed by Tukey’s Test. When the non-parametric Kruskal-Wallis Test was
significant at the α=0.05, it was followed with Dunn’s Test.

36

Results
Impact of Grafting on Biomass Partitioning After 50 Days Under Field Conditions
There were significant differences in stem/leaf dry, root fresh, and root dry weights among the
six treatments of 50-day-old tomato plants maintained under field conditions (Table 6). The
stem/leaf dry weight was significantly different among the six treatments (F5,14=9.286, p=0.001).
Non-grafted Rutgers stem/leaf dry weight was similar to non-grafted Anahu. Non-grafted Anahu
had the greatest amount of stem/leaf dry weight among the six treatments. There was, however,
no significant difference between the non-grafted treatments. Homo-grafted Anahu and homografted Rutgers also produced similar stem/leaf dry weights. However, stem leaf dry weights for
both homo-grafted Rutgers and Anahu were significantly less than non-grafted Rutgers and
Anahu (Table 6). Hetero-grafted Rutgers and hetero-grafted Anahu stem/leaf dry weight was
similar. Stem/leaf dry weight was significantly less for hetero-grafted Rutgers than either nongrafted Rutgers or Anahu. On the other hand, hetero-grafted Anahu stem/leaf dry weight was
not significantly different than non-grafted Rutgers, but it was significantly less than non-grafted
Anahu (Table 6).
The root fresh weight was significantly different among the six treatments (F5,14=7.263,
p=0.001). Non-grafted Rutgers and non-grafted Anahu had a similar amount of root fresh weight
(Table 6). Both non-grafted treatments, however, had significantly greater amounts of root fresh
weight than all other grafted treatments. Homo-grafted and hetero-grafted treatments had a
similar amount of root fresh weight.
The root dry weight was significantly different among the six treatments (F5,15=4.014, p=0.003).
Non-grafted Rutgers produced similar amount of root dry weight compare to non-grafted Anahu.
Homo-grafted Anahu had the lowest root dry weight among the six treatments and it was not
significantly different than homo-grafted Rutgers. However, homo-grafted Anahu produced less
37

root dry weight then non-grafted Anahu, but similar to non-grafted Rutgers. On the other hand,
homo-grafted Rutgers was not significantly different than non-grafted treatments. Heterografted Rutgers and hetero-grafted Anahu had similar amount of root dry weight and both were
significantly less than non-grafted Anahu but similar to non-grafted Rutgers.
Impact of Grafting on Tomato Plant Biomass Partitioning at After 100 Days Under Field
Conditions
Non-grafted Rutgers fruit fresh weight was the lowest among all six treatments (Table 7). Nongrafted Anahu produced a similar amount of fruit fresh weight as non-grafted Rutgers. Homografted Anahu produced the greatest amount of fruit fresh weight among all six treatments.
Homo-grafted Rutgers produced a similar amount of fresh fruit weight as home-grafted Anahu.
Hetero-grafted Rutgers produced the second greatest amount of fruit fresh weight. Hetero-grafted
Anahu produced much lower fruit fresh weight than hetero-grafted Rutgers. However, there was
no significant difference between hetero-grafted Rutgers and hetero-grafted Anahu.
The root fresh weight was not significantly different among all six treatments. Non-grafted
Rutgers produced the lowest amount of root fresh weight. Non-grafted Anahu root fresh weight
was not significantly different than non-grafted Rutgers. Homo-grafted Anahu produced the
greatest amount of root fresh weight. Homo-grafted Rutgers produced a similar amount of root
fresh weight to homo-grafted Anahu and it was the second greatest among the six treatments.
Hetero-grafted Rutgers root fresh weight was similar to hetero-grafted Anahu root fresh weight
(Table 7).
The stem/leaf dry weight was not significantly different among the six treatments (Table 8).
Non-grafted Rutgers stem/leaf dry weight was similar to non-grafted Anahu’s stem/leaf dry
weight. There was no significant difference between non-grafted Rutgers and non-grafted
Anahu. Homo-grafted Anahu stem/leaf dry weight was the greatest among the six treatments

38

and homo-grafted Rutgers stem/leaf dry weight was the lowest. There was, however, no
significant difference in stem/leaf dry weight between the two homo-grafted treatments. Heterografted Rutgers’s stem/leaf dry weight was the second greatest among the six treatments.
Hetero-grafted Anahu stem/leaf dry weight was not significantly different than hetero-grafted
Rutgers’s stem/leaf dry weight.
The root dry weight was not significantly different among the six treatments. Non-grafted
Rutgers root dry weight was numerically higher than non-grafted Anahu’s root dry weight. Nongrafted Anahu’s root dry weight was lowest amount all six treatments. The root dry weights of
the two non-grafted treatments were not significantly different. Homo-grafted Anahu produced
the greatest amount of root dry weight among the six treatments fallowed by homo-grafted
Rutgers. There was, however, no significant difference in root dry weight between the two
homo-grafted treatments. Hetero-grafted Rutgers and hetero-grafted Anahu also produced a
similar amount of root dry weight (Table 8).
Discussion
The impact of grafting tomato cultivars on fruit tissue yield has been studied in detail under
various abiotic and biotic stress conditions (Turhan et al. 2011; Schwarz et al. 2010; MartinezRodriguez et al. 2008; Venema et al. 2008). Grafting of tomato cultivars Anahu and Rutgers,
however, have not been studied under field condition. In addition, most grafting studies have not
included both partial and full-season biomass partitioning data. In our research, the stem/leaf
dry, root fresh, and root dry weights of 50-day-old homo-grafted and hetero-grafted tomato
plants were significantly less than those of non-grated plants. This is most likely due to the
wound healing process that took time and slowed plant growth in the early days of vegetative
growth. With mature plants, however, there were no significant differences for stem/leaf dry,
root fresh, and root dry among the grafted or non-grafted treatments. This result agrees with
39

Khah (2006b) findings where grafted plants dry and fresh weights were not significantly
different than those of non-grafted plants. In other studies, grafted plants have better water and
nutrient uptake compared to non-grafted plants (Di Gioia et al. 2013; Lee 1994). In this study,
homo-grafted Anahu biomass was slightly larger than other treatments due to water and nutrient
uptake by the plants at harvest. Homo-grafted Anahu, however, produced the lowest among the
six treatments amounts of fresh and dry weight of stem/leaf and root as well as fruit yield under
greenhouse conditions (Chapter 1). Even though other studies have found significant increases
in tomato fruit yield on grafted plants compared to non-grafted plants (Rivard and Louws 2008),
this was not the case in our research. Moreover, there was no negative effect on grafted plants
compared to non-grafted plants. We suspect that a lack of difference in biomass of plants at
harvest was due to a lack of biotic or abiotic environmental stress factors. Other studies,
however, have demonstrated grafting impacts on the fresh and dry weight and as well as
increased the yield under conditions of stress (Kumar et al. 2015; N. Liu et al. 2009; Di Gioia et
al. 2013; Rivard and Louws 2008; Savvas et al. 2009).
Conclusion
Grafting has excellent potential for use in integrated pest management due to increased concern
about environment quality and human safety associated with the use of pesticides. In this study,
there was no significant difference in mature tomato plant dry weight and yield between grafted
and non-grafted treatments. This was likely due to the fact there were insufficient biotic or
abiotic stress factors associated with the site until the arrival of Late Blight. Thus, future work is
needed for investigation of the benefits of homo and hetero-grafting of tomato cvs Anahu and
Rutgers under adverse conditions. Albeit, there were no significant effects in this study, grafting
has potentially great benefits, including: increase tomato fruit yield, increase plant vigor,
overcomes soil borne diseases. For instance, root-knot nematode species are soil-borne
40

pathogens that cause major economic losses in tomato production. Anahu is a root-knot
nematode (Meloidogyne incognita) resistant variety (Hu et al. 2015; Williamson et al. 1994;
Reddy, Tikoo, and Anand 1987) that can be used as a rootstock grafted to susceptible tomato
cultivars to overcome root-knot nematode damage under both field and greenhouse growing
conditions. Anahu also is resistant to both potato aphids and sweet potato whitefly (Bemisia
tabaci) (Nombela, Williamson, and Muñiz 2003; Goggin, Williamson, and Ullman 2001). Thus,
future research is required to evaluate use of tomato cv Anahu as a rootstock to confer resistance
to the grafted plant.

41

APPENDICES

42

Appendix A. Statistical Tables
Table 6. Influence of Solanum lycopersicum L. grafting on stem/leaf dry, root fresh, and
root dry tissue biomass partitioning after 50 days under field conditions.
Stem/leaf dry weight (g)

Grafting
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

Root fresh weight (g)

Root dry weight (g)

Mean

SE

Mean

SE

Mean

SE

c1

1.70

14.90 b

1.04

3.14 b

0.09

10.30 a

1.56

11.46 a

0.87

2.69 a

0.07

9.94 a

0.82

11.22 a

0.75

2.74 a

0.07

12.12 ab

1.06

11.34 a

0.87

2.77 a

0.07

11.63 a

0.90

11.53 a

0.61

2.81 ab

0.06

17.25 bc

1.43

16.19 b

0.70

3.01 ab

0.14

19.33

Date analyzed with One Way ANOVA. Significantly different means at p-value ≤ 0.05
followed by Tukey’s test. Same letter in the column are not different at p-value › 0.05.
1

Table 7. Impact of grafting on biomass partitioning of Solanum lycopersicum cvs Rutgers
and Anahu after 100 days under field conditions.
Fruit tissue weight
Non-grafted

Root tissue weight

Mean (kg)

SE

Mean (g)

SE

Rutgers

4.4a1

0.60

38.6a

6.13

Anahu

4.8a

0.75

57.6a

6.54

Anahu/Anahu

5.7a

0.61

77.7a

7.34

Rutgers/Rutgers

5.2a

0.67

65.6a

11.576

Rutgers/Anahu

5.7a

0.33

58.3a

4.21

Anahu/Rutgers

4.5a

0.4.8

59.9a

11.33

Homo-grafted

Hetero-grafted

Date analyzed with One Way ANOVA. Significantly different means at p-value ≤ 0.05
followed by Tukey’s test. Same letter in the column are not different at p-value › 0.05.
1

43

Table 8. Influence of Solanum lycopersicum L. grafting on stem/leaf and root dry weight
biomass partitioning after 100 days under field conditions.
Grafting
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

Stem/leaf tissue weight
Mean (g)
SE

Root tissue weight
Mean (g)
SE

140.10a1

23.03

4.09a

0.64

211.32a

22.16

7.81a

0.67

162.89a

14.55

6.37a

0.60

145.11a

14.18

6.29a

1.38

136.89a

23.93

6.49a

1.15

143.72a

19.94

5.69a

0.74

Date analyzed with One Way ANOVA. Significantly different means at p-value ≤ 0.05
followed by Tukey’s test. Same letter in the column are not different at p-value › 0.05.
1

44

Appendix B. Figures
Figure 7. Aerial view of the 2015 tomato grafting field research site at the Department of
Entomology Research Farm at Collins Road, Michigan State University.

Figure 8. Tomato grafting research site at the Michigan State University, Department of
Entomology Collins Road Research Farm.

45

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46

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Kubota, C., Mcclure, M. A., Kokalis-Burelle, N., Bausher, M. G., & Rosskopf, E. N. (2008).
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49

CHAPTER 3: INFLUENCE OF GRAFTING SOLANUM LYCOPERSICUM L. CVS
ANAHU AND RUTGERS ON PLANT BIOMASS PARTITIONING IN THE PRESENCE
AND ABSENCE OF MELOIDOGYNE INCOGNITA (NEMATODA): WITH THE
SPECIFIC REFERENCE TO CENTRAL ASIA.
Abstract
Understanding plant biomass partitioning is important for selection of appropriate rootstocks and
scions for grafting vegetable crops. Biomass allocation has not been studied in regards to
grafting Solanum lycopersicum L. The primary objective of this research was to determine the
impact of self-grafting (homo-grafting) and rootstock grafting (hetero-grafting) on tomato plant
fresh weight partitioning of stem and leaf, fruit, and root tissue following three types of pruning
(none, light, and heavy), in the presence and absence of Meloidogyne incognita at initial
population densities of Pi=1000, Pi=5000 and Pi=0. Two additional objectives were to evaluate
grafting M. incognita resistant tomato cv Anahu to the susceptible tomato cv. Rutgers for control
of M. incognita under greenhouse conditions, and determine if the function of the M. incognita
resistance gene (Mi) is retained in grafted tomato rootstocks. In addition to biomass partitioning,
homo-grafting and hetero-grafting were evaluated in regards to their impact on the final female
population density (Pf) of M. incognita. Non-grafted and homo-grafted S. lycopersicum were
used as controls. In the presence of M. incognita (Pi=1000) and no pruning, non-grafted Anahu
and non-grafted Rutgers possessed the same biomass partitioning signatures as in the absence of
M. incognita. With extensive pruning and M. incognita at Pi=0, Pi=1000, Pi=5000, all grafted
treatments possessed biomass partitioning signatures similar to non-grafted Rutgers. Most of the
biomass was partitioned to fruit tissue, compared to stem and leaf or root tissue. Resistant
tomato cv Anahu had a significant effect on the Pf of M. incognita in root tissue. Over all,
Anahu was not completely resistant to M. incognita. In the non-pruned experiment, there was an
11-fold difference in Pf between Anahu and Rutgers. Homo-grafted Anahu followed the same

50

trend and produced 19-fold fewer M. incognita at Pf than Rutgers. Nevertheless, hetero-grafted
Rutgers produced only 1-fold less Pf compare to Anahu. In the case of heavy and light pruning,
Pf was high on both non-grafted Anahu and homo-grafted Anahu, 10-fold and 15-20-fold,
respectively. When the plants were not pruned in the presence of M. incognita (Pi=1000), fruit
yield was 31 % less, compared to plants maintained in the absence of M. incognita. In
conclusion, Anahu rootstock grafting may not be beneficial to commercial tomato growers due to
its biomass partitioning signature where Anahu produced more leaves and stem under
greenhouse conditions. Moreover, Anahu resistance to M. incognita is not complete according to
these research results. Thus, Anahu should not be used as a rootstock to manage M. incognita
under greenhouse and possibly field conditions.
Introduction
Tomato (Solanum lycopersicum L.) is the second most valuable fruit/vegetable crop on a global
basis. Uzbekistan produces 80% of the tomatoes grown in Central Asia (FAO.STAT, 2010).
More than three million tons are produced annually. This is about four-fold less than tomato
production in the USA, yet Uzbekistan exports most of the tomato fruit to neighboring countries
and Russia. Soil-borne diseases represent key limiting factors for tomato production.
Sometimes farmers lose nearly 100% of expected yield due to soil-borne pests/pathogens. One
of these is Meloidogyne spp. (Nematoda: Heteroderidae), a sedentary-endoparasite of numerous
key food crops worldwide and one of the most damaging nematode pests in agriculture (Dong et
al, 2013, Davis et al, 2014). Meloidogyne incognita (Kofoid and White1919) Chitwood 1949 is
a common pest under both field and greenhouse tomato production conditions. During the past
60 years, control of Meloidogyne spp. has been based mainly on the use of soil fumigants, nonfumigant nematicides and rotation with non-host crops.

51

Due to environmental and human health risks, Integrated Pest Management (IPM) is
recommended for M. incognita control. IPM is designed to keep pest damage under threshold
levels using good monitoring plus all available control tactics, including, biocontrol agents,
cultural controls, host plant resistance, and when it is needed or required, chemical pesticides
(Bird, 2003). Not all IPM strategies work for M. incognita management under greenhouse
conditions. Use of pest resistant tomato varieties and hybrids can be an important aspect of IPM.
Some tomato varieties are resistant to M. incognita. These varieties carry the Mi gene. Mi
stands for resistance to M. incognita. Not all resistant tomato varieties are suitable for tomato
production, or preferred by farmers and consumers. Grafting has the potential to bring all the
essential traits together.
Vegetable grafting began early in the 20th century in Japan and Korea (J.-M. Lee & Oda, 2002)
and is now practiced on a global basis (J. Lee, 1994). Vegetable grafting is used to reduce risk to
detrimental abiotic and biotic factors (Pradeep et al., 2015; Charles et al., 2012; Theodore
McAvoy et al, 2012; Na Liu et al., 2009; Alfonso et al., 2008). Grafting is a key practice for
control of soil-borne diseases in East and South Asia (J.-M. Lee & Oda, 2002). In some cases,
both grafting and use of a resistant plant will enhance the crop’s overall vigor and protect it from
pests (Khah et al, 2006b). In several countries, vegetables are grafted to enhance overall plant
vigor (Estañ, 2005, Venema 2008). Grafting, however, can increase plant vigor, yet at the same
time, reduce yield potential.
It is important to understand biomass allocation in grafted plants in order to select optimal
rootstock and scion combinations. In tomato biomass resource allocation, it is well known that
increasing one type of tissue impacts other parts of the plant (Brian and Karl, 2002; Heuvelink,
1999). Plant resource partitioning has been extensively researched on non-grafted tomato plants

52

under greenhouse conditions (Albacete et al., 2008; Heuvelink, 1999). The impact of grafting on
resource partitioning is not well understood under severe or sub-optimal biotic stress conditions
(Marcelis, 1991). Tomato biomass partitioning has been studied extensively in regards to fruit
yield (Marcelis et al., 1998; Heuvelink, 1996), but much less is known about overall biomass
partitioning (Dimitrios et al., 2009, Charles and Xin, 2012). In particular, tomato has not been
studied in detail in regards to biomass partitioning after grafting and when grafted plants are
under stress, especially under greenhouse conditions. In addition, plant biomass partitioning has
not been studied in relation to homo-grafted (self-grafted), and hetero-grafted (rootstock grafted)
plants.
The specific objective of this research was to determine the impact of homo and hetero-grafting
on tomato plant fresh weight partitioning to stem and leaf, fruit and root tissues in the presence
and absence of the biological stress factor, M. incognita. The second objective was to evaluate
M. incognita resistant tomato cv Anahu grafted with susceptible tomato cv. Rutgers for future
use in greenhouse and field condition; including identification of the extent the Mi gene
mechanism is retained in grafted tomato plants in the case of Anahu resistant and Rutgers
susceptible grafted plants.
Materials and Methods
The research was conducted under greenhouse conditions at Michigan State University to
evaluate the impact of grafting S. lycopersicum on biomass partitioning. The Materials and
Methods Section describes the procedures used to determine tomato plant biomass partitioning
and nematode resistance evaluation of Anahu and Rutgers plants before and after grafting. It
includes the experimental design, plant materials, grafting procedures, greenhouse conditions,
final M. incognita population density and data analysis.

53

Experimental Design
The research included three initial population densities of M. incognita (Pi=0, Pi=1000, and Pi=5000
eggs per experimental unit) and grafting. There were six grafting treatments: 1) non-grafted
Anahu, 2) non-grafted Rutgers, 3) homo-grafted Anahu, 4) homo-grafted Rutgers, 5) heterografted Anahu (comprised of Anahu scion, Rutgers rootstock) and 6) hetero-grafted Rutgers
(comprised of Rutgers scion, Anahu rootstock). Each treatment was replicated five times. The
plants were maintained in a randomized complete block design under greenhouse conditions for
125 to 135 days after being transplanted into 5-gallon plastic pots.
Plant materials
A single tomato cv Anahu (University of Hawaii Seed Program) or cv Rutgers (Sustainable Seed
Co.) was sown in each cell in 50 or 72 cell seedling trays containing plant growth media
(SUREMIX professional growing media, Michigan Grower Products Inc.) and held in a growth
chamber maintained at 26 C for 16 hours and 20 C for eight hours, day and night, respectively.
The seedlings were watered every other day. Anahu seeds were planted two days before Rutgers
seeds to ensure the same seedling diameter for grafting. This time-interval was based on
preliminary research results. Approximately 30-day-old tomato plants were transplanted into
five-gallon plastic pots filled with sterilized sandy soil and placed in a greenhouse maintained at
26-28 C. Plants were watered as necessary and the ripe fruit (90% or more of the skin red in
color) harvested weekly. At the end of each experiment, all remaining tomato fruit were
harvested. Combined stem and leaf fresh weights were recorded and the tissue placed in a
drying oven at 30-35 C the same day. The tomato plant intact root system was removed from the
soil by washing with pistol nozzles in tap water and fresh root tissue weight recorded. The root
tissue was placed in a drying oven at the same temperature used for drying the stem and leaf
tissue. After seven days, stem and leaf and root dry weights were recorded.
54

Pruning
Pruning treatments consisted of three pruning levels including: non-pruned, lightly pruned and
heavily pruned. Simple pruning methods were used to remove suckers from the nodes. Each
sucker was pinched off entirely by hand when the sucker was small or with garden clippers when
suckers were larger. In heavy pruning, all of the suckers were removed from the entire plant and
leaves were removed up to a 3-4-foot height. Under the lightly pruned conditions, all the suckers
and leaves were removed up to first fruit clusters. The reason for pruning tomato plants was to
accommodate potential limited space in the greenhouse. In the non-pruned experiment (Figure
9), there were 60 tomato plants, five plants per greenhouse bench (60 in widths x 66 in length).
In Experiment 3, the plants were lightly pruned and there were 90 tomato plants, 15 tomato
plants per bench. The lightly pruned experiment was conducted three times. Heavily pruned
experiment was conducted only once and the numbers of plants were identical to the lightly
pruned experiments.
Grafting
After 14 days in the growth chamber, the seedlings were of appropriate size for grafting.
Rootstocks and scions were cut below the cotyledon, at a 45-65 degree angle using a Miter-Cut
Grafting Knife (Johnny's Selected Seeds). Scions and rootstocks were clipped together with
silicon tubes used to hold the rootstock and scion together until the grafting wound healed.
Silicon clipper (Hydro-Gardens, Colorado Springs, CO) size varied from 1.5-2.5 mm depending
on seedling diameter. During the graft wound healing process, a humidifier (Air Innovations
Model # HUMID06 1.37-gallons Ultrasonic Digital Humidifier) was used to control relative
humidity (RH). The grafted plants were maintained in a growth chamber for seven days at 24-26
C. The RH was decreased gradually from 90% at three days, to 85% at two days, and then to
75% one day and 65% the final day. After seven days, the grafted seedlings were moved to the
55

laboratory for three days of acclimatization. The six treatments included: 1) Non-grafted Anahu,
2) Non-grafted Rutgers, 3) Homo-grafted Anahu, 4) Homo-grafted Rutgers, 5) Hetero-grafted
Anahu (Anahu scion and Rutgers rootstock) and 6) Hetero-grafted Rutgers (Rutgers scion and
Anahu rootstock).
Meloidogyne. incognita Population Density
Only the root systems of plants inoculated with M. incognita and expressing root galls, were
used for estimating final female population densities of M. incognita (Figure 11). From each
experimental unit, five grams of root tissue were used for estimation of the final female
population density. The root tissue for analysis was cut at random from different parts of each
root system. The root staining procedure of Daykin and Hussey (1985) was used. The staining
helps to see female nematode in root system under microscope. Five grams of root tissue were
wrapped with a fine mashed nylon cloth tagged with corresponding treatment label. In this
procedure, nylon cloth was used for the first time in root staining. This prevented loss of root
tissue during the staining procedure and allowed for the staining of 20-40 roots at a time (Figure
12). Staining several roots together shortens the time spent for staining and increases staining
efficacy. Moreover, staining several roots together increases the staining quality by accurately
timing the staining procedure. Fixed roots were placed in 600 ml beakers containing 1:1 ratio of
8.25% NaOCl (Clorox Regular Bleach 3.78qt) and tap water for four minutes, stirring several
times. The roots were rinsed with tap water and soaked for 15 minutes in tap water. They were
then drained and washed under tap water several times in order to remove the rest of the bleach
from roots. After washing, the roots were soaked in water for 15 minutes. Next, the roots were
placed into a 600-ml beaker and filled with water until all the fixed roots were submerged. Stock
acid-fuchsine-stain solution was added at a concentration of 1.0 ml for each 30-50 ml water
required to fill the beaker. The beaker was placed on a hot plate, brought to a boil for one-two
56

minutes and cooled at room temperature. The cooled roots were drained, rinsed with tap water
and dried with a paper towel. The fixed roots were then filled with acidified glycerin with a few
drops of 5N HCl and boiled on a hot plate for 30-60 seconds. After the roots cooled, the Pf of M.
incognia were determined under a binocular microscope.
Data Analysis
The data were analyzed using SPSS 24 Grad Primmum Pack. First, data distribution normality
was checked with a histogram plot for skewness of the distribution. If the data were positively
skewed, natural Log or Log10 was used to transform the data. If the data were negatively
skewed, the Square Root Method was applied. If data were not normalized, a non-parametric
Kruskal-Wallis Test was used as the substitute for One-Way ANOVA. If the One-Way ANOVA
showed a significant difference at the α=0.05, means were separated using the Tukey’s Test. If
the non-parametric Kruskal-Wallis Test was significant at the α=0.05, means were separated
using the Dunn’s Test.
Results
Grafting Impact on Fruit Fresh Tissue Weight
When left unpruned, grafting did not increase the fruit tissue biomass (p=0.128), compared to
non-grafted tomato cultivars under M. incognita densities of Pi=0 and Pi=1000. Non-grafted
Rutgers produced the greatest amount of fruit tissue in both Pi=0 and Pi=1000 (Table 9A). Nongrafted Anahu, homo-grafted Anahu and hetero-grafted Rutgers produced least fruit weight at
Pi=1000. Overall fruit tissue was significantly less in Pi=1000, compared to Pi=0 (t49=2.105 P=0.04).
With light pruning, fruit tissue biomass was not significantly different among any treatments at
the Pi (Table 9B). This also was the case for tomato fruit tissue compared among Pi of M.
incognita (F2,6= 3.018, P=0.055).

57

With heavy pruning, grafting did not alter tomato fruit tissue biomass in any treatments with Pi=0.
There are significant differences among the six treatments for Pi=1000 (F5,24=5.075, P=0.003)
when the plants were heavily pruned (Table 9C). Rutgers produced the greatest amount of fruit
tissue, while the lowest amount of fruit tissue was observed on homo-grafted Anahu. There also
were significant differences among the six treatments for Pi=5000 (F5,24=3.544, P=0.015) when the
plants were heavily pruned (Table 9C). The greatest amount of fruit tissue was produced by
homo-grafted Rutgers, while homo-grafted Anahu produced the least amount of fruit tissue.
There was no significant difference on overall fruit yield among any Pis.
Impact of Pruning on Plant Population Density Under Greenhouse Conditions
In the non-pruned experiment, there were only two initial population densities of M. incognita
(Pi=0 and Pi=1000), due to greenhouse space. Sixty tomato plants were accommodated. With light
or heavy pruning, we were able to place 180 tomato plants in the same amount of space required
for 60 tomato plants in the non-pruned experiment.
Lightly and heavily pruned experiments had three level of M. incognita initial population
densities (Pi=0, Pi=1000, and Pi=5000). With heavy pruning, 90 tomato plants were placed in half of
the space required for the non-pruned experiment. Moreover, there were significant differences
in total fruit weight among non-pruned and lightly or heavily pruned experiments (H2=7.041,
P=0.03). Lightly pruned plants had 100 kg of fruit tissue, which was significantly more than
plants in the non-pruned experiment (P=0.043). With heavy pruning, tomato plants yielded 120
kg of fruit, which was significantly more compared to non-pruned plants which produced 68 kg
(P=0.012). In the heavily pruned experiment, however, total fruit tissue per plant was
significantly less compared to non-pruned plants (P=0.038) with Pi=0 and Pi=1000 of M. incognita.

58

Overall, tomato fruit production was increased by heavy pruning due to accommodating three
times more plants per unit area in the greenhouse.
Grafting Impact on Stem/Leaf Fresh Weight
Tomato stem/leaf fresh tissue weight was significantly different among treatments of non-pruned
plants in the absence of the M. incognita (H5=19.13, P=0.02) (Table 10). The greatest amount of
stem/leaf fresh tissue weight was produced on hetero-grafted Anahu and it was significantly
different than hetero-grafted Rutgers without pruning and in the absence of M. incognita. In the
absence of pruning and M. incognita, stem/leaf fresh weight was not significantly different on
non-grafted Anahu compared to non-grafted Rutgers. Similarly, homo-grafted Anahu and homografted Rutgers produced a similar amount of stem/leaf fresh tissue weight without pruning and
in the absence of M. incognita. Tomato stem/leaf fresh tissue weight was not impacted by any
grafting treatments of lightly pruned or heavily pruned plants (Table 10).
Grafting Impact on Root Fresh Weight
The root fresh weight of the six treatments was significantly different on lightly pruned plants
with a nematode initial population density of 1000 (F5,24=2.692, P=0.046). The greatest amount
of root fresh tissue weight was observed on homo-grafted Rutgers and it was significantly
different than the root fresh tissue weight on homo-grafted Anahu (P=0.023). Hetero-grafted
Anahu root fresh tissue weight was not significantly different than hetero-grafted Rutgers. Nongrafted Anahu produced a similar amount of root fresh tissue to non-grafted Rutgers (Table
11B).
Meloidogyne incognita Population Density
In the absence of pruning, resistant tomato cv Anahu had a significant negative effect on M.
incognita female population density in root tissue. There was an 11-fold reduction (P=0.014) in
female density in non-grafted Anahu compared to non-grafted Rutgers (Table 12A). Homo59

grafted Anahu followed a similar trend and produced 19-fold fewer females than homo-grafted
Rutgers (P=0.014). Hetero-grafted Rutgers had only a 1-fold reduction in female density
compared to hetero-grafted Anahu.
Under light pruning and a M. incognita density of Pi=1000, resistant tomato cv Anahu Pf increased;
however, the change was not significantly different than susceptible tomato cv Rutgers (Table
12B). Non-grafted Anahu produced 8% more Pf than non-grafted Rutgers. Homo-grafted Anahu
produced 30% less Pf compared to homo-grafted Rutgers. Hetero-grafted Rutgers, however,
increased Pf by 6% compared to hetero-grafted Anahu.
Under a higher density of Pi=5000 and with light pruning, non-grafted Anahu produced 2-fold
more Pf compared to non-grafted Rutgers. Homo-grafted Anahu produced 15-fold more Pf
compared to homo-grafted Rutgers and it was the highest Pf among all six treatments. Heterografted Rutgers produced 30% more Pf compared to hetero-grafted Anahu (Table 12B).
The lightly pruned experiment was conducted three times. In the first experiment, there were
significantly more tomato fruit than in the second experiment (stem/leaf tissue: P=0.140, fruit
tissue: P=0.02, root tissue: P=0.176). In the third experiment, all measurements were
significantly less than in the first lightly pruned experiment (stem/leaf tissue: P=0.049, fruit
tissue: P=0.031, root tissue: P=0.003). The data for the second and third experiments were not
analyzed due to plant growth issues that made it difficult to accurately record the results and
abnormal tomato fruit yield production. In particular, foliar necrosis took place during the
growth period, resulting in a lot of outlier data (Data is not presented).
With heavy pruning and a M. incognita density of Pi=1000, resistant cv. Anahu did not have a
significant impact on the population density of females in root tissue, compared to susceptible cv
Rutgers under the same conditions (Table 12C). Non-grafted Anahu produced one-fold fewer

60

females compare to non-grafted Rutgers. Homo-grafted Anahu reduced Pf by 44 %. Nematode
density did not differ on hetero-grafted Rutgers compared to hetero-grafted Anahu, supporting
2101 and 2487 M. incognita females, respectively. Overall, with heavy pruning there was no
significant difference in Pf among the six treatments at Pi=1000. There was no statistical
significance among six grafted treatments after heavy pruning at Pi=5000 (Table 12C). The
greatest Pf was on non-grafted Rutgers. It was 3.5-fold more, compared to non-grafted Anahu.
Hetero-grafted Rutgers produced the lowest Pf, with close to eight-fold fewer M. incognita
females compared to non-grafted Rutgers (P=0.032).
Biomass Partitioning
Non-pruned Experiment
In the absence of M. incognita (Pi=0), non-grafted Anahu and non-grafted Rutgers possessed
unique biomass partitioning signatures (Figure 13A). Non-grafted Anahu plants allocated more
of their fresh weight to stem and leaf tissue, compared to non-grafted Rutgers (P=0.029). Nongrafted Rutgers, however, allocated more biomass to fruit tissue, compared to non-grafted Anahu
(p=0.038). Root tissue was not statistically significant among the non-grafted tomato cvs Anahu
and Rutgers (p=0.226). Homo-grafting Anahu did not change the biomass partitioning signature
compared to non-grafted Anahu. Homo-grafted Anahu, however, slightly shifted (9%) biomass
towards to fruit tissue, compared to non-grafted Anahu (Figure 13A, B). Biomass partitioning
for homo-grafted Rutgers, was not significantly different compared to non-grafted Rutgers,
although it shifted slightly (9%) towards to stem/leaf tissue. Hetero-grafted Anahu did not
change the biomass partitioning signature, compared to non-grafted Anahu (Figure 13A, C).
Hetero-grafted Rutgers biomass shifted towards stem and leaf tissue (12%), compared to nongrafted Rutgers.

61

In the presence of a moderate density of M. incognita (Pi=1000), non-grafted Anahu and nongrafted Rutgers possessed the same unique biomass partitioning signatures as in the absence of
M. incognita (Figure 13AFigure 14A). Non-grafted Anahu allocated more of its fresh weight to
stem and leaf tissue, compared to non-grafted Rutgers (P=0.012). Non-grafted Rutgers,
however, allocated more biomass to fruit tissue, compared to non-grafted Anahu (P=0.043).
Root tissue fresh weights were not statistically different among the non-grafted tomato cvs
Anahu and Rutgers (P=0.760). Homo-grafting Anahu did not change the biomass partitioning
signature of fruit, stem/leaf, and root tissue, compared to non-grafted Anahu (Figure 14A, B).
Homo-grafted Rutgers biomass partitioning was not significantly different from that of nongrafted Rutgers, shifting slightly (4 %) from fruit tissue to stem and leaf tissue in comparison to
non-grafted Rutgers. The biomass partitioning signature of hetero-grafted Anahu was the same
as that of non-grafted Anahu (Figure 14A, C). Non-grafted Anahu, however, shifted its biomass
signature to fruit tissue (12%) in comparison to non-grafted Anahu. Hetero-grafted Rutgers s
allocated it biomass differently than non-grafted Rutgers, with hetero-grafted Rutgers shifting
more of its fresh weight to stem and leaf tissue than fruit tissue (P=0.05). Fruit tissue was
significantly less compared to non-grafted Rutgers. In all of the experimental treatments, the
amount of fresh weight allocated to root tissue was less than that allocated to fruit or stem/leaf
tissues.
Lightly-pruned Experiment
Under light pruning and in the absence of M. incognita (Pi=0), non-grafted Anahu and nongrafted Rutgers possessed unique biomass partitioning signatures (Figure 15A). There were no
significant differences among the grafting treatments. Non-grafted Anahu allocated more fresh
weight to stem and leaf tissue, compared to non-grafted Rutgers. Root tissue was not
significantly different in the non-grafted tomato cvs Anahu and Rutgers. Non-grafted Rutgers,
62

however, allocated more biomass to fruit tissue, compared to non-grafted Anahu. Homo-grafting
Anahu did not change the biomass partitioning signature compare to non-grafted Anahu. Homografted Anahu biomass shifted more to stem and leaf tissue after grafting, compared to nongrafted Anahu (Figure 15A, B). Homo-grafted Rutgers biomass partitioning, however, was not
significantly different than non-grafted Rutgers, although there were slight shifts toward stem
and leaf tissue (9%) and root tissue (5%), compared to non-grafted Rutgers. Hetero-grafted
Anahu did not change the partitioning signature, compared to non-grafted Anahu (Figure 15A,
C). Hetero-grafted Rutgers biomass did not change the biomass partitioning signature, compared
to non-grafted Rutgers (Figure 15A, C).
Under light pruning and in the presence of a moderate density of M. incognita (Pi=1000), nongrafted Anahu and non-grafted Rutgers, again, possessed the same unique biomass partitioning
signatures (Figure 16A). Non-grafted Anahu allocated more of its fresh weight to stem and leaf
tissue, compared to non-grafted Rutgers. Non-grafted Rutgers, however, allocated more biomass
to fruit tissue, compared to non-grafted Anahu. Root tissue was not significantly different for the
non-grafted tomato cvs Anahu and Rutgers. Homo-grafted Anahu biomass partitioning signature
was the same as for non-grafted Anahu (Figure 16A, B). Homo-grafted Rutgers biomass
partitioning was also not significantly different from non-grafted Rutgers, but did shift towards
stem and leaf tissue (11%) and root tissue (9%), in comparison to non-grafted Rutgers. Heterografted Anahu did not alter the partitioning signature, compared to non-grafted Anahu (Figure
16A, C). Hetero-grafted Rutgers allocated biomass differently than non-grafted Rutgers.
Hetero-grafted Rutgers allocated most of its fresh weight to fruit tissue.
Under light pruning and in the presence of a high density of M. incognita (Pi=5000), non-grafted
Anahu and non-grafted Rutgers possessed similar unique biomass partitioning signatures as in

63

the absence of M. incognita (Figure 17A). Non-grafted Anahu allocated more of its fresh weight
to stem and leaf tissue, compared to non-grafted Rutgers. Non-grafted Rutgers, however,
allocated more biomass to fruit tissue compared to non-grafted Anahu. Root tissues were not
significantly different for the non-grafted tomato cvs Anahu and Rutgers (Figure 17A). Homografting Anahu did not change its biomass partitioning signature in comparison to non-grafted
Anahu (Figure 17A, B). Homo-grafted Rutgers did not change its biomass partitioning signature
compared to non-grafted Rutgers. Likewise, hetero-grafted Anahu did not change the
partitioning signature compared to non-grafted Anahu, but shifted slightly toward root tissue
(5%) (Figure 17A, C). Hetero-grafted Rutgers did not change the biomass partitioning compare
non-grafted Rutgers.
Heavily-pruned Experiment
After heavy pruning and in the absence of M. incognita (Pi=0), non-grafted Anahu and nongrafted Rutgers possessed similar biomass partitioning signatures (Figure 18A). All six
treatments allocated most of their biomass to fruit tissue, compared to stem and leaf tissue. Root
tissue had the least fresh weight in all six treatments. In the presence of M. incognita (Pi=1000),
non-grafted Anahu and non-grafted Rutgers possessed similar biomass partitioning signatures
(Figure 19 A). All six treatments allocated more of the biomass to fruit tissue than to stem and
leaf tissue. Root tissue had the least weight in all six treatments. In the presence of M. incognita
(Pi=5000), non-grafted Anahu and non-grafted Rutgers possessed similar biomass partitioning
signatures (Figure 20A). All treatments allocated more of the biomass to fruit tissue than to stem
and leaf tissue. Root tissue had the least weight in all treatments.
Discussion
Understanding how a plant partitions its biomass is critical to choosing root-stocks for
commercial tomato grafting (Khah, Kakava, Mavromatis, Chachalis, & Goulas, 2006a). Tomato
64

cultivars differ in how they allocate biomass and often very little is known about the effect of
grafting on biomass. Rootstocks can allocate plant biomass differently and allocation of biomass
on the scion can be manipulated by rootstocks (Kudo & Harada, 2007; Yang et al., 2015). Plant
biomass allocation has been studied in the past by a number of researchers (Marcelis, 1991;
Heuvelink, 1997; Marcelis, Heuvelink, & Goudriaan, 1998; Heuvelink, 1999; Enquist & Niklas,
2002). However, none of studies investigated biomass partitioning of grafted vegetable plants.
This is the first paper describing biomass partitioning of grafted tomato plants with comparison
to non-grafted plants. Moreover, this research includes biomass allocation with three types of
pruning in the presence and absence of the southern root-knot nematode, M. incognita.
In the absence of M. incognita and pruning, non-grafted Anahu and non-grafted Rutgers,
possessed unique biomass partitioning signatures. Non-grafted Anahu allocated more of its fresh
weight to stem and leaf tissue; whereas, non-grafted Rutgers allocated more of its fresh weight to
fruit tissue. Homo-grafted Rutgers, shifted its biomass from fruit to stem and leaf tissue. This is
consistent with other studies demonstrating that grafting makes plants more vigorous (Estañ,
Martinez-Rodriguez, Perez-Alfocea, Flowers, & Bolarin, 2005; Venema, Dijk, Bax, van Hasselt,
& Elzenga, 2008), but does not increase fruit tissue (Romano & Paratore, 2000). In contrast,
hetero-grafted Anahu and hetero-grafted Rutgers biomass signatures were the same as the nongrafted Anahu biomass allocation signature. One possible explanation for the increased vigor is
that the Anahu scion is not influenced by the Rutgers biomass partitioning signature. In contrast,
the Rutgers biomass signature is manipulated by the Anahu biomass partitioning signature in
hetero-grafted Rutgers. A second possibility is that wounding during grafting triggers plant
defensive mechanisms. As a result, the grafted tomato cultivar scion becomes more vigorous
without producing more fruit. Consistently, the literature indicates that plants become more

65

vigorous after grafting (Khah et al., 2006b). Contradictory to this, we found that non-grafted
Anahu allocated more biomass towards to stem and leaf tissue than fruit tissue. Thus, the Anahu
biomass allocation signature appears to be unique, along with its ability to control scion biomass
partitioning. Authors who reported plant vigor or fruit increase after grafting have neglected to
compare and contrast the biomass partitioning with the non-grated cultivar’s biomass
partitioning signature.
In the presence of a moderate density of M. incognita (Pi=1000) both non-grafted Anahu and nongrafted Rutgers retained their biomass partitioning signatures. Nematode infection resulted in
less fruit tissue (Table 9), rather less than stem and leaf tissue (Table 10). Thus, biomass
partitioning shifted towards stem/leaf tissue (Figure 13AFigure 14A) and there was a significant
decrease (P=0.027) in yield between Pi=0 and Pi=1000. Root galling increased root tissue weight
slightly in some treatments with Pi=1000 (Table 11A-C). In the heavy pruned experiment with all
Pi, biomasses partitioning shifted from stem and leaf tissue to fruit tissue due to the heavy
pruning. Despite the nematode infection resulting in a low amount of fruit tissue, fruit tissue
biomass remained high compared to other parts of the plant biomass.
In this research, the Anahu Mi gene for resistance to M. incognita (Williamson, Ho, Wu, Miller,
& Kaloshian, 1994) did not provide complete resistance to this nematode. While Anahu
produced females in the root systems, the Pf was reduced up to 19-fold, compared to Rutgers
when plants were not pruned. Reddy, Tikoo and Anand (1987) found Anahu to be susceptible to
M. incognita at a Pi=20 per 100 grams of soil, supporting my findings. Another possibility for the
susceptibility of Anahu is that a resistance breaking M. incognita population was present in our
culture (Eddaoudi, Ammati, & Rammah, 1997). An additional possible explanation for Anahu
supporting M. incognita is that the Mi gene losses resistance above 28 C to M. incognita

66

(Dropkin, 1969; Cap, Roberts, & Thomason, 1993; Veremis, Van Heusden, & Roberts, 1999; Hu
et al., 2015). Greenhouse temperature in our study, however, was kept at 26 C during the winter
when tomato plants were inoculated with M. incognita eggs. Thus, Mi gene segregation due to
high temperature was unlikely. With heavy and light pruning, M. incognita Pf increased
compared to that in the non-pruned experiments, up to 10-fold and 20-fold on non-grafted Anahu
and homo-grafted Anahu, respectively. This is evidence that pruning can have a negative impact
on resistance.
Root galling was different in Pi=1000, compared to Pi=5000. Roots were more heavily galled with
Pi=1000 than Pi=5000 (Table 12B, C). Moreover, the root system was healthier and produced larger
galls on the root systems of susceptible tomato cultivars with Pi=1000. Also, inoculating tomato
seedling with large numbers of eggs resulted in secondary disease infection with symptoms of
root necrosis. High Pf in Anahu was observed in the both heavily pruned and lightly pruned
experiments. There were no significant differences on Pf between resistant and susceptible
cultivars following inoculation with M. incognita egg Pi=1000 or Pi=5000, per experimental unit.
Increase in Pf of M. incognita after pruning has not been reported in previous research.
Heavily and lightly pruned tomato plants fit into a smaller space in the greenhouse than nonpruned plants, resulting in a higher yield of fruit per unit area. Heavily pruned tomato plants (90
experimental units) and lightly pruned plants (90 experimental units) were placed in the same
area that accommodated only 60 non-pruned plants. The total yield was 68kg of fruit for the 60
non-pruned plants, while a combined tomato yield 220 kg was recorded for the 180 heavily
pruned and lightly pruned plants. The findings suggest that pruning can be used as a means of
increasing production of greenhouse-grown tomatoes. Pruning plants, however, decreased
tomato fruit tissue per plant due to removing most of the leaves, the energy and matter syntheses

67

factory of the plants (Ferrandino, 1999). This finding was similar to that of Olson (1989) and
Kanyomeka and Shivute (2005), where heavily pruned tomato fruit production was less than
non-pruned and removal of leaves was negatively correlated with fruit yield.
The results of this study indicate that it is important to understand resource partitioning before
selecting a rootstock for commercial tomato production. While resistant rootstocks can suppress
soil-borne disease, choosing a resistant rootstock might not be in the best interest of a grower if
the plant allocates biomass towards stem and leaf tissue, instead of fruit tissue. A grower must
have access to rootstocks that allocate biomass towards fruit tissue and not stem and leaf tissue.
This is consistent with the conclusions of Lopez-Perez et al (2006) where homo-grafted plants
decreased yield compare to non-grafted or hetero-grafted plants.
Grafting has been reported to result in scion control over the rootstock in fruit trees, including:
apricot, peach, apple, pear etc. To eliminate this phenomenon Lopez-Perez et al (2006)
compared susceptible scion grafted to resistant rootstock with the same resistant non-grafted
tomato cultivar where the number of galls and final egg population density of M. incognita was
not statistically significant. In contrast, we found in the non-pruned experiment that non-grafted
Anahu had a five-fold greater Pf when resistant tomato cv. Anahu was used as the rootstock. In
this case, the resistant rootstock was not manipulated by the susceptible scion.
Conclusion
Grafting is known to improve plant salt tolerance, yield, and plant vigor. These have been very
well studied. Increased biomass in one part of the plant, however, can lead to decreases in other
parts of the plant. Different tomato cultivars allocate biomass differently. Thus, biomass
partitioning in tomato is cultivar specific. Based on the biomass partitioning signature results I
conclude that Anahu rootstock can controlled biomass partitioning signature of Rutgers when the
Rutgers was used as a scion (Figure 13C). My findings provide an understanding of tomato
68

biomass partitioning that will be useful in the selection of rootstocks favorable for commercial
tomato fruit production under greenhouse conditions.
Anahu does not appear to be a good rootstock for grafting for several reasons. First, Anahu is
not completely resistant to M. incognita. Second, biomass partitioning in Anahu does not favor
tomato fruit production. Third, heavily infested Anahu roots are susceptible to secondary
diseases that will cause early mortality of tomato plants.
The results demonstrate that pruning tomato plants provides more space to grow additional
tomato plants under greenhouse conditions, resulting in greater fruit yields.

69

APPENDICES

70

Appendix A. Statistical Tables

Table 9. Influence of Solanum lycopersicum L. grafting on fruit tissue in the presence and
absence of Meloidogyne incognita.at initial nematode population densities of Pi=0, Pi=1000,
and Pi=5,000 eggs per experimental unit. A. Non-pruned, B. Lightly pruned, C. Heavily
pruned.
A
Grafting
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

B
Grafting
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

C
Grafting
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers
1

Non-pruned
Pi=0
Mean (g)
1441.5
1760.1
1216.7
1345.3
1678.1
2161.0

Pi=1000
SE
760.7
802.1
1282.9
1041.5
1207.6
1043.9

Mean (g)
623.2
675.5
540.6
1100.2
1565.2
1691.2

Difference (g)
SE
488.2
619.5
243.5
833.9
696.9
767.5

0 vs 1000
818.3
1084.6
676.1
245.0
113.0
469.8

Lightly pruned
Pi=0
Mean (g)
1262.0
841.9
1686.4
1219.7
1076.3
1690.8

Pi=1000
SE
683.0
352.5
668.9
273.9
480.0
717.4

Mean (g)
988.4
1026.7
1141.5
969.3
879.9
1168.2

Pi=5000
SE
460.1
358.1
275.2
490.4
281.6
504.0

Mean (g)
719.5
1008.9
1330.4
722.1
1179.6
1058.4

Difference (g)
SE
415.9
446.4
972.3
449.3
471.7
345.1

0 vs 1000
273.7
-184.8
544.8
250.4
196.4
522.7

0 vs 5000
542.5
-167.0
356.0
497.6
-103.3
632.4

Heavily pruned
Pi=0
Mean (g)
1269.4
1180.2
1810.2
1222.3
1738.8
1433.9

Pi=1000
SE
496.8
607.4
332.3
538.3
709.7
479.8

Mean (g)
1117.2 a
947.4 a
1306.8 ab
932.2 a
1740.0 ab
2052.3 b

Pi=5000
SE
679.6
400.4
192.7
239.7
668.7
252.9

Mean (g)
1134.8 ab1
768.8 a
1297.9 ab
927.7 ab
1641.4 b
1410.3 ab

Difference (g)
SE
367.5
216.4
506.5
234.3
560.4
246.9

0 vs 1000
152.2
232.8
503.4
290.1
-1.2
-618.3

Column means followed by different letters are significantly (P = 0.05) different.

71

0 vs 5000
134.6
411.4
512.3
294.5
97.4
23.7

Table 10. Influence of Solanum lycopersicum L. grafting on stem/leaf tissue in the presence
and absence of Meloidogyne incognita at initial nematode population densities of Pi=0,
Pi=1000, and Pi= 5000 eggs per experimental unit. A. Non-pruned, B. Lightly pruned, C.
Heavily pruned.
A

Non-pruned
Pi=0

Pi=1000

Difference (g)

Grafting

Mean (g)

SE

Mean (g)

SE

Anahu

2382.4 ab

416.3

2299.8

255.8

82.6

Anahu/Anahu

1870.4 ab

484.7

2257.6

486.6

-387.2

Rutgers/Anahu

1661.6 a

346.1

1752.4

341.4

-90.8

Anahu/Rutgers

2515.2 b

633.5

2082.8

600.6

432.4

Rutgers/Rutgers

2006.8 ab

435.6

1885.2

170.4

121.6

Rutgers

1717.0 ab

49.1

1696.0

245.0

21.0

B
Grafting

0 vs 1000

Lightly pruned
Pi=0

Pi=1000

Pi=5000

Difference (g)

Mean (g)

SE

Mean (g)

SE

Mean (g)

SE

0 vs 1000

0 vs 5000

Anahu

1707.8

420.4

1128.2

452.2

865.8

532.3

579.6

842.1

Anahu/Anahu

1718.0

538.1

1230.8

670.4

1093.0

540.3

487.2

625.0

Rutgers/Anahu

1652.2

473.9

1067.4

651.9

849.2

479.7

584.8

803.0

Anahu/Rutgers

1653.2

234.1

1164.6

181.6

802.8

392.4

488.6

850.4

Rutgers/Rutgers

1692.0

120.2

1019.0

571.6

585.4

105.3

673.0

1106.6

Rutgers

1569.6

240.2

694.8

153.9

603.2

227.9

874.8

966.4

C
Grafting

Heavily pruned
Pi=0

Pi=1000

Pi=5000

Difference (g)

Mean (g)

SE

Mean (g)

SE

Mean (g)

SE

0 vs 1000

0 vs 5000

Anahu

1055.8

288.7

738.6

217.1

822.2

207.9

317.2

233.6

Anahu/Anahu

871.2

169.0

971.4

350.1

903.0

387.4

-100.2

-31.8

Rutgers/Anahu

905.6

196.2

1077.8

154.8

689.4

261.3

-172.2

216.2

Anahu/Rutgers

859.4

162.2

775.4

341.6

681.4

154.5

84.0

178.0

Rutgers/Rutgers

955.2

250.1

737.8

118.7

646.8

104.0

217.4

308.4

Rutgers

893.6

123.6

749.6

261.8

834.0

167.6

144.0

59.6

72

Table 11. Influence of Solanum lycopersicum L. grafting on root tissue in the presence and
absence of Meloidogyne incognita at initial nematode population densities of Pi=0, Pi=1000,
and Pi= 5000 eggs per experimental unit. A. Non-pruned, B. Lightly pruned, C. Heavily
pruned.
Non-pruned

A

Pi=0
Grafting

Pi=1000

Difference (g)

Mean (g)

SE

Mean (g)

SE

0 vs 1000

Anahu

217.6

70.7

118.8

33.5

98.8

Anahu/Anahu

142.0

69.6

141.6

80.5

0.4

Rutgers/Anahu

120.0

39.5

179.2

100.8

-59.2

Anahu/Rutgers

195.2

41.6

248.8

124.6

-53.6

Rutgers/Rutgers

158.4

31.6

205.2

69.2

-46.8

Rutgers

159.6

33.1

224.0

119.4

-64.4
Lightly pruned

B

Pi=0
Grafting

Pi=1000

Pi=5000

Difference (g)

Mean (g)

SE

Mean (g)

SE

Mean (g)

SE

0 vs 1000

0 vs 5000

Anahu

156.8

113.7

181.4 a

75.2

165.1

92.9

-24.7

-8.4

Anahu/Anahu

128.2

71.0

207.8 a

90.7

289.0

105.9

-79.6

-160.8

Rutgers/Anahu

136.0

64.0

158.4 a

73.8

208.4

102.8

-22.4

-72.4

Anahu/Rutgers

214.0

128.7

215.4 a

74.3

241.2

132.5

-1.4

-27.2

Rutgers/Rutgers

285.8

201.1

380.6 b

206.9

168.6

60.7

-94.8

117.2

Rutgers

151.8

19.9

164.4 a

94.0

190.6

83.8

-12.6

-38.8

Heavily pruned

C

Pi=0
Grafting

Pi=1000

Pi=5000

Difference (g)

Mean (g)

SE

Mean (g)

SE

Mean (g)

SE

0 vs 1000

0 vs 5000

Anahu

90.8

63.6

105.6

52.8

139.8

67.6

-14.8

-49.0

Anahu/Anahu

88.0

14.7

111.4

60.4

153.2

76.8

-23.4

-65.2

Rutgers/Anahu

73.0

32.4

91.4

54.5

106.2

59.5

-18.4

-33.2

Anahu/Rutgers

76.4

25.0

107.8

67.5

170.8

48.7

-31.4

-94.4

Rutgers/Rutgers

93.6

51.0

175.6

96.1

166.6

48.5

-82.0

-73.0

Rutgers

82.8

25.5

158.8

46.5

210.6

68.7

-76.0

-127.8

73

Table 12. Influence of Solanum lycopersicum L. grafting on final female population
densities of Meloidogyne incognita associated with initial M. incognita egg population
densities of Pi=1,000 and Pi=5,000 per experimental unit. A. Non-pruned, B. Lightly pruned, C.
Heavily pruned.
Non-pruned

A

Pi=0

Grafting

Pi=1000

Difference

Mean (g)

SE

Mean (g)

SE

0 vs 1000

Anahu

0.0

0.0

297.1

226.9

-297.1

Anahu/Anahu

0.0

0.0

243.0

289.3

-243.0

Rutgers/Anahu

0.0

0.0

1547.4

1665.7

-1547.4

Anahu/Rutgers

0.0

0.0

3370.2

2951.0

-3370.2

Rutgers/Rutgers

0.0

0.0

4618.2

3758.3

-4618.2

Rutgers

0.0

0.0

3329.2

2677.5

-3329.2

Lightly pruned

B

Pi=0
Grafting

Pi=1000

Pi=5000

Difference

Mean (g)

SE

Mean (g)

SE

Mean (g)

SE

Pi=0 vs Pi=5000

Anahu

0.0

0.0

3042.8

3808.8

5749.4

5635.6

-2706.5

Anahu/Anahu

0.0

0.0

4778.2

4440.5

11590.8

17613.7

-6812.7

Rutgers/Anahu

0.0

0.0

3988.8

2768.7

7754.3

8264.6

-3765.5

Anahu/Rutgers

0.0

0.0

3733.5

4558.6

5469.4

5514.1

-1735.8

Rutgers/Rutgers

0.0

0.0

6732.3

5762.3

778.8

712.6

5953.6

Rutgers

0.0

0.0

2800.8

2948.4

2895.3

4460.0

-94.5

Heavily pruned

C

Pi=0
Grafting

Pi=1000

Pi=5000

Difference

Mean (g)

SE

Mean (g)

SE

Mean (g)

SE

Pi=0 vs Pi=5000

Anahu

0.0

0.0

2837.3

3141.4

3769.5

4840.1

-932.2

Anahu/Anahu

0.0

0.0

3748.2

3681.8

6420.3

5692.3

-2672.1

Rutgers/Anahu

0.0

0.0

2100.7

2627.0

1667.4

1163.8

433.3

Anahu/Rutgers

0.0

0.0

2486.8

3238.6

7804.6

6257.2

-5317.8

Rutgers/Rutgers

0.0

0.0

6642.8

3896.5

4495.4

2814.6

2147.4

Rutgers

0.0

0.0

5884.8

1095.1

12886.0

10066.0

-7001.2

74

Appendix B. Figures

Figure 9. Grafted and non-pruned Solanum lycopersicum L. experiment under greenhouse
conditions.

Figure 10. Grafted Solanum lycopersicum L. heavily pruned and lightly pruned greenhouse
trials. Left: heavily pruning experiment, Right: lightly pruned experiment. There were 15
experiment units per bench.

75

Figure 11. Solanum lycopersicum L. root systems: Left. Susceptible tomato cv Rutgers
under initial egg population density (Pi) of 1000 per experimental unit. Right. Susceptible
tomato cv Rutgers under initial egg population densities (Pi) of 0 per experimental unit.

76

Figure 12. Root staining method with nylon fine mashed cloth to prevent loss of small
pieces of root tissue.

77

Figure 13. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers in the absence of M. incognita (Pi=0). A. Non-grafted, B. Homo-grafted, C. Heterografted

78

Figure 14. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers in the presence of M. incognita (Pi =1000). A. Non-grafted, B. Homo-grafted, C.
Hetero-grafted.

79

Figure 15. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers in the absence of M. incognita (Pi = 0) and with lightly pruning. A. Non-grafted, B.
Homo-grafted, C. Hetero-grafted.

80

Figure 16. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers in the presence of M. incognita (Pi =1000) and with lightly pruning. A. Non-grafted,
B. Homo-grafted, C. Hetero-grafted.

81

Figure 17. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers with M. incognita (Pi = 5000) and with lightly pruning. A. Non-grafted, B. Homografted, C. Hetero-grafted.

82

Figure 18. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers with heavily pruning and in the absence of M. incognita (Pi = 0). A. Non-grafted, B.
Homo-grafted, C. Hetero-grafted.

83

Figure 19. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers in the presence of M. incognita (Pi =1000) with heavily pruned. A. Non-grafted, B.
Homo-grafted, C. Hetero-grafted.

84

Figure 20. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers with heavily pruning and in the absence of M. incognita (Pi = 5000). A. Non-grafted,
B. Homo-grafted, C. Hetero-grafted.

85

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86

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benefits. HortScience, 29(4), 235–239.
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López-Pérez, J.-A., Le Strange, M., Kaloshian, I., & Ploeg, A. T. (2006). Differential response of
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CHAPTER 4: IMPACT OF SOLANUM LYCOPERSICUM L. GRAFTING ON
TRIALEURODES VAPORARIORUM WESTWOOD (INSECTA) DEVELOPMENT
UNDER GROWTH CHAMBER CONDITIONS: WITH SPECIAL REFERENCE TO
THE MI GENE, TYPE-D TRICHOMES AND CENTRAL ASIA.
Abstract
Solanum lycopersicum L. resistance to Trialeurodes vaporariorum Westwood has not been studied
in regards to the Mi gene which confers resistant to Meloidogyne incognita (Kofoid & White)
Chitwood, Macrosiphum euphorbiae (Thomas) and Bemisia tabaci (Gennadius).

Grafting

susceptible cultivar scions to resistant rootstocks with the Mi gene should control, M. incognita,
but it is not known if the resistance to foliar feeding insects will be conferred to the scion. In this
research Anahu with the Mi gene and Rutgers without the Mi gene were homo-grafted, heterografted and evaluated in regards to resistance to T. vaporariorum. Glandular trichomes are
responsible for mechanical and chemical resistance, thus Type-D Trichome densities differences
were evaluated in grafted plants. T. vaporariorum population density associated with non-grafted
Anahu was significantly less that that associated with homo-grafted Rutgers. Type-D Trichome
density was significantly different among three plant heights. The greatest Type D-Trichome
density was on leaves immediately basipetal to the apical meristem. Anahu had significantly more
Type-D Trichomes than Rutgers. In this research, the Anahu Mi gene carrying tomato cultivar
was not resistant to T. vaporariorum. Grafting did not impact Type-D Trichome density.
Introduction
Solanum lycopersicum L. is a major vegetable crop in Uzbekistan, where greenhouse whitefly
Trialeurodes vaporariorum Westwood, 1856 (Hemiptera: Aleyrodidae) is a devastating pest of
this plant. In addition to direct damage, T. vaporariorum vectors viruses of economic
significance (Jones 2003). For instance, T. vaporariorum transmits Tomato Infectious Chlorosis
Virus (Zalom 2012; Duffus, Liu, and Wisler 1996). In Uzbekistan, whitefly management is

90

based on insecticidal control in both greenhouse and field production systems. Farmers apply
insecticides weekly (personal conversation/observation) under greenhouse conditions. This can
be harmful to the environment and result in excessive chemical residues. Moreover, overuse of
pesticides increases the selection of populations of T. vaporariorum that are resistant to specific
active ingredients (Gorman et al. 2007; Gorman et al. 2002). Using resistant varieties is an
environmentally friendly management strategy for control of T. vaporariorum in tomato
production. For instance, the tomato cultivar Anahu has the Mi gene that confers resistant to
Meloidogyne incognita (Kofoid & White) Chitwood and also cross-resistance to Macrosiphum
euphorbiae (Thomas) (Goggin, Williamson, and Ullman 2001; Vos et al. 1998) and sweet potato
whitefly, Bemisia tabaci (Gennadius) (Nombela, Williamson, and Muñiz 2003; Goggin,
Williamson, and Ullman 2001). There are several Mi genes in cultivated and wild tomatoes.
These include Mi-1 to Mi-9. Mi-1 has seven homologues, Mi-1.1 through Mi-1.7 (Seah, Telleen,
and Williamson 2007). Among these, the Mi-1.2 homologue is the only functional nematode
resistant gene for M. incognita (Milligan et al. 1998). There is no literature indicating that the
Anahu Mi gene confers resistant to T. vaporariorum. Plants infested with T. vaporariorum,
however, trigger the same resistance reaction as that of B. tabaci infection (Puthoff et al. 2010).
Tomato leaves possess trichomes which can provide both mechanical and chemical defenses
against insects. Seven types of trichomes exist in tomato plants (Tissier 2012). They are divided
in to two groups based on glandular cell presence at the tip of the trichome. Glandular trichomes
are responsible for plant resistant and repellency to pests in tomato (Goffreda, Mutschler, and
Tingey 1988). Among glandular trichomes, Type-D, otherwise known as Type VI, is
responsible for mechanical and chemical resistance (Aina, Rodriguez, and Knavel 1972).

91

Tomato cultivars preferred in Central Asia do not possess the Mi gene. Grafting local cultivar
scions to tomato rootstocks with the Mi gene should control, M. incognita, but whether foliar
feeding insect resistance can also be conferred to the scion has not been determined. The
objective of this research was to determine if the Mi insect resistance trait of cv. Anahu can be
transferred to grafted cv. Rutgers in the absence of M. incognita. The second objective of this
research was to evaluate the relationship of tomato trichomes Type-D to T. vaporariorum
resistance after grafting.
Materials and Methods
The research was conducted under growth chamber conditions at Michigan State University to
evaluate the impact of grafted S. lycopersicum resistant rootstock on the population development
of T. vaporariorum. The Materials and Methods Section describes the procedures, including the
experimental design, plant materials, grafting procedures, growth chamber conditions, T.
vaporariorum population density bioassay, trichome Type-D density, and data analysis.
Experimental Design
There were six treatments: 1) non-grafted Anahu, 2) non-grafted Rutgers, 3) homo-grafted
Anahu, 4) homo-grafted Rutgers, 5) hetero-grafted Anahu (Anahu scion and Rutgers rootstock)
and 6) hetero-grafted Rutgers (Rutgers scion and Anahu rootstock). Each treatment was
replicated eight times. The plants were maintained in a completely randomized design under
growth chamber conditions for 20 to 25 days after being transplanted into 32 oz styrofoam cups.
Plant materials
A single tomato cv Anahu (University of Hawaii Seed Program) or cv Rutgers (Sustainable Seed
Co.) was sown in each cell in 50 cell seedling trays containing plant growth media (SUREMIX
professional growing media, Michigan Grower Products Inc.) and maintained under growth
chamber conditions. Growth chamber temperature was maintained at 26 C for 16 hours and 20
92

C for eight hours, day and night, respectively. The seedlings were watered every two days.
Anahu seeds were planted two days before Rutgers seeds to ensure the same seedling diameter
for grafting. This time-interval was based on preliminary research results.
Grafting
After 14 days under growth-chamber conditions, the seedlings were of appropriate size for
grafting. Rootstocks and scions were cut below the cotyledon, at a 45-65 degree angle using a
Miter-Cut Grafting Knife (Johnny's Selected Seeds). Scions and rootstocks were clipped
together with silicon tubes used to hold the rootstock and scion together until the grafting wound
healed. Silicon clipper (Hydro-Gardens, Colorado Springs, CO) size varied from 1.5-2.5 mm
depending on seedling diameter. During graft-wound healing, a humidifier (Air Innovations
Model # HUMID06 1.37-gallon Ultrasonic Digital Humidifier) was used to control relative
humidity. The grafted plants were maintained in a growth chamber for seven days at 24-26 C at
90% RH. The relative humidity was decreased gradually from 90% at three days, to 85% the
next two days, 75% the next day and 65% on the final day. After 7 days, the grafted seedlings
were moved to the laboratory for three days for acclimatization.
Growth Chamber
Following acclimatization, the tomato plants were transplanted into 32 oz styrofoam cups, filled
with steam sterilized sandy soil. Growth chamber temperature was maintained at 26 C for 16
hours and 20 C for eight hours, day and night, respectively. Plants were watered every two days
and as necessary.
T. vaporariorum Population Density Bioassay
A colony of T. vaporariorum was maintained on Rutgers, a susceptible tomato cultivar, under
growth chamber conditions. For each experimental unit, five female and at least one male T.
vaporariorum were introduced into mini-insectaries in two-dram vials paced under each tomato
93

plant. The insectaries were made from clear plastic jars with an opening of 3.5 inches designed to
fit snugly into the 32 oz cup containing the tomato plant (Figure 21). The insectaries had openings
on each side and on the top, covered with fine mashed nylon cloth to prevent white fly escape and
allowing for air exchange. The insectaries and tomato plants were removed from the growth
chamber after 20-25 days.

The abaxial side of leaves of each plant was observed for T.

vaporariorum eggs, nymphs, pupa, and pupa cases. Pupa and pupa cases were combined and
presented as pre-adults. They were counted using a binocular microscope at 60X magnification
and recorded.
Type-D trichome density
Glandular trichomes, known as Type-D or Type VI trichomes, were counted under a 60x
binocular microscope at three levels of tomato plant height: Type-D trichome density was
determined on the abaxial side of the leaves using one leaf from the second leaf branch (lower
height), one from top branch (upper height), and one from a mid-branch (mid height) of each
plant. The detached leaf adaxial surface was fixed to a glass slide with tape to make the leaf
surface flat, making it easy to counting Type-D trichomes under a 60 X binocular (Figure 22).
Data Analysis
The data were analyzed using SPSS 24 Grad Primmum Pack. First, the data distribution normality
was checked with a histogram plot for skewness. If the data were positively skewed, a natural Log
or Log10 transformation was used to normalize the data. If the data were negatively skewed, the
Square Root Method was applied.

If data were not normalized after transformation, non-

parametric Kruskal-Wallis Test was used as the substitute for ANOVA. In the case of the T.
vaporariorum population density bioassay, two identical bioassay experiments were conducted
separately under growth chamber conditions. Growth chamber conditions were as described
above. The results were compared using an Independent Sample T Test to identify significance
94

differences in total population density. Since there were no statistically significant differences in
the results of Experiment 1 and 2, the data were combined to increase the number of replicates. A
natural square root transformation was used to normalize the data. Since Leaven’s Test of
Homogeneity was violated for ANOVA, the non-parametric Kruskal-Wallis Test was used to
analyze the data. In the trichome density experiment, the data were positively skewed. A natural
log transformation was used to normalize the data. For all data, a one-way ANOVA was used to
test significant treatment effects. This was followed with Fisher’s Least Significance Difference
(LSD) test for mean separation.
Results
Nymphs
Nymph population density was significantly different among the six treatments (F5,90=2.788,
P=0.022). It was significantly less on non-grafted Anahu, compared to non-grafted Rutgers
(P=0.047) (Table 13). Homo-grafted Anahu had significantly fewer nymphs compared to homografted Rutgers (P=0.007). Hetero-grafted Anahu nymph density was numerically lower than
hetero-grafted Rutgers, but it was not significantly different (Table 13). The highest density of
nymphs developed on homo-grafted Rutgers, followed by non-grafted Rutgers.

Both had

significantly higher densities than all other grafted treatments.
Pre-adults and Hatched Pupa Cases
Pre-adult density was not statistically significant among the six treatments (F5,90=1.713, P=0.140).
The greatest density of pre-adult life stages were recorded on homo-grafted Rutgers, 70.25 (Table
13). The lowest pre-adult life stage population density was present on non-grafted Anahu, 43.25.
Total Population Density of T. vaporariorum
Total greenhouse population density was significantly different among the six treatments
(F5,90=2.141, P=0.068). The lowest total population density of T. vaporariorum was observed on
95

non-grafted Anahu and it was not significantly different from non-grafted Rutgers (Table 13).
The highest total population density of T. vaporariorum was observed on homo-grafted Rutgers
and it was significantly different than homo-grafted Anahu (P=0.024). Total population density
of T. vaporariorum on hetero-grafted Anahu and hetero-grafted Rutgers was not significantly
different, 61.50 and 76.69, respectively (Table 13)

Type-D Trichome Density after 7 and 25 days of Transplant into Plastic Pots
Leaves from the upper part of the plants had significantly more Type-D trichomes than those at
the mid or lower plant heights after seven days (Table 14). There was a significant effect of leaf
position on trichome density after seven days (F2,105=5.870, P=0.004). Type-D trichome numbers
were significantly higher on upper leaves than on lower leaves (P=0.007) or mid-height leaves
after seven days (P=0.002) (Table 14). When the data were analyzed between treatments there
was not significant difference after seven days (H5=9.019, P=0.108). Non-grafted Rutgers had
the least number of Type-D trichomes among all treatments and it was almost two-fold less than
Anahu’s Type-D trichome density, 31.89 and 55.50, respectively (Table 15) seven days after
transplanting. The highest density of Type-D trichomes was recorded on hetero-grafted Rutgers
and it was two-fold greater than homo-grafted Anahu seven days after transplanting. Type-D
trichome density was similar between homo-grafted Anahu and homo-grafted Rutgers, 49.94 and
39.17, respectively after seven days of transplant (Table 16).

Type-D trichome density was significantly different at the three heights after 25 day of
transplanting (F2, 105=9.527, P=0.001), with trichome density increasing with increasing height.
Leaves at the low height had the lowest Type-D trichome density and it was significantly
different than densities at the mid or upper heights, P=0.02 and P=0.001 respectively. Mid
96

region Type-D trichome density was significantly less then Type-D trichome density on upper
region leaves (P=0.046).
Twenty five days after transplanting, Type-D trichome density did not differ significantly among
the six treatments (F5,102=1.652, P=0.153). Numerically, the lowest Type-D trichome density
was observed on non-grafted Rutgers and it was similar to Type-D trichome density on nongrafted Anahu, 31.11 and 43.72, respectively (Table 17). Type-D trichome density on homografted Anahu and Rutgers was intermediate, 58.44 and 37.72, respectively. The greatest density
of Type-D trichomes was recorded on hetero-grafted Rutgers, 70.67; however, Type-D trichome
density was not significantly different than hetero-grafted Anahu, 49.39.
Discussion
Solanum lycopersicum has been studied extensively for insect pest resistance. Many resistant
wild tomato species and cultivated tomato varieties have been reported to have resistance to
tomato pests including: Tetranychus urticae, Macrosiphum euphorbia, Bemisia tabaci, and
Trialeurodes vaporariorum. Pest resistance in Solanum lycopersicum is based on a number of
factors including: trichomes, chemical exudes and overall genetics. In this research, we
evaluated two T. vaporariorum resistance characteristics, the Mi gene and abundance of Type-D
trichomes in regards to S. lycopersicum cultivars, Anahu and Rutgers. The results indicate that
that the Mi gene carrying S. lycopersicum cv Anahu was not resistant to T. vaporariorum.
Moreover, Type-D trichome density did not impact T. vaporariorum resistance.
Several Mi gene carrying tomato cultivars have significantly lowered Q-biotype of B. tabaci
population densities (Nombela, Beitia, and Muñiz 2001). In contrast, our findings document that
the Mi gene in cultivar Anahu did not confer resistant to T. vaporariorum and there was no
difference in population density of T. vaporariorum between Anahu and the susceptible cultivar
Rutgers. Lucatti et al (2010) found that two different Mi gene carrying tomato cultivars responded
97

differently to T. vaporariorum infestation. Tomato cultivar LC 138 was susceptible and tomato
cultivar Uco Plata resistant, producing significantly fewer adult T. vaporariorum, 43 and 10,
respectively. Kumar et al. (1995), however, found that the tomato cultivar Anahu, with the Mi
gene, was significantly less attractive to T. vaporariorum, compared to the tomato cultivar Rey de
los Tempranos, without the Mi gene. Moreover, Anahu showed moderate resistant to killing or
repelling 50% of inoculated Tetranychus urticae on a leaf disc experiment (Aina, Rodriguez, and
Knavel 1972).
The mini-insectary used in this research to evaluate T. vaporariorum population development
enabled me to test for resistance on intact tomato plants. This procedure was used because it is
difficult and not very accurate to use population density models to estimate whitefly population
densities (Rumei 1982). Mini-insectaries, however, can prevent direct light from reaching the
plant. As a result, plants in the mini-insectaries might have received lower levels of light, which
could impact photosynthesis and impacted plant growth and its overall physiology (CraftsBrandner and Chu 1999). Based on over observation we do not believe the light intensity had an
impact on over research.
In wild tomato Solanum pennellii and Solanum peruvianum resistance to Macrosiphum
euphorbiae is attributed to their sticky glandular trichromes and trichome density, respectively
(Gentile and Stoner 1968). Gentile et al (1968) later reported that Solanum pennellii and Solanum
hirsutum Humb. and Bonpl. had the same sticky glandular hairs that were death traps to T.
vaporariorum. However, these two reports did not specify which types of the trichomes are
responsible for insect pest resistance. Trichome types are discussed by Tissier (2012) after
Luckwill (1943), describing the presence of glandular trichomes in wild species and cultivated
tomatoes. According to this article, Type-D trichomes produce sticky exudes that are death traps

98

to several tomato pests. Rodriguez et al., found that Type-D trichomes were repellent and toxic to
mites. De Ponti and Hogenboom (1975) reported that resistance to T. vaporariorum in Solanum
hirsutum and Solanum hirsutum glabratum is based on internal factors in addition to glandular
sticky trichomes. This factor in Solanum hirsutum f. glabratum is reported as 2-tridiconon, a TypeD trichome exude which is toxic to various pests of wild tomato species (Williams et al. 1980).
Moreover, Williams et al. (1980) reported that wild tomato species have 72 times more of 2tridecanon chemical then cultivated tomatoes. Kevin and Frank (1995), however, screened 20
commercial cultivars of processing tomato and seven species of wild tomatoes. They found that
Bemisia tabaci B-biotype (Bemisia argentifolii) resistance was not due to trichome density. This
is consistent with the results of my findings where there was a weak relationship between trichome
density and T. vaporariorum population density (Figure 23).
The numbers of Type-D trichomes increased in density with increasing plant height. I also visually
observed that T. vaporariorum laid eggs preferentially on the lower region leaves where tomato
trichomes Type-D are less prevalent. Hargreaves (1915) mentioned that T. vaporariorum lays
eggs on younger leaves and trichome density interfered with oviposition rate and fecundity. This
characteristic, however, was not consistent with earlier observations on this research. The eggs
were distributed randomly among leaves in various positions when the density of ovipositing
adults was high. Gentile et al. (1968) failed to find significant difference of eggs and nymphs of
T. vaporariorum at four plant heights of wild tomato plants, using 16-mm-diam leaf discs. I
counted trichome Type-D density in each leaf and concluded that this should not be done where
leaf veins are present. Moreover, trichome Type-D density was not equal in each half of the single
leaf. Thus, the entire leaf Type-D trichomes were counted and samples were taken from three
heights of the plant.

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Conclusion
The primary objective for this research was to determine if the Mi gene in Anahu confers resistant
to T. vaporariorum. This was not supported by the results. This may have been due to the large
population density of T. vaporariorum on Anahu. Moreover, Anahu was developed through the
embryo rescue method by breading Solanum lycopersicum cv Michigan State Forcing and
Solanum peruvianum PI128657 (Liharska and Williamson 1997). Thus, a lack of resistance in
tomato cv Anahu can possibly be explained with S. peruvianum not being resistant to T.
vaporariorum (De Ponti, Pet, and Hogenboom 1975; Gentile, Webb, and Stoner 1968).
Additionally, plants exhibit high whitefly resistance at temperatures above 28 C under greenhouse
conditions. An additional possible explanation for Anahu not being resistant to T. vaporariorum
is that the Mi gene segregates above 28 C (Dropkin 1969; Cap, Roberts, and Thomason 1993;
Veremis, Van Heusden, and Roberts 1999; Hu et al. 2015). This is another clue that the Mi gene
is not responsible for T. vaporariorum resistance in tomato. Interestingly, Bas et al. (1992) found
a larger difference between genotypes in older plants. In this study, all bioassays were conducted
using 20-25 days old tomato plants. Bas et al. (1992) and Byrne and Draeger (1989) reported that
tomato plant resistance increases with plant age in some wild tomato species. Therefore, it would
be of interest to investigate the impact of plant age on the Mi gene carrying tomato cultivar’s
resistance to T. vaporariorum.

100

APPENDICES

101

Appendix A. Statistical Tables
Table 13. Influence of Solanum lycopersicum L grafting on T. vaporariorum life stage
population densities.

Grafting Treatment
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

Nymph

SE

Pre-adult

SE

Total

SE

1

12.50 a *

3.32

43.25

5.34

55.75 a

5.51

10.50 a

2.44

55.00

10.41

65.50 a

12.48

13.75 ab

2.01

62.94

7.61

76.69 ab

8.35

12.75 a

2.38

48.75

6.79

61.50 a

8.00

21.06 b

3.24

70.25

8.03

91.31 b

10.17

17.88 b

2.71

58.44

7.10

76.31 ab

7.41

1

Data were natural log transformed. T. vaporariorum population density was recorded at the end
of each experiment in entire plant. None of the data were violated Homogeny of variance after
natural log transformation. The data analyzed by One Way ANOVA and followed by Fisher’s
LSD for mean separation. Column means fallowed by the same letter are not significantly
different at α=0.10
Table 14. Solanum lycopersicum L Type-D trichome density at three plant heights seven
days after transplanting into plastic pot. Lower region (leaves from second branch), Midregion (leaves from a mid-region branch), and Upper region (leaves from last branch one
before merry stem).

Lower region
Mid region
Upper region

N

Type-D Trichomes

SE

36

42.72 a1

6.81

36

43.19 a

7.53

36

71.42 b

8.80

1

Data were natural log transformed. The data did not violated Homogeny of variance after
natural log transformation. The data analyzed by One Way ANOVA and followed by Fisher’s
LSD for mean separation.
* Different letter show significant difference at α=0.05

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Table 15. Impact of Solanum lycopersicum L grafting on Type-D Trichrome density seven
days after transplanting into plastic pot. Means for six replicates of each grafting
treatments. Three leaf samples were taken from three leaves of the each tomato plant.

Grafting

N

Mean

SE

Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

18

55.501

12.71

18

49.94

11.29

18

85.33

17.59

18

52.83

5.75

18

39.17

7.61

18

31.89

4.17

1

Data violated the Homogeneity of variance. Data natural log transformation for fixing normality of the
data. Homogeneity of variance was violated. Thus, the data were analyzed by using Kruskal–Wallis test
non parametric test equivalent of One-way analysis of variance.

Table 16. Impact of Solanum lycopersicum L grafting on Type-D trichome density at three
plant heights 25 days after transplanting into plastic pot. Lower region (leaves from second
branch), Mid-region (leaves from a mid-region branch), and Upper region (leaves from last
branch one before merry stem).

Lower region
Mid region
Upper region

N

Type-D Trichomes

SE

36

26.44 a1

3.80

36

42.25 b

5.19

36

76.83 c

11.08

1

Data were natural log transformed. The data did not violated Homogeny of variance after
natural log transformation. The data analyzed by One Way ANOVA and followed by Fisher’s
LSD for mean separation. Different letter show significant difference at α=0.05

103

Table 17. Impact of Solanum lycopersicum L grafting on Type-D Trichrome density 25
days after transplanting into plastic pot. Means for six replicates of each grafting
treatments. Three leaf samples were taken from three leaves of the each tomato plant.

Grafting

N

Mean

SE

Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

18

10.59

18

43.721
58.44

18

70.67

12.81

18

49.39

8.03

18

37.72

7.48

18

31.11

7.22

1

17.95

Data were natural log transformed. The data did not violated Homogeny of variance after
natural log transformation. The data analyzed by One Way ANOVA.

104

Appendix B. Figures
Figure 21. Mini-insectary for T. vaporariorum population density bioassay. A: Young
grafted tomato seedling in a mini-insectary. B: Experimental units after inoculation with T.
vaporariorum in a completely randomized design under growth chamber conditions.
B

A

105

Figure 22. Tomato leaf fixed with tape to make the leaf flat to assist counting whiteflies and
tomato trichomes.

106

Trialeurodes vaporariorum population density.

Figure 23. Relationship between lower region Type-D trichome density and total T.
vaporariorum population density.

Type-D trichome population for lower region leaves.

107

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108

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Gorman, K., Devine, G., Bennison, J., Coussons, P., Punchard, N., & Denholm, I. (2007). Report of
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CHAPTER 5: JOINT IMPACT OF MELOIDOGYNE INCOGNITA (KOFOID AND
WHITE1919) CHITWOOD 1949 AND WESTWOOD ON GRAFTED SOLANUM
LYCOPERSICUM L. CULTIVARS: WITH SPECIAL REFERENCE TO THE MI GENE.
Abstract
The impact of root-scion grafting combinations of the Mi gene containing Solanum lycopersicum
L cv, Anahu with cv Rutgers (Mi gene absent) on Trialeurodes vaporariorum population density
was evaluated under growth chamber conditions in no choice assays in the presence of
Meloidogyne incognita. The treatments included homo-grafted Anahu, homo-grafted Rutgers,
hetero-grafted Anahu and hetero-grafted Rutgers. Non-grafted Anahu and Rutgers served as the
controls. The hypotheses were that the presence of M. incognita triggers T. vaporariorum
resistance conferred by the Mi gene, or an increase in Type-D trichome density. In this research,
there were no significant differences in Type-D trichome density between cvs Anahu and
Rutgers. The presence of M. incognita triggered resistant to T. vaporariorum on non-grafted
Anahu and homo-grafted Anahu, compared to non-grafted Rutgers and homo-grafted Rutgers.
Moreover, the presence of M. incognita delayed the life cycle of T vaporariorum in all grafted
treatments. In our previous study in the absence of M. incognita, the Mi gene in Anahu did not
impact T. vaporariorum population density. Anahu was partially resistant to T. vaporariorum in
the presence of the M. incognita, while also expressing partial resistance to M. incognita. In
conclusion, S. lycopersicum cv Anahu does not appear to be an optimal cultivar for T.
vaporariorum or M. incognita management, despite being the parent of most Mi gene carrying
commercial S. lycopersicum cultivars and hybrids.
Introduction
Trialeurodes vaporariorum (Westwood) is a key pest of Solanum lycopersicum L, especially
under greenhouse conditions. T. vaporariorum is difficult to control because of its rapid rate of
population growth and the fact that most currently available chemical pesticides are only highly
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effective against adults. It’s high reproduction increases the risk associated with rapid
development of resistance to insecticides. Furthermore, T. vaporariorum causes indirect damage
by transmitting plant viruses during feeding and probing of a plant (Jones, 2003). According to
Duffus et al. (1996) T. vaporariorum is a vector of Tomato Infectious Chlorosis Virus (TICV).
Therefore, it is of major interest to have plants that are not only resistant to T. vaporariorum, but
also repellant in order to prevent the spread of TICV during white fly probing.
Tomato leaves possess trichomes which can provide both mechanical and chemical defenses
against insects. Seven types of trichomes exist in tomato plants (Tissier 2012). They are divided
in to two groups based on glandular cell presence at the tip of the trichome. Glandular trichomes
are responsible for plant resistant and repellency to pests in tomato (Goffreda, Mutschler, and
Tingey 1988). Among glandular trichomes, Type-D, otherwise known as Type VI, is
responsible for mechanical and chemical resistance (Aina, Rodriguez, and Knavel 1972).
Tomato breeding programs have identified and extensively studied several wild species of S.
lycopersicumI that are resistant to T. vaporariorum. De Ponti et al. (1975) discovered and
evaluated several wild tomato species, including: Lycopersicon hirsutum, L. hirsutum glabratum
and S. pennellii that are resistant to T. vaporariorum. Bas et al. (1992) mentioned that older L.
hirsutum f. glabratum plants were highly resistance to T. vaporariorum. Moreover, some wild
tomato species possess the Mi gene which was introduced into S. lycopersicum through the
embryo rescue method (Smith 1944). Among Mi-1 homologues, the Mi-1.2 gene has conferred
resistance to several organisms including Meloidogyne incognita (Kofoid & White) Chitwood
(Milligan et al., 1998; Seah, Telleen, & Williamson, 2007), and cross-resistance to both
Macrosiphum euphorbiae (Thomas) (Goggin, Williamson, & Ullman, 2001; Vos et al., 1998)
and sweet potato whitefly, Bemisia tabaci (Gennadius) (Nombela, Williamson, & Muñiz, 2003;

113

Goggin et al., 2001). Utilization of resistant S. lycopersicum has not, however, proven to be an
achievable way to control T. vaporariorum.
Breeding for new hybrids or cultivars is time consuming and not all favored traits of the parents
are transferred to the next generation. Grafting provides an alternative approach to integrate both
the pest resistant and food quality traits desired for successful S. lycopersicum production under
greenhouse or field conditions. The objective of this research is to evaluate the impact of a
grafted S. lycopersicum resistant rootstock on the population development of T. vaporariorum in
the presence of the nematode, M. incognita. Furthermore, to determine if the Mi insect
resistance trait of cv. Anahu can be transferred to grafted cv. Rutgers in the presence of M.
incognita. Additionally, the research involves determination of the relationship between M.
incognita and S. lycopersicum Type-D trichome density at three plant heights: at lower region,
mid region and upper region.
Materials and Methods
The research was conducted under growth chamber conditions at Michigan State University.
This section describes the procedures used to determine the population density evaluation of T.
vaporariorum associated with Anahu and Rutgers plants, before and after grafting. It includes
the experimental design, plant materials, grafting procedures, growth chamber conditions, T.
vaporariorum population density bioassay, trichome type density, data analysis and
abbreviations.
Experimental Design
The research consists of six treatments: 1) non-grafted Anahu, 2) non-grafted Rutgers, 3) homografted Anahu, 4) homo-grafted Rutgers, 5) hetero-grafted Anahu (Anahu scion and Rutgers
rootstock) and 6) hetero-grafted Rutgers (Rutgers scion and Anahu rootstock). Two other
treatments were the presence of T. vaporariorum, replicated three times, and the absence of T.
114

vaporariorum, replicated five times. The plants were maintained in a completely randomized
design under growth chamber conditions for 20 to 25 days after being transplanted into 32 oz
styrofoam cups.
Plant Materials
A single tomato cv Anahu (University of Hawaii Seed Program) or cv Rutgers (Sustainable Seed
Co.) was sown in each cell of 50 cell seedling trays containing plant growth media (SUREMIX
professional growing media, Michigan Grower Products Inc.) and maintained under growth
chamber conditions at 26 C for 16 hours and 20 C for eight hours, day and night, respectively.
The seedlings were watered every two days. Anahu seeds were planted two days before Rutgers
seeds to ensure the same seedling diameter for grafting. This time-interval was based on
preliminary research results. At the termination of the experiment, after 20-25 days, the fresh
and dry weights of the combined stem and leaf tissues, as well as the fresh and dry weights of the
root tissue were determined using a 200 g scale.
Grafting
After 14 days under growth-chamber conditions, the seedlings were grafted. Rootstocks and
scions were cut below the cotyledon, at a 45-65 degree angle using a Miter-Cut Grafting Knife
(Johnny's Selected Seeds). Scions and rootstocks were clipped together with silicon tubes used
to hold the rootstock and scion together until the grafting wound healed. Silicon clipper (HydroGardens, Colorado Springs, CO) size varied from 1.5-2.5 mm, depending on seedling diameter.
During graft-wound healing, a humidifier (Air Innovations Model # HUMID06 1.37-gal.
Ultrasonic Digital Humidifier) was used to control relative humidity (RH). The grafted plants
were maintained in a growth chamber for seven days at 24-26 C. The RH was decreased
gradually from 90% for the first three days, 85% the next two days, 75% the next day and 65%

115

on the final day. After seven days, the grafted seedlings were moved to the laboratory for three
days for acclimatization.
Growth Chamber
The tomato plants were transplanted into 32 oz styrofoam cups, filled with steam sterilized sandy
soil. Growth chamber temperature was maintained at 26 C for 16 hours and 20 C for eight hours,
day and night, respectively. The plants were watered every two days, or as necessary.
Trialeurodes vaporariorum Population Density Bioassay
A colony of T. vaporariorum was maintained on Rutgers, a susceptible tomato cultivar, under
growth chamber conditions. For each experimental unit, five females and at least one male T.
vaporariorum were introduced into mini-insectaries in two-dram vials placed under each tomato
plant. The insectary was constructed from a clear plastic jar with an opening of 3.5 inches designed
to fit snugly into the 32 oz cup containing the tomato plant. The insectary had openings on each
side and on the top. The openings were covered with fine mashed nylon cloth to prevent white fly
escape and allow for air flow. The insectaries and tomato plants were removed from the growth
chamber after 20-25 days. The abaxial sides of leaves on each plant were observed for T.
vaporariorum eggs, nymphs, pupa, and pupa cases. They were counted using a binocular
microscope at 60X magnification and the data recorded.
Meloidogyne incognita Population Density
The root systems of plants inoculated with M. incognita and expressing root galls were used for
estimating final female population densities of M. incognita. From each experimental unit, three
grams of root tissue were used to estimate the final female population density. Root tissue for
nematode analysis was cut at random from various parts of each root system. A slightly modified
root staining procedure of Bybd Jr et al. (1983) was used to determine nematode female final
population density. Three grams of root tissue were wrapped with a fine mashed nylon cloth
116

tagged with corresponding treatment label. This prevented loss of root tissue and allowed for the
staining of 20-40 roots at a time. Staining several roots together shortened the time spent for
staining and increased staining efficacy. Moreover, staining several roots together increased the
staining quality by accurately timing the procedure. Fixed roots were placed in 600 ml beakers
containing 1:1 ratio of 8.25% NaOCl (Clorox Regular Bleach 3.78qt) and tap water for four
minutes, stirring several times. The roots were rinsed with tap water and soaked for 15 minutes in
tap water. They were then drained and washed under tap water several times to remove the rest of
the bleach from roots. After washing, the roots were soaked in water for 15 minutes. Next, the
roots were placed into a 600-ml beaker and filled with water until all the fixed roots were
submerged. Based on how much water was used to fill the beaker, 1.0 ml of stock acid-fuchsinestain solution was added for each 30-50 ml water. The beaker was placed on a hot plate, brought
to a boil for one-two minutes and cooled at room temperature. The room-temperature roots were
drained, rinsed with tap water and dried with a paper towel. They were then filled with acidified
glycerin with a few drops of 5N HCl and boiled on a hot plate for 30–60 seconds. After cooling,
the final population density (Pf) of M. incognita infesting the roots was determined under a
binocular microscope.
Trichome Types
Glandular trichomes, known as Type-D or Type VI trichomes (Tissier 2012), were counted on
the abaxial side of the leaves under a 60X binocular microscope. The determinations were made
at three plant height levels: upper, mid and lower. Lower leaves were selected from a position
immediately above the first leaf branch. The upper leaves from the upper most leaf branch and
mid leaves from a branch in the middle of the plant.

117

Data Analysis
The data were analyzed using SPSS 24 Grad Primum Pack. First, data normality was checked with
a histogram plot for skewness of the distribution. If the data were positively skewed, natural Log
or Log10 was used to transform the data. If the data were negatively skewed, the Square Root
Method was applied. Normalized data after transformation were analyzed with a One Way
ANOVA test, followed by a Fisher’s LSD pairwise test for mean separation if the ANOVA p value
was equal or less than α=0.05. If the data were not normalized after transformation, the nonparametric Kruskal-Wallis Test was used as the substitute for the One Way ANOVA, followed by
a Dunn pairwise test for mean separation if the Kruskal-Wallis Test p-value was equal or less than
α=0.05.
Results
Greenhouse Whitefly
After 20-25 days under growth chamber conditions at 26 C for 16 hours and 20 C for eight hours,
day and night, respectively, T. vaporariorum population densities were significantly different
(F5,90=2.788, P=0.022) among the six treatments. Homo-grafted Anahu had the lowest population
density of T. vaporariorum and it was significantly lower than all other treatments, except nongrafted Anahu (Table 18). Non-grafted Anahu had a significantly lower population density of T.
vaporariorum than Non-grafted Rutgers. Hetero-grafted Anahu had the highest population density
of T. vaporariorum, although it was not significantly different than hetero-grafted Rutgers.
Type-D Trichome Density in the Presence of M. incognita
There were no Type-D trichome density differences among the six treatments after seven days of
plant growth in the presence of M. incognita (F5,102=1.286, P=0.276). The highest Type-D
trichomes density was observed on hetero-grafted Rutgers (Table 19). The lowest number of
Type-D trichomes density was associated with non-grafted Rutgers. There were significant
118

differences in Type-D trichome densities relative to the vertical location of leaves (F2,102=11.235,
P=0.001). The greatest density of Type-D trichomes, were found on the most recently formed
leaves (Upper region leaves) (Table 20). Leaves at this location had significantly more Type-D
Trichomes than those at the mid and lower positions (P=0.004 and P=0.001, respectively).
Twenty-five days after inoculation with M. incognita, trichome density did not differ significantly
among the six treatments (F5,102=0.889, P=0.491). Numerically, the greatest number of Type-D
trichome was observed on hetero-grafted Anahu plants. The lowest numbers were associated with
the homo-grafted Anahu plants (Table 21). There was a significant difference in Type-D trichome
density on leaves at different heights (F2,105=7.774, P=0.001). Type-D trichome density observed
on the upper region leaves was statistically greater compared to those observed on the lower region
(P=0.001) and mid region (P=0.007) leaves (Table 22).

Stem/leaf Biomass in the Presence and Absence of Trialeurodes vaporariorum
Fresh weight of stem/leaf tissue was not significantly different among the six treatments when T.
vaporariorum was absent (H5=1.139, P=0.951). Numerically, the greatest fresh weight was
observed on non-grafted Anahu. The lowest stem/leaf tissue fresh weight was observed on heterografted Anahu (Table 23).
Dry weight of stem/leaf tissue was not significantly different among six grafted treatments when
T. vaporariorum was absent (F5.24=1.778, P=0.156).

Numerically, the highest amount of

stem/leaf biomass was observed on non-grafted Rutgers and the lowest amount on homo-grafted
Anahu (Table 24).
When T. vaporariorum was present, the fresh weight of stem/leaf tissue was substantially greater
than in the absence of this insect (Table 24, Table 25). The fresh weight, however, was not
significantly different among the six treatments (F5,12=1.152, P=0.387). Numrically, the greatest

119

amount stem/leaf fresh weight was observed on hetero-grafted Anahu. The lowest amount of
stem/leaf tissue fresh weight was observed on homo-grafted Anahu.
Dry weights of stem/leaf tissue also were not significantly different amount the six treatments
(F5,12=0.518, P=0.758). Numerically, the greatest amount of dry weight of stem/leaf tissue was
observed on hetero-grafted Anahu (Table 26). The lowest amount of dry weight of stem/leaf tissue
was observed on homo-grafted Anahu. While the fresh weight of stem/leaf tissue was not
significantly different in the presence or absence of T. vaporariorum, dry weight of stem/leaf tissue
was significantly less (P=0.03) in the presence of T. vaporariorum compared to when this insect
was absent (Table 23 and Table 26).
Root Biomass in the Presence and the Absence of the Trialeurodes vaporariorum
In the absence of T. vaporariorum, there were significant differences in root fresh weights
among the treatments (F5.24=3.183, P=0.024). Non-grafted Rutgers had the greatest root fresh
weight and it was not significantly different than non-grafted Anahu (Table 27). Homo-grafted
Anahu had the lowest root tissue fresh weight among the six treatments and it was significantly
less than homo-grafted Rutgers (P=0.029). Hetero-grafted Anahu and hetero-grafted Rutgers
had similar root tissue fresh weights.

In the presence of T. vaporariorum, root tissue fresh weights were not significantly different
among the six treatments (H5=8.196, P=0.146). Numerically, hetero-grafted Anahu produced the
greatest amount of root tissue fresh weight in the presence of the T. vaporariorum (Table 28).
Hetero-grafted Anahu had the lowest amount of root tissue fresh weight.
There were significant differences in dry weights of root tissue among the six treatments in the
absence of T. vaporariorum (F5,24=6.073, P=0.01). The greatest amount of root tissue dry weight
was associated with non-grafted Rutgers and it was significantly more than non-grafted Anahu

120

(P=0.026) (Table 29). Homo-grafted Anahu produced the lowest amount of root tissue dry
weight and it was significantly less than homo-grafted Rutgers (P = 0.003). Hetero-grafted
Anahu and hetero-grafted Rutgers had similar amounts of root tissue dry weight.
In contrast, In the presence of T. vaporariorum there were no significant differences among the
six treatments (H5=9.117, P=0.102). numerically, the greatest amount of root tissue dry weight
was associated with non-grafted Rutgers and the lowest root tissue dry weight was observed on
hetero-grafted Rutgers (Table 30). Overall, there were no significant differences in fresh and dry
root weights when T. vaporariorum was present or absent.

Meloidogyne incognita Population Development in the Presence and the Absence of
Trialeurodes vaporariorum
In the absence of T. vaporariorum, M. incognita final female population densities were not
significantly different among the six treatments (F2,24=2.410, P=0.066) (Table 30). Numerically,
the highest number of M. incognita females was observed on homo-grafted Rutgers. The lowest
number M. incognita females were observed on homo-grafted Anahu. When T. vaporariorum
was present, the M. incognita final female population densities also were not significantly
different (Table 32). The highest and lowest population densities were observed on non-grafted
Rutgers and hetero-grafted Rutgers, respectively.

Biomass Partitioning in the Presence and Absence of Trialeurodes vaporariorum and
Meloidogyne incognita
Plants associated with all six treatments allocated greater biomass towards to stem/leaf compared
to root tissue (Figure 24). After grafting, homo-grafted Anahu and Hetero-grafted Anahu shifted
their biomass allocation more towards stem/leaf tissue than root tissue, compared to non-grafted
Anahu. Grafting also impacted homo-grafted Rutgers and hetero-grafted Rutgers, where

121

biomass allocation shifted to stem/leaf compared to root tissue associated with non-grafted
Rutgers. When T. vaporariorum was present, non-grafted Anahu shifted its biomass towards
stem/leaf rather than root tissue (Figure 25). The same effect was observed on non-grafted
Rutgers where 2% of biomass shifted from root to stem/leaf in the presence of T. vaporariorum.
Homo-grafted Anahu did not follow this pattern, but rather shifted 1% towards to root fresh
weight in the absence compared to the presence of T. vaporariorum. On homo-grafted Rutgers,
biomass shifted more towards to stem/leaf in the presence of T. vaporariorum. Hetero-grafted
Anahu biomass partitioning was identical to homo-grafted Anahu, but compared to heterografted Anahu in the absence it partitioned 1% less of its biomass to stem/leaf than root fresh
weight. Hetero-grafted Rutgers had the highest biomass partitioned towards stem/leaf among the
six treatments. Hetero-grafted Rutgers shifted 4% of the biomass from root fresh to stem/leaf
weight in the presence of T. vaporariorum.
Relationship between Solanum lycopersicum L Root Fresh Weight and Trialeurodes
vaporariorum
Fresh root weight was used as a predictor variable and T. vaporariorum population density used
as response variable. When 18 data points were pooled together from the six treatments, fresh
root weight used to predict T. vaporariorum population density with the increase of the root fresh
weight the T. vaporariorum population density increased (Figure 26). When data points were
grouped by grafting treatments, the results were very different. Non-grafted Anahu had a
negative impact on T. vaporariorum population density (Figure 27,Figure 28). An increase in
root fresh weight of non-grafted Rutgers had no impact on the T. vaporariorum population
density (Figure 27Figure 28). Homo-grafted Anahu did not impact the T. vaporariorum
population density with its increase in root fresh weight. Homo-grafted Rutgers, however, had a
positive impact on the T. vaporariorum population density associated with the root fresh weight

122

increase (Figure 28). T. vaporariorum population density increases for hetero-grafted Anahu and
hetero-grafted Rutgers were associated with the increase in root fresh weight (Figure 29).
Discussion
This study was carried out to test the impact of grafting on T. vaporariorum in the presence of M.
incognita. Not all Mi gene resistant S. lycopersicum cultivars possess complete resistant to T.
vaporariorum. Partial resistance, however, is important in pest control and can be responsible for
decreases in chemical pesticide application. Thus, we evaluated the impact of grafting on
resistance of two S. lycopersicum cultivars, Anahu (Mi gene resistance reported to Bemisia tabaci)
and Rutgers (Mi gene absent) after 20-25 days under growth chamber conditions.
Mini insectaries were used to prevent T. vaporariorum dispersal from the plants. This mini
insectary system had both advantages and disadvantages. The advantage is that the mini insectary
prevented T. vaporariorum dispersal, allowing us to accurately determine the exact population
density of T. vaporariorum. The disadvantage is that the material used to build the insectaries can
prevent direct light and also increase temperature inside the insectary (Crafts-Brandner & Chu,
1999). In our five-female oviposition system, the total number of eggs produced by five female
of T. vaporariorum ranged from 25-129, with a mean of 72 per female at 26 C. This is consistent
with the approximately 30 eggs per female at 27 C and 124 eggs per female at 24 C reported by
Burnett (1949). T. vaporariorum oviposition rate is known to decrease with temperature above 18
C when reared on tomato plants (Burnett 1949). Thus, the number of eggs laid per female in our
study appeared to not be impacted by our caging method.
Our findings suggest that the rootstock Anahu transfers its resistance to the susceptible Rutgers
scion. Although T. vaporariorum population densities on hetero-grafted Rutgers (Rutgers scion
and Anahu rootstock) were not significantly different than those associated with non-grafted
Rutgers or homo-grafted Rutgers, there was a pattern indicating a decrease in the T. vaporariorum
123

population density on hetero-grafted Rutgers plants (Table 18). The results of our study are in
agreement with findings of Lucatti (2010), where the Mi gene carrying S. lycopersicum cv Uco
Plata supported a lower T. vaporariorum population density in a free choice assay, compared to
plants without the Mi gene. In our research, however, all experimental units were inoculated with
M. incognita Pi=5000.

T. vaporariorum developed significantly less on non-grafted Anahu,

compared to non-grafted Rutgers. A possible explanation for this is the Mi gene or related gene
responsible for resistance to T. vaporariorum was dormant or turned-off in the absence of M.
incognita and functional in the presence of M. incognita, making the plants more resistant to T.
vaporariorum. Interestingly, T. vaporariorum population density was less on homo-grafted Anahu
and homo-grafted Rutgers compared to their non-grafted peers. A possible explanation for this is
that grafting made the plant more vigorous after homo-grafting (Khah, Kakava, Mavromatis,
Chachalis, & Goulas, 2006).
Type-D trichomes are partially responsible for resistant to pests in tomato (Aina, Rodriguez, and Knavel
1972). In this study, the trichomes were not evenly distributed on the leaves of S. lycopersicum cvs

Anahu and Rutgers in the presence of M. incognita. The Type-D trichome density was greatest in
the upper region of both cultivars. In addition, we did not find significant differences in trichome
Type-D density between Anahu and Rutgers. Thus, we propose that Type-D trichome is not
responsible for T. vaporariorum resistance in Anahu. Nombela et al. (2000), similarly, found no
association between Type-D trichome density and resistance to Bemisia argentifolii and low TypeD trichome density was associated on Mi gene carrying S. lycopersicum cultivars compared to
cultivars without the Mi gene. The presence of M. incognita did not impact the overall trichome
Type-D density.

124

Stem and leaf tissue fresh and dry weights were not impacted by the presence of T. vaporariorum.
In contrast, Pofu (2012) found that the presence of T. vaporariorum and M. javanica decreased the
dry weight of shoot and root tissues by 86 % and 60 %, respectively. We found that root tissue
fresh and dry weight differed when T. vaporariorum was absent, however when T. vaporariorum
was present, the weights were not significantly different.

In addition, the presence of T.

vaporariorum increased the M. incognita final female population density in all grafted treatments
except hetero-grafted Rutgers. Similarly, Pofu et al. (2012) found that M. javanica population
density increased in wild watermelon in the presence of T. vaporariorum.
Biomass partitioning of S. lycopersicum cvs Anahu and Rutgers was not impacted by the presence
of T. vaporariorum. When all of the root tissue fresh weight data from the six grafted treatments
were pooled, and regressed against the T. vaporariorum population density, there was a positive
correlation (Table 20). One possible explanation is that homo-grafted Rutgers, hetero-grafted
Anahu, and hetero-grafted Rutgers had positive correlations, with the correlation being close to
1.00, thus the pooled date became a positively correlated (Figure 28,Figure 29). The facts that
non-grafted Anahu and non-grafted Rutgers had negative correlations and homo-grafted Anahu
had a correlation that was close to 0.00 decreased the positive correlation coefficient of the pooled
data. This is an excellent example why it is important to evaluate both pooled date and from the
individual grafting treatment.
Conclusion
S. lycopersicum cv Anahu is only partially resistant to T. vaporariorum in the presence of M.
incognita. In this research, we proved experimentally that the presence of the M. incognita delayed
life cycle of the T. vaporariorum population density. Nevertheless, partial resistance can be
beneficial in the management of T. vaporariorum when used with conjunction of chemical
insecticides. The action threshold for T. vaporariorum is ten females or eight nymphs per tomato
125

leaf (Polack, 2005). In our research, the initial population density of T. vaporariorum did not
exceed Polack’s threshold in any of our grafting treatments. Thus, we conclude that it is necessary
to conduct future research that will involve multiple generations of T vaporariorum. Moreover,
the presence of T. vaporariorum increased the M. incognita final female population density.

126

APPENDICES

127

Appendix A. Statistical Tables
Table 18. T. vaporariorum population density on grafted Solanum lycopersicum after 25
days in the presence of Meloidogyne incognita.
Grafting Treatment
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

N

Mean

SE

3
3
3
3
3
3

48.00 ab1
26.67 a
74.67 b
100.00 c
87.67 bc
93.00 c

5.03
1.20
7.22
19.47
23.25
12.42

Data analyzed by One Way ANOVA. If the ANOVA is statistically significant in α=0.05
followed by Fisher’s LSD for separation of means. Different letter in a column are statistically
significant at α=0.05.
1

Table 19. Grafted Solanum lycopersicum Type-D trichome density after seven days in the
presence of Meloidogyne incognita.
Grafting Treatment
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers
1

N

Mean

SE

18
18
18
18
18
18

56.831
32.89
45.50
66.61
27.00
39.72

12.69
6.67
11.06
18.42
6.94
7.05

Data violated the Homogeny of variance. Natural log transforming the data normalized. The
data analyzed by One Way ANOVA. If the ANOVA is statistically significant in α=0.05
followed by Fisher’s LSD. Different letter in a column are statistically significant.

128

Table 20. Solanum lycopersicum L Type-D Trichome density at three plant heights after
seven days in the presence of M. incognita.
Leaf location

N

Mean

SE

Lower region
Mid region
Upper region

36
36
36

22.83 a1
38.81 a
72.64 b

2.83
6.37
10.77

1

Data violated the Homogeny of variance. Natural log transforming the data normalized. The
data analyzed by One Way ANOVA. If the ANOVA is statistically significant in α=0.05
followed by Fisher’s LSD. Different letter in a column are statistically significant.
Table 21. Grafted Solanum lycopersicum Type-D trichome density after 25 days in the
presence of Meloidogyne incognita.
Grafting Treatment
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

N

Mean

SE

18
18
18
18
18
18

61.221
49.78
65.17
60.00
51.72
30.61

13.25
13.17
15.83
16.56
12.36
4.78

1

Data violated the Homogeny of variance. Natural log transforming the data normalized. The
data analyzed by One Way ANOVA. If the ANOVA is statistically significant in α=0.05
followed by Fisher’s LSD. Different latter on a column are statistically significant.
Table 22. Solanum lycopersicum L Type-D Trichome density at plant three plant heights
after 25 days in the presence of Meloidogyne. incognita.
Leaf location

N

Mean

SE

Lower region
Mid region
Upper region

36
36
36

31.81 a1
39.00 a
88.44 b

4.72
4.06
13.15

1

Data violated the Homogeny of variance. Natural log transforming the data normalized. The
data analyzed by One Way ANOVA. If the ANOVA is statistically significant in α=0.05
followed by Fisher’s LSD. Different letter in a column are statistically significant.

129

Table 23. Grafted Solanum lycopersicum fresh weight of stem and leaf tissue after 24 days
in the presence of Meloidogyne incognita.
Grafting Treatment
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

N

Mean (g)

SE

5
5
5
5
5
5

48.261
47.39
46.37
46.06
46.43
46.95

2.31
2.44
3.25
1.84
1.56
0.35

1

Data violated the Homogeny of variance. Natural log or log 10 transforming the data did not
normalize it. Means of experiment analyzed by Kruskal-Wallis Test. If there was statistical
significant at α=0.05 followed by Mann-Whitney U.
Table 24. Grafted Solanum lycopersicum dry weights of stem and leaf tissue after 24 days in
the presence of with Meloidogyne incognita.
Grafting Treatment
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

N

Mean (g)

SE

5
5
5
5
5
5

6.841
6.37
7.60
6.46
6.87
7.79

0.16
0.43
0.65
0.56
0.30
0.34

The data analyzed by One Way ANOVA. If the ANOVA is statistically significant in α=0.05
followed by Fisher’s LSD. Different letter in a column are statistically significant.
1

130

Table 25. Impact of Trialeurodes vaporariorum to grafted Solanum lycopersicum fresh
weight of stem/leaf after 24 days in the presence of Meloidogyne incognita.
Grafting Treatment
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

N

Mean (g)

SE

3
3
3
3
3
3

49.811
44.78
48.83
51.49
49.00
48.45

2.29
2.69
1.13
2.32
2.22
1.22

The data analyzed by One Way ANOVA. If the ANOVA is statistically significant in α=0.05
followed by Fisher’s LSD. Different letter in a column are statistically significant.
1

Table 26. Impact of Trialeurodes vaporariorum on grafted Solanum lycopersicum dry
weight of stem and leaf tissue after 24 days in the presence of Meloidogyne incognita.
Grafting Treatment
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

N

Mean (g)

SE

3
3
3
3
3
3

6.121
5.71
6.19
6.48
6.36
5.83

0.40
0.64
0.35
0.21
0.51
0.19

Data analyzed by One Way ANOVA. If the ANOVA is statistically significant in α=0.05
followed by Fisher’s LSD. Different letter in a column are statistically significant.
1

Table 27. Grafted Solanum lycopersicum fresh weight of root tissue after 24 days in the
presence of Meloidogyne incognita.
Grafting Treatment
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

N

Mean (g)

SE

5
5
5
5
5
5

14.94 abc1
12.76 a
15.16 bc
13.22 ab
15.23 bc
16.30 c

0.73
0.67
0.62
0.61
0.88
0.94

The data analyzed by One Way ANOVA. If the ANOVA is statistically significant in α=0.05
followed by Fisher’s LSD. Different letter in a column are statistically significant.
1

131

Table 28. Grafted Solanum lycopersicum dry weight of root tissue after 24 days in the
presence of Meloidogyne incognita.
Grafting Treatment
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

N

Mean (g)

SE

5
5
5
5
5
5

0.85 bc1
0.65 a
0.90 cd
0.67 ab
0.95 cd
1.07 d

0.07
0.06
0.06
0.05
0.06
0.08

The data analyzed by One Way ANOVA. If the ANOVA is statistically significant in α=0.05
followed by Fisher’s LSD. Different letter in a column are statistically significant.
1

Table 29. Impact of Trialeurodes vaporariorum on grafted Solanum lycopersicum root fresh
weight after 24 days in the presence of Meloidogyne incognita.
Grafting Treatment
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers
1

N

Mean

SE

3
3
3
3
3
3

14.401
13.45
12.66
15.61
14.70
15.39

0.24
1.22
0.69
0.31
1.09
0.39

Data violated the Homogeny of variance. Natural log or log 10 transforming the data did not
normalize it. Means of experiment analyzed by Kruskal-Wallis Test. If there was statistical
significant at α=0.05 followed by Mann-Whitney U.

132

Table 30. Impact of Trialeurodes vaporariorum on grafted Solanum lycopersicum root dry
weight of root tissue after 24 days in the presence of Meloidogyne incognita.
Grafting Treatment
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

N

Mean (g)

SE

3
3
3
3
3
3

0.731
0.64
0.62
0.87
0.87
0.87

0.04
0.13
0.02
0.04
0.08
0.02

1

Data violated the Homogeny of variance. Natural log or log 10 transforming the data did not
normalize it. Means of experiment analyzed by Kruskal-Wallis Test. If there was statistical
significant at α=0.05 followed by Mann-Whitney U.
Table 31. Meloidogyne incognita final female population density on six grafted S.
lycopersicum L. treatments.
Grafting Treatment
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

N

Mean

SE

5
5
5
5
5
5

459.00
347.66
898.01
1144.21
1149.32
893.52

291.42
204.09
254.72
95.84
137.81
261.33

Table 32. Meloidogyne incognita final female population density on six grafted S.
lycopersicum L treatments in the presence of Trialeurodes vaporariorum.
Grafting Treatment
Anahu
Anahu/Anahu
Rutgers/Anahu
Anahu/Rutgers
Rutgers/Rutgers
Rutgers

133

N

Mean

SE

3
3
3
3
3
3

1134.40
688.05
493.52
1759.56
1522.29
2153.44

587.04
648.71
474.41
437.62
539.14
357.74

Appendix B. Figures
Figure 24. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers in the presence of M. incognita (Pi = 5000).

134

Figure 25. Biomass partitioning signatures of Solanum lycopersicum L. cvs Anahu and
Rutgers in the presence of M. incognita (Pi =5000) and T. vaporariorum.

135

Figure 26. Relationship between S. lycopersicum root fresh weight and T. vaporariorum
population density per plant.

136

Figure 27. Relationship between S. lycopersicum L cv Anahu and cv Rutgers root fresh
weight and T. vaporariorum population density per plant.

137

Figure 28. Relationship between S. lycopersicum L homo-grafted Anahu and homo-grafted
Rutgers root fresh weight and T. vaporariorum population density per plant.

138

Figure 29. Relationship between S. lycopersicum L hetero-grafted Anahu and heterografted Rutgers root fresh weight and T. vaporariorum population density per plant.

139

Appendix C. Specimens
RECORD OF DEPOSITION OF VOUCHER SPECIMENS

The specimens listed below have been deposited in the named museum as samples of those
species or other taxa, which were used in this research. Voucher recognition labels bearing the
voucher number have been attached or included in fluid preserved specimens.
Voucher Number: _____2017-03__________
Author and Title of thesis:
Bahodir Ruzumboevich Eshchanov
“Influence of Mi Genes on Grafted Solanum Lycopersicum L. Cultivars for Control of
Trialeurodes Vaporariorum (Insecta) in the Presence and Absence of Meloidogyne
Incognita (Nematoda).”
Museum(s) where deposited:
Albert J. Cook Arthropod Research Collection, Michigan State University (MSU)
Table 33. List of voucher specimens.
Family

Genus/Species

Life Stage

Quantity

Preservation

Aleyrodidae

Trialeurodes vaporariorum

Adult

5

Ethanol

Aleyrodidae

Trialeurodes vaporariorum

Pupa

5

Ethanol

Aleyrodidae

Trialeurodes vaporariorum

Nymph

5

Ethanol

140

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141

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CONCLUSION
A significant number of important conclusions can be drawn from the results of my Ph.D.
dissertation research. Some of these findings are completely new; whereas, others support or
contradict the discoveries of other scientists. The major findings of this Ph.D. research program
are:
•

Solanum lycopersicum cultivars Anahu and Rutgers have unique biomass
partitioning signatures and allocate their resources differently under greenhouse
conditions.

•

Anahu biomass allocation is not favorable for greenhouse S. lycopersicum
production systems.

•

Lightly and heavily pruned S. lycopersicum allow for more plants per unit area
and S. lycopersicum yield production is increased by increased plant numbers.

•

With the lack of environmental stress, grafted plants produced similar yields
under field conditions.

•

Fresh and dry biomass of homo-grafted and hetero-grafted plant were
significantly less than non-grafted plants at the early stages of the growth period.
This difference, however, was not present at the end harvest.

•

Biomass allocation is a cultivar specific characteristic and the biomass
partitioning signature of a rootstock can manipulate scion biomass allocation.

•

Understanding the biomass signature of a cultivar is important before selecting
rootstocks for vegetable grafting.

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•

Anahu, with the Mi resistant gene for M. incognita, was not completely resistant
to M. incognita.

•

Pruning significantly increased the final population density of M. incognita in
both resistant and susceptible plants.

•

Young Anahu plants with the Mi gene are not resistant to T. vaporariorum.

•

Anahu rootstock influenced Rutgers scion resulting in T. vaporariorum
population density being significantly less on hetero grafted Rutgers than nongrafted Rutgers and hetero-grafted Anahu.

•

The presence of M. incognita improved the Anahu’s resistance to T.
vaporariorum in 20 to 25 day-old plants.

•

T. vaporariorum resistance is not related to Type-D Trichome density in Anahu.

•

Type-D Trichome density was greater on upper, compared to lower or mid-region
leaves. This was not impacted by M. incognita.

•

Future research recommendations

•

Hetero-grafted and homo-grafted S. lycopersicum cultivars should be tested in the
presence of M. incognita under field conditions.

•

The entire life cycle of T. vaporariorum has to be tested under a distractive
experimental design in the presence of M. incognita on S. lycopersicum cultivar
Anahu that caries Mi gene.

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•

Anahu plants older than 20-25 days need to be tested for resistance to T.
vaporariorum.

We researched Anahu for biomass allocation and resistance to M. incognita and T.
vaporariorum. The results indicate that Anahu is not completely resistant to M. incognita and T.
vaporariorum. Furthermore, Anahu did not show any significant difference in response to T.
vaporariorum in the absence of the M. incognita. We proved that the Mi gene or other genes
that lowered the T. vaporariorum population significantly were turned on in the presence of M.
incognita. The most interesting finding is that the Anahu rootstock significantly impacted T.
vaporariorum population density on Rutgers scion in the presence of the M. incognita.

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