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dissertation entitled

The Influence of Mites and a Photosynthetic
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of Sour Cherry (Prunus cerasus L. 'Montmorency')

 

presented by

Mark Anthony Hubbard

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THE INFLUENCE OF MITES AND A PHOTOSYNTHETIC
INHIBITOR (TERBACIL) ON PHOTOSYNTHESIS AND YIELD OF
SOUR CHERRY (PRUNUS CERASUS L. 'MONTMORENCY')
By

Mark Anthony Hubbard

A DISSERTATION

Submitted to
Michigan State University
in partial fulﬁllment of the requirements
for the degree Of

DOCTOR OF PHILOSOPHY
Department of Horticulture

1995

ABSTRACT
THE INFLUENCE OF MITES AND A PHOTOSYNTHETIC INHIBITOR (TERBACIL)
ON PHOTOSYNTHESIS, CARBON ACCUMULATION AND YIELD OF SOUR
CHERRY (PRUNUS CERAS US L. 'MONTMORENCY')

By

Mark Anthony Hubbard

Orchard and container-grown sour chen'y (Prunus cerasus L. 'Montmorency')
trees were infested with two—spotted spider mites (Tetranychus urticae) to study mite—
feeding effects on photosynthetic parameters, cold hardiness, starch accumulation and
reproduction effort (ﬂowering and yield). In the ﬁeld, mite populations reached 1764
average cumulative miteOdays (CMD) for mite-infested trees (MIT) and 1093 CMD for
controls. Chlorophyll was reduced in MIT compared to controls late in the growing
season; however, there was no reduction during the season in measured photosynthetic
parameters, stem cambium cold hardiness or shoot starch levels. Additionally, miticides
used commercially in Michigan tart cheny orchards were determined to have no effect on
photosynthetic parameters. There was an increase in spring root starch levels of MIT the
spring following infestation, but no effect on return fruit set or return yield. In the
container grown trees, average CMD reached 917 for MIT and 90 for controls. Reductions
were observed in single leaf A, whole tree A and chlorophyll ﬂuorescence. This suggests
that mite populations sustained in the orchard in 1993 were not high enough to reduce
subsequent year yield, and therefore, control measures were of no economic beneﬁt. The

ﬁeld trees demonstrated an ability to buffer the effects of mite feeding and maintain

reproductive capacity. Reduction in photosynthesis does occur, as in the container study,
and that reduction in photosynthesis could ultimately affect yield and hardiness. In order to
study photosynthetic reduction thresholds that affect tree hardiness and yield, trees were
treated with 50-63 ppm terbacil to limit photosynthesis for speciﬁc periods of time.
Terbacil was applied to orchard trees at 4 physiological stages: fruit growth stages I and II,
fruit harvest, and prior to leaf fall; at each date, A was reduced up to 60% within 24 h and
showed 100% recovery within 7-10 days. Reductions were also observed in whole tree A
and chlorophyll ﬂuorescence, but not in chlorophyll, shoot cold hardiness, fruit yield or
fruit quality. Root and shoot starch levels did vary slightly dependent on the timing of

terbacil application but there was no effect on return yield.

DEDICATION

This dissertation is dedicated to Joanna for her sacriﬁce, love and encouragement. I
was told years ago that the spouse of the Ph.D candidate does as much work for the degree
than the candidate, and I believe it. This is also dedicated to Sarah who is a blessing from
God and is due a lot of time from her father who spent too long in the lab and ﬁeld during

her ﬁrst years.

iv

ACKNOWLEDGMENTS

I am forever indebted to Jim Flore for his guidance, patience, constant support and
his great help in completing this work. Thanks to Jim I have learned much more than could
ever be written down or told concerning pomology, horticulture and science in general.

Jim has also taught me many "cultural" things as well.

I am also grateful to my committee members for their encouragement and support.
Dr. Ken Poff has always been available for discussion as well as keeping things in
perspective with a sharp wit. Dr. Don Dickmann has provided much wisdom and
encouragement and I am grateful to him for all his advice. Dr. Stan Ries has taught me
many things and has provided much encouragement and advice. Also, I am thankful to Dr.
M. John Bukovac for his friendship and his ability to inspire, as well as his support.

This work would never have been started had it not been for Lynne Sage and it
would never have been ﬁnished without Sarah Breitzkreitz. I am grateful to both!!

There have been a host of people at Michigan State who have in one way or another
contributed to my education and this work in particular. I am thankful to Des, Tom, Mark,
Edgardo, Moreno, Ricardo, Claudio, Maurizio I, Maurizio 11, Brent, John, Rebecca, Beth,
Dave, Jon and the personnel at Clarksville and the Hort Farm. I have no doubt left
someone out.

Finally, I must acknowledge that my motivation and source of strength are in my

friend and savior Jesus Christ! His constant love saw this work through!

TABLE OF CONTENTS

LIST OF TABLES ................................................................................. x
LIST OF FIGURES ............................................................................. xii
ABBREVIATIONS .............................................................................. xiv
INTRODUCTION ................................................................................. 1

Literature Cited ............................................................................ 3

THE INFLUENCE OF MITES AND A PHOTOSYNTHETIC INHIBITOR (TERBACIL)
ON PHOTOSYNTHESIS AND YIELD OF SOUR CHERRY (PRUN US

CERAS US L. 'MONTMORENCY') ................................................... 6
Abstract ..................................................................................... 7
Materials and Methods .................................................................. 12
Plant Materials .................................................................. 12
Photosynthesis .................................................................. 12
Chlorophyll Fluorescence ..................................................... 14
Chlorophyll ..................................................................... l4
Carbohydrate Status ............................................................ 14
Cold Hardiness ................................................................. 15
Mite-Feeding Studies .......................................................... 16
Vegetative and Reproductive Growth ........................................ 17

vi

Miticide Study .................................................................. 17

Terbacil Studies ................................................................. l8
Vegetative and Reproductive Growth ........................................ 18
Statistical Protocol .............................................................. 19
Results .................................................................................... 19
Mite-Feedin g Studies .......................................................... 19
Orchard Study .................................................................. 19

Mite Populations ....................................................... 19

Gas Exchange and Fluorescence .................................... l9

Chlorophyll ............................................................ 2O

Vegetative and Reproductive Growth ............................... 20
Carbohydrate Status ................................................... 20

Cold Hardiness ........................................................ 20

Container Study ................................................................ 21

Mite Populations ....................................................... 21

Gas Exchange and Fluorescence .................................... 21

Effects of Miticides .................................................... 21

Terbacil Studies ................................................................. 22
Discussion ................................................................................ 24
Literature Cited .......................................................................... 34
SUMMARY AND CONCLUSIONS .......................................................... 49
APPENDICES .................................................................................... 53

Appendix A. Regression Figures of 1993 Orchard Single Leaf C02 Assimilation,
Whole Tree C02 Assimilation, and Chlorophyll Fluorescence against Mites Per

Leaf and Cumulative MiteODays ................................................... 51

vii

Appendix B. Cold Hardiness of Orchard Sour Cherry Trees Infested with Mites in
1993 .................................................................................. 58
Appendix C. Effects of Miticides on C02 Assimilation, Stomatal Conductance,
Transpiration and Chlorophyll Fluorescence of Greenhouse Grown Sour
Cherry Trees ......................................................................... 61
Appendix D. Effects of Differing Concentrations of Terbacil on C02 Assimilation
and Chlorophyll Fluorescence of Sour Cherry Trees ........................... 65
Appendix E. Shoot Extension and Chlorophyll Levels of Orchard Sour Cheny
Trees Treated with 63 ppm Terbacil on Speciﬁed Dates in 1993 .............. 68
Appendix F. Cold Hardiness of Orchard Sour Cherry Trees Treated with 63 ppm

Terbacil on Speciﬁed Dates in 1993 .............................................. 70
Appendix G. 1994 Terbacil Spray Methods and Results .......................... 73
Appendix H. Pesticide Residue Study ............................................... 77
Appendix I. Pesticide Residues Extraction Protocols .............................. 91
Appendix J. Data for 1993 Mite Population Counts ................................ 95

Appendix K. Data for 1993 Chlorophyll Fluorescence Measurements of Field
Study on Mites ...................................................................... 97
Appendix L. Effect of Mites on Shoot Growth and Flowering ................... 99
Appendix M. Effect of Terbacil on Fruit Yield and Quality in 1993 and 1994. 100
Appendix N. Chlorophyll Data Collected in 1993 on Mite Treated Trees ...... 102

Appendix 0. Yield Data Collected in 1993 and 1994 ............................. 104
Appendix P. Starch Data Collected in 1993 and 1994 ............................ 105
Appendix Q. Data for Container Grown Mite Study .............................. 106

Appendix R. Daily Minimum and Maximum Temperatures for 1993-1994 Dormant
Period at Clarksville Michigan ................................................... 1 11
Appendix S. Air ﬂow generating fan and inlet setup for whole plant photosynthetic

C02 assimilation chambers ....................................................... 114

Appendix T. Schematic drawing of whole plant photosynthetic C02 assimilation
chambers ............................................................................ 117

LIST OF TABLES

Table 1. The effect of mite-infestation on single leaf C02 assimilation of sour cherry leaves

Table 2.

Table 3.

Table 4.

Table 5.

Table 6.

Table 7.

Table 8.

from Clarksville Michigan orchard trees on speciﬁed dates in 1993 .......... 44
The effect of mite infestation on Fv/Fm ratio, Fo, Fm and Fv from chlorophyll

ﬂuorescence of sour cherry leaves from orchard trees on selected dates in 1993

........................................................................................ 45
The effect of mite-infestation and cumulative mite'days (CMD) on C02

assimilation (A) and Fv/Fm ratio of fully expanded sour cherry leaves of

container grown trees on specific dates in 1994 ................................. 46

The effect of mites and cumulative mite°days (CMD) on whole tree C02
assimilation, single leaf C02 assimilation, transpiration (E), Stomatal
conductance (gs), water use efﬁciency (W UE), photosynthetic yield, and
Fv/Fm ratio of expanded sour cherry leaves of container—grown trees on 2
August 1994 ......................................................................... 47

The effect of application of 63 ppm terbacil on shoot starch levels on 1 November
1993 and shoot and root starch levels on 14 April 1994 of orchard sour cherry
trees ................................................................................... 48

The effect of miticide application on shoot length increase and chlorophyll of
greenhouse grown sour cherry trees after application .......................... 64

The effect of application of 63 ppm terbacil on sheet extension and total
chlorophyll on selected dates of orchard sour cherry trees in 1993 ........... 69

The effect of application of 63 ppm terbacil on fruit weight, soluble solids and

removal force for orchard sour cherry trees in 1994 ............................ 76

Table 9. Pesticide applications made to peach orchards in 1992 ........................... 87

Table 10.
Table 11.

Table 12.

Table 13.
Table 14.

Table 15.

Table 16.

Table 17.

Table 18.
Table 19.
Table 20.

Pesticide applications made to peach orchards in 1993 ......................... 88
The effect of different levels of chemical pesticide input on pesticide residues of
peach fruit in 1992 .................................................................. 89
The effect of different levels of chemical pesticide input on pesticide residues of
peach fruit in 1993 .................................................................. 90
Data for mite population counts at Clarksville Michigan in 1993 .............. 96

Data for chlorophyll ﬂuorescence measurements for control and mite infested
trees at Clarksville Michigan in 1993 ............................................. 98
The effect of mites on 1993 shoot growth and 1994 ﬂowering and fruit set for
orchard trees infested in 1993 ..................................................... 99
The effect of application of 63 ppm terbacil on 1993 fruit yield, diameter,
weight, soluble solids and removal force; and 1994 return fruit set and yield of
orchard sour cherry trees at Clarksville Michigan .............................. 101
Chlorophyll data from control and mite infested trees at Clarksville Michigan in
1993 ................................................................................. 103
Yield data collected in 1993 and 1994 from orchard sour cherry trees ....... 104
Starch data collected from orchard sour cherry trees in 1993 and 1994 ...... 105

Data collected from container grown control and mite trees in 1994 ......... 107

LIST OF FIGURES

Figure 1. Whole plant photosynthetic C02 assimilation chamber ......................... 40
Figure 2. Calculated cumulative mite-days on control and mite-infested orchard sour
cherry trees on speciﬁed dates during the 1993 season at Clarksville Michigan .
........................................................................................ 41
Figure 3. The effect of spraying 63 ppm terbacil on CO2 assimilation and Fv/Fm ratio of
orchard sour cherry leaves at Clarksville Michigan in 1993 .................. 42
Figure 4. The effect of spraying 63 ppm terbacil on whole tree C02 assimilation of orchard
grown sour cherry trees at Clarksville Michigan in 1993. .................... 43
Figure 5. Single leaf C02 assimilation (ug0m'2¢3'1) for orchard sour cherry trees in 1993
at Clarksville Michigan plotted against (A): mites per leaf, and (B) cumulative
mite-days ............................................................................ 53
Figure 6. Whole tree C02 assimilation (ugom'ZOS'l) for orchard sour cherry trees in 1993
at Clarksville Michigan plotted against (A): mites per leaf, and (B) cumulative
mite°days ............................................................................ 54
Figure 7. Fv/Fm ratio from chlorophyll ﬂuorescence for orchard sour cherry trees in 1993
at Clarksville Michigan plotted against (A): mites per leaf, and (B) cumulative
mite'days ............................................................................ 55
Figure 8. Effect of cumulative mite day populations for orchard sour cherry trees in 1993
at Clarksville Michigan (A) 1994 yield (kg per tree), and (B) cold hardiness
(T 50) of 1993-94 dormant season ............................................... 56

Figure 9. Effect of cumulative mite day populations for orchard sour cherry trees in 1993

Figure 10.

Figure 11.

Figure 12.

Figure 13.

Figure 14.

Figure 15.

Figure 16.

Figure 17.

at Clarksville Michigan on (A) 1993 fall shoot starch concentrations, (B) 1994
spring shoot starch concentrations, and (C) 1994 spring root starch
concentrations ...................................................................... 57
Stem tissue cold hardiness as determined by T(5()) values for controls and
mites-infested trees on the speciﬁed dates in the 1993-94 dormant season at
Clarksville Michigan ................................................................ 60
The effect of miticide application on (A): C02 assimilation (A), (B): Stomatal
conductance (gs), (C): transpiration (E), and (D): Fv/Fm of expanded sour
cherry leaves in East Lansing Michigan greenhouse in 1994 .................. 63
The effect of terbacil concentration on (A): C02 assimilation (A) and (B):
Fv/Fm ratio of expanded leaves of container grown sour cherry trees ....... 67
Stem tissue cold hardiness as determined by T(5()) values for controls and trees
treated with 63 ppm terbacil on the speciﬁed dates in the 1993-94 dormant
season at Clarksville Michigan .................................................... 72
The effect of single and multiple spray applications of 63 ppm terbacil on (A):
C02 assimilation and (B): Fv/Fm ratio of expanded sour cherry leaves from
orchard trees in 1994 ............................................................... 75
Daily minimum and maximum temperatures for 1993-1994 dormant period at
Clarksville Michigan .............................................................. 113.
Air ﬂow generating fan and inlet setup for whole plant photosynthetic C02
assimilation chambers ............................................................. 116
Schematic drawing of whole plant photosynthetic C02 assimilation chambers

................................................................................... 119

ABBREVIATIONS

A ....................................................... carbon dioxide assimilation
CMD .......................................................... cumulative mitesdays
Conventional ............................... conventional level of chemical input
EPA ............................................ Environmental Protection Agency
Fv/ Fm ...................... variable ﬂuorescence to maximal ﬂuorescence ratio
GC ............................................................. gas chromatography
gm .......................................................... mesophyll conductance
gs .............................................................. Stomatal conductance
E ......................................................................... transpiration
IPM ................................................... integrated pest management
Low .................................................... low level of chemical input
Moderate ......................................... moderate level of chemical input
MPL .................................................................... mites per leaf
Mylar .......................... polyvinylidene coated polyethylene terephthalate

0mite.... propargite [2-(p—tert-buty1phenoxy) cyclohexyl 2-propynyl sulﬁte]
Vendex... fenbutin-oxide[Hexakis(2-methyl-2-phenylpropyl)-distannoxane]
WUE ........................................................... water use efﬁciency

X-77 ...... alkylarylpolyoxyethylene, gycol, fatty acids and isopropanol mix

xiv

Guidance Committee:

The journal—article format was adopted for this dissertation in accordance with
departmental and university requirements. The manuscript was prepared for publication in
the Journal of the American Society for Horticultural Science. The manuscript in Appendix

H was prepared for publication in HortScience.

Introduction

Insects, mites and diseases present on leaves of fruit crop plants can have a number
of deleterious effects on both vegetative growth and fruit development. These pests
damage the photosynthetic machinery and reduce the potential of carbohydrates produced
by those leaves, and a threshold is ultimately reached at which vegetative and reproductive
growth is reduced. Mites (Acari: Tetranychidae) and other insects feeding on apple (Malus
sp.) leaves reduce photosynthesis (Ferree, et al., 1986; Hall and Ferree, 1975; Proctor et
al., 1982), growth, yield and return fruiting (Hull and Beers, 1990; Beers and Hull, 1987;
Bailey, 1979). Reductions in C02 assimilation as a result of mite-feeding have also been
demonstrated in peach, Prunus persica (Anderson and Mizell, 1987; Mobley and Marini,
1990), almond, Prunus amygdalus (Youngman, et al., 1986), and grape, Vitis (Candolﬁ,
et al., 1992). However, only at very high population levels do mites affect photosynthesis,
vegetative growth and yield of apple (Hull and Beers, 1990; Beers and Hull, 1987), and
there was little effect on yield, fruit quality and vegetative growth of peach (McCleman and
Marini, 1986). Photosynthesis was not affected by feeding of European red mite
(Panonychus ulmi) on grape, although it was reduced by two—spotted spider mite
(Tetranychus urticae) (Candolﬁ, er al., 1993).

Speciﬁc low levels of pest populations that do not affect total yield and fruit quality
can presumably be tolerated in an orchard. The reduction of photosynthesis and the
resulting effects on cropping have not been studied on sour cherry, and therefore,
biological, action and economic thresholds have not been established. Relatively low
numbers of insects or mites feeding on the leaves does not result in lower yield or fruit

quality (Mobley and Marini, 1990). The mechanism of tolerance may be that the tree either

2
alters carbon assimilation primarily toward fruit production at the expense of vegetative

growth and storage or leaves compensate for lost photosynthetic tissues by increasing
photosynthesis in the remaining tissues (Layne and Flore, 1992).

It is not clear how reduced vegetative growth and carbohydrate storage from mite or
insect feeding affects physiological processes such as cold acclimation and hardiness of
tissues and subsequent—year ﬂower initiation and fruit development. These factors are
particularly important with regard to sour cherry as insect and mite populations are often
highest after sour cherry harvest when ﬂower bud development and carbohydrate storage
are occurring. Howell and Stackhouse (197 3) have demonstrated that severe defoliation
caused by cherry leaf spot can reduce subsequent years yield. Therefore, establishing
insect and mite feeding thresholds that do not adversely affect these vegetative and
reproductive processes for sour cherry orchards would be an important tool for orchard
pest management. Also, as carbohydrate demand is lower after sour cherry harvest (Flore,
1994), thresholds for pest damage could be considerably higher than during the period
prior to harvest, and considerably higher than thresholds-established for apples.
Additionally, it would be of value to growers to develop a system that could be applied in
the ﬁeld for early detection of photosynthetic injury and ultimately a photosynthetic
threshold for management of phytophagus pests.

The application of chemical pesticides can be costly and may act on beneﬁcial
organisms as well as the targeted pest. It has also been demonstrated that many insecticides
can reduce photosynthesis (Ayers and Barden, 1975; Andersen, et al., 1986). Therefore,
the use of pesticides can act to inhibit photosynthesis, reduce populations of natural
predators and increase the levels of residues in the fruit. This calls into question the
necessity of some pesticide applications, particularly for mites on sour cherry, in light of
the lack of information on the effects of mite feeding on sour cherry trees.

The objectives of this research were to characterize the effects of mite-feeding on

photosynthetic parameters, starch accumulation, cold hardiness and cropping; study mite

3
population thresholds that have no adverse effect on tree growth; determine the effects of

miticide application on photosynthesis; and characterize the effects of photosynthetic
reduction on these parameters and establish photosynthetic reduction thresholds that have

no adverse effect on tree growth.

LITERATURE CITED

Anderson, PC. and R.F. Mizell, III, 1987. Impact of the peach silver mite, Aculus
cornutus (Acari: Eriophyidae), on leaf gas exchange of 'Flordaking' and 'June

Gold' peach trees. Environ. Entomol. 16:660-663.

Andersen, P.C., R.F. Mizell, W.J. French and J .H. Aldrich, 1986. Effect of multiple
applications of pesticides on leaf gas exchange of peach. HortScience 21(3):508-
510.

Ayers, Jr., J .C. and J.A. Barden, 1975. Net photosynthesis and dark respiration of apple
leaves as affected by pesticides. J. Amer. Soc. Hort. Sci. 100(1):24-28.

Bailey, P. 1979. Effect of late season populations of twospotted mite on yield of peach

trees. J. Econ. Entomol. 72:8-10.

Beers, E.H. and Lil. Hull. 1987. Effect of European red mite (Acari: Tetranychidae)
injury on vegetative growth and ﬂowering of four cultivars of apple. Environ.

Entomol. 16(2):569-574.

4
Candolﬁ, M.P., E.F. Boller, and B. Wermelinger, 1992. Inﬂuence of the twospotted

spider mite, Tetranychus urticae, on the gas exchange of Pinot noir grapevine

leaves. Vitis 31 :205-212.

Candolﬁ, M.P., B. Wermelinger, and ER Boller, 1993. Photosynthesis and
transpiration of "Riesling x Sylvaner" grapevine leaves as affected by European red
mite (Panonychus ulmi Koch) (Acari, Tetranychidae) feeding. J. Appl. Ent.

1 15:233-239.

Ferree, D.C., F.R. Hall, and M.A. Ellis, 1986. Inﬂuence of mites and diseases on net
photosynthesis of apple leaves. Regulation of Photosynthesis of Fruit Trees

Symposium Proceedings, NY State Agr. Exp. Sta., Geneva, NY, p. 56—62.

Flore, J .A., 1994. Stone Fruit. in Schaffer, B. and RC. Andersen (ed.), Handbook of
Environmental Physiology of Fruit Creps, Volume 1: Temperate Crops, CRC
Press, Inc., Boca Raton, pp.233-270.

Hall, F. R. and D. C. Ferree, 1975. Inﬂuence of twospotted spider mite populations on

photosynthesis of apple leaves. J. Econ. Entomol. 68(4):517-520.

Howell, GS and 8.8. Stackhouse, 1973. The effect of defoliation time on acclimation

and dehardening in tart cherry. J. Amer. Soc. Hort. Sci. 98(2): 132-136.

Hull, LA. and EH. Beers, 1990. Validation of injury thresholds for European red mite

(Acari: Tetranychidae) on 'Yorking' and 'Delicious' apple. J. Econ. Entomol.
83(5): 2026-2031.

5
Layne, DR. and J.A. Flore, 1992. Photosynthetic compensation to partial leaf area

reduction in sour cherry. J. Amer. Soc. Hort. Sci. 117(2):279-286.

McClernan, W.A. and RP. Marini, 1986. European red. mite on yield, fruit quality, and
growth of peach trees. HortScience 21(2):244-246.

Mobley, K. N. and R. P. Marini, 1990. Gas exchange characteristics of apple and peach

leaves infested by European red mite and twospotted Spider mite. J. Amer. Soc.
Hort. Sci. 115:757—761.

Proctor, J.T.A., J.M. Bodnar, W.J. Blackburn and R.L. Watson, 1982. Analysis of the
effects of the spotted tentiforrn leaf minor (Phyllonorycter blancardella) on the
photosynthetic characteristics of apple leaves. Can. J. Bot. 60:27 34-2740.

Youngman, R.R., V.P Jones, S.C. Welter, and M.M. Barnes, 1986. Comparison of
feeding damage caused by four Tetranychid mite species on gas-exchange rates of

almond leaves. Environ. Entomol. 15:190-193.

THE INFLUENCE OF MITES AND A PHOTOSYNTHETIC INHIBITOR
(TERBACE) ON PHOTOSYNTHESIS AND YIELD OF SOUR CHERRY
(PRUNUS CERASUS L. 'MONTMORENCY')

Mark A. Hubbard1 and James A. Flore2
Department of Horticulture, Michigan State University, East Lansing, MI 48824-1325

Received for publication

1Graduate Research Assistant

2Professor

Cellular and Whole Plant Physiology

The Inﬂuence of Mites and a Photosynthetic Inhibitor (T erbacil) on Photosynthesis,

Carbon Accumulation and Yield of Sour Cherry (Prunus cerasus L. 'Montrnorency')
Additional index words. IPM, pest management, miticides, chlorophyll ﬂuorescence.

Abbreviations: Carbon dioxide assimilation, A; transpiration, E; stomatal conductance, gs;
mesophyll conductance, gm; water use efﬁciency, WUE;.variable ﬂuorescence to maximal
ﬂuorescence ratio, Fv/Fm; cumulative mite-days, CMD; mites per leaf, MPL; mite—infested
trees, MIT; Omite: Propargite [2-(p-tert-butylphenoxy) cyclohexyl 2-propynyl sulﬁte];
Mylar: polyvinylidene coated polyethylene terephthalate; Vendex, Fenbutin-
oxide[Hexakis(2-methyl-Z-phenylpropyl)-distannoxane]; Omite, Propargite [2-(p-tert-
butylphenoxy) cyclohexyl 2-propynyl sulﬁte]; X-77, alkylarylpolyoxyethylene, gycol, free
fatty acids, isopropanol proprietary mix (Valent).

Abstract. Orchard and container—grown sour cherry (Prunus cerasus L. 'Montrnorency')
trees were infested with two—spotted spider mites (Tetranychus urticae) to study mite—
feeding effects on photosynthetic parameters, cold hardiness, starch accumulation and
reproduction effort (ﬂowering and yield). In the ﬁeld, mite populations reached 1764
average cumulative mite-days (CMD) for mite-infested trees (MIT) and 1093 CMD for
controls. Chlorophyll was reduced in MIT compared to controls late in the growing
season, yet there was no reduction during the season in measured photosynthetic
parameters, stem cambium cold hardiness or shoot starch levels. Additionally, miticides
used commercially in Michigan tart cherry orchards were determined to have no effect on
photosynthetic parameters. There was an increase in spring root starch levels of MIT the

spring following infestation, but no effect on return fruit set or retum yield. In the

8
container grown trees, average CMD reached 917 for MIT and 90 for controls. Reductions

were observed in single leaf A, whole tree A and chlorophyll ﬂuorescence. This suggests
that mite populations sustained in the orchard in 1993 were not high enough to reduce
subsequent year yield, and therefore, control measures were of no economic benefit. The
ﬁeld trees demonstrated an ability to buffer the effects of mite feeding and maintain
reproductive capacity. Reduction in photosynthesis does occur, as in the container study,
and that reduction in photosynthesis could ultimately affect yield and hardiness. In order to
study photosynthetic reduction thresholds that affect tree hardiness and yield, trees were
treated with 50-63 ppm terbacil to limit photosynthesis for speciﬁc periods of time.
Terbacil was applied to orchard trees at 4 physiological stages: fruit growth stages I and II,
fruit harvest, and prior to leaf fall; at each date, A was reduced up to 60% within 24 h and
showed 100% recovery within 7 -10 days. Reductions were also observed in whole tree A
and chlorophyll ﬂuorescence, but not in chlorophyll, shoot cold hardiness, fruit yield or
fruit quality. Root and shoot starch levels did vary slightly dependent on the timing of

terbacil application but there was no effect on return yield.

The infestation of fruit orchards with mite species has increased signiﬁcantly in
recent years with the increased use of pyrethroid insecticides to control insect pests as those
insecticides signiﬁcantly reduce the populations of predatory mites that feed on
phytophagus mites in the orchard. The result has been an increase in damage to the orchard
trees as a result of higher populations of two—spotted mite and European red mite
(Panonychus ulmi). Growers are currently using a number of pesticide applications aimed
speciﬁcally at phytophagus mites. We estimated that one miticide application to 80% of
Michigan sour chenies could cost growers between $2.4 and 3.0 million and result in the

use of 18,000 to 36,000 pounds of active ingredient, depending on the compound chosen.

9
The effects of phytophagus mites on fruit trees such as apple and peach have been

well documented (Avery and Briggs, 1968; Bailey, 1979; Beers and Hull, 1987; Lienk, et
al., 1956; Mobley and Marini, 1990; Zwick, et al., 1976) while research on the effects of
phytophagus mite feeding on sour cherry has been limited. Jones (1990a) conducted a
study on predator: prey relationships in sour cherry and later considered effects of
pesticide applications on mite activity (Jones, 1990b). There have been no reports on the
effects of phytophagus mite feeding on photosynthetic parameters and yield components of
sour cherry. As damage to sour cherry leaves is visibly similar and proximate in time to
apple leaf damage, the abundant research conducted on mite—feeding and apple tree
photosynthesis has been extrapolated to management of sour cherries. Unlike apple and
peach, sour cherry orchards experience high mite populations and damage primarily after
the current season's crop has been harvested. The current season‘s yield of peach and
apple can be adversely affected by mid- and late-summer mite populations (Ames, et al.,
1984; Bailey, 1979; McCleman and Marini, 1986) as can pear (Westigard, et al., 1986)
and grape (Welter, et al., 1989). Additionally, this late season mite feeding has been
shown to reduce apple ﬂowering and yield in subsequent years (Beers and Hull, 1987;
Beers and Hull, 1990). Utilizing these factors, thresholds have been developed for mite
populations in apple and grape production in particular (Hull and Beers, 1990; Candolﬁ, er
al., 1993).

Several distinct thresholds for management purposes can be developed. The
biological threshold is that population of mites that causes a biological response, such as a
reduction in A, and is therefore reached at relatively low population of mites. The action
threshold is the population level that dictates an action on part of the orchard manger to
prevent adverse effects on the total crop. It presumably would occur at mite populations at
least as high as those for the biological threshold. Ultimately, the economic threshold is

reached, which is the mite population that results in an economic loss to the orchard

10
production system. Ideally, the biological threshold can be tolerated but is used as a guide

to predict the action threshold which is then used to avoid reaching the economic threshold.

The question arises as to whether sour cherry can be affected by late season mite—
feeding to the extent that ﬂowering and yield are reduced in the subsequent year as can
occur with apple (Beers and Hull, 1990). If yield is adversely affected, are the similarities
with apple trees sufﬁcient to warrant using apple threshold levels? Sour cherry growers are
currently applying miticides at a substantial cost, and as no research has been conducted on
thresholds for subsequent-year fruiting of sour cherry, the assumption is that sour cherries
are damaged by late season mite—feeding to the same extent as apples.

In addition to damage caused by the pests themselves, some pesticides have been
shown to limit photosynthesis of sour cherry (Murphy, 1937) as well as apple and peach
(Ayers and Barden, 1975; Andersen, et al., 1986). Repeated applications of pesticides can
also reduce photosynthesis (Ferree and Hall, 1978). It is unclear if miticide chemicals have
an adverse effect on tree photosynthesis and growth apart from any beneﬁcial effect on pest
populations.

Photosynthetic reduction in the orchard can occur as a result of any number of
abiotic or biotic stresses apart from mite damage. Plants can compensate for moderate
degrees of photosynthetic loss or for partial defoliation by an increase in photosynthetic rate
of the undamaged portions of the leaves. Layne and Flore (1992) suggested that a loss of
up to 20% of sour cherry leaf area resulted in no net reduction of photosynthesis. Similar
results have been shown with other plants (Bassman and Dickmann, 1982; Proctor, et al.,
1982; Satoh, et al., 1977). Increased damage, particularly from insects or mites, will
ultimately reduce photosynthesis to a point that the leaf or tree can no longer compensate
and net photosynthesis will decline. Reduced photosynthesis can have a variety of effects
on the tree including reduced growth and yields ( Bailey, 1979; Hull and Beers, 1990;
Beers and Hull, 1987), and reduced carbohydrate levels and cold hardiness (Flore, et al.,

1983). Tissue hardiness has been correlated with starch concentration in peach (Flore er

1 1
al., 1987; Lasheen et al., 1970), but low starch concentration does not necessarily result in

lower hardiness. The same factors that promote starch concentrations may also promote
cold hardiness (Flore and Howell, 1987). Of obvious importance is a reduction in yield,
which can also occur in years subsequent to the season in which the photosynthetic
reduction actually occurred. N 0 studies have considered the amount of photosynthetic
reduction that can occur in sour chen'y before yield or tree hardiness is affected, or how the
timing of any photosynthetic reduction could effect yield and hardiness.

Terbacil is the active ingredient of a commercial herbicide that inhibits
photosynthesis by blocking the ﬂow of electrons in photosystem II (Gardiner, 1981). At
the labeled concentration of 2-4 kg-ha'l of concentrated spray volume (7676-15353 ppm
terbacil), terbacil is an effective herbicide used in production of many fruit crops, but has
been used to limit photosynthesis in apples for the purpose of fruit thinning (Byers et al.
1990). We hypothesized that at these signiﬁcantly reduced concentrations, 50—200 ppm,
terbacil would inhibit photosynthesis of sour cherry for a limited time after which the trees
would recover to their full photosynthetic capacity. This temporary inhibition could mimic
the effect of mite, insect or disease damage on plant photosynthesis, after which, control
measures relieve the pest-induced inhibition and the tree typically recovers to full
photosynthetic capacity.

A series of experiments were undertaken to address those unknowns and obtain
information useful in sour cherry orchard management. A study was initiated to determine
sour cherry leaf mite populations that reduced chlorophyll levels, photosynthesis and
chlorophyll ﬂuorescence. As any reduction in these parameters would not affect the current
season's crop, consideration was to be given to the effects on hardiness and cropping in the
following season. Therefore, measurements were made on the effect of mite populations
on stem cambium cold hardiness, tissue starch levels, return fruiting characteristics and
return yield. 0f additional interest was the use of chlorophyll ﬂuorescence measurements

as an indicator of plant stress due to mite damage. Chlorophyll ﬂuorescence, the Fv/Fm

12
ratio in particular, is a sensitive indicator of the physiological status of the photosynthetic

apparatus (Krause and Weis, 1991). As continuous mite-feeding increasingly damages the
photosynthetic apparatus of the leaf (Avery and Briggs, 1968), the ﬂuorescence parameters
should exhibit corresponding alterations. It was hypothesized that periodic chlorophyll
ﬂuorescence measurements would detect the mite damage to the leaf in advance of other
measurable parameters. If such were the case, this technique would allow growers a
method of early detection of mite injury - an early biological threshold — and become a
valuable tool in orchard management in predicting action thresholds. A ﬁnal objective was
to reduce photosynthesis in a sour cherry orchard via application of a dilute terbacil
concentration to study how the resulting photosynthetic reductions affect yield and tree

hardiness.
Materials and Methods

Plant Materials. A sour cherry orchard established in 1982 at Clarksville Michigan
was utilized for the mite—feeding and terbacil application experiments . The container
studies utilized one-year-old sour cherry trees obtained from Newark Nurseries (Hartford,
MD that were potted in 12 L pots with a ﬁeld soil mix, consisting of 7: 1 : l,
soilzsandzorganic matter and the trees pruned to two buds on a single stem. The mite study
trees were grown outdoors under natural conditions in March—August 1994 for 45-60 days
by which time there were ten or more fully expanded leaves before infesting with mites.
The plants used for miticide studies were grown in a greenhouse on the Michigan State
University campus with day temperatures of 25—30°C and night temperatures of 17-21°C
under natural light conditions in March through May 1994. All trees were watered as
needed, and fertilized biweekly with soluble 20—20—20 fertilizer at the rate of 60 g'L'l.

Photosynthesis. All single leaf photosynthesis was measured from the 2 most

recently expanded leaves using portable infrared gas analyzer units (ADC, Hoddesdon,

13
UK, model LCA—2, and RP. Systems, Haverhill, Mass, model CIRAS) and Parkinson

leaf chambers. The units measured and recorded net change in chamber C02, leaf and air

temperatures, ambient and leaf chamber humidiﬁes, humidity increase due to transpiration,

and incident light radiation. Net A; transpiration, E; stomatal conductance, gs; mesophyll
conductance, gm; and water use efﬁciency, WUE were calculated using a BASIC computer
program developed by Moon and Flore (1986), or when using the PP Systems unit, these
parameters were calculated automatically by the unit.

Whole tree photosynthesis was measured with whole tree gas exchange chambers
(Figure 1) that utilized a fan-forced air inlet; C02 concentrations were monitored at the inlet
and the outlet using portable infrared gas analyzer units described above. The 44.9 m3
chambers were transparent Mylar (DuPont) and the seems were sealed with transparent tape
(3M Corp.) Industrial type fans were utilized that introduced ﬂow rates ranging from
2400 to 3600 L-min'l. This ﬂow rate was sufﬁciently high to minimize temperature
increase inside the chamber and maximize C02 differential, approximately 1 complete
exchange of. Flow rate was continuously monitored at the inlet using an anemometer
(Cole-Parmer, Tri—sense model 37000-00) and taking the average of measurements across
the inlet duct. The chambers were comparable to that described by Corelli-Grappadelli and
Magnanini (1993). Smaller chambers were used for whole tree photosynthesis
measurements of container trees, and are described in detail elsewhere (Miller, et al., in
press). Flow rates for the container tree chambers ranged from 36 to 70 L/min.
Assimilation of CO2 was calculated from the differential CO2 concentrations and the air
ﬂow through the chamber based on the calculations of Moon and Flore (1986).
Assimilation was determined on a leaf area basis using an estimation of the whole tree leaf
area from data collected by Teichman and Flore (unpublished) for orchard trees and by
actual leaf area measurements for container trees. Additionally, A was calculated on a trunk

cross sectional area basis.

l4
Chlorophyll Fluorescence. Chlorophyll a ﬂuorescence was measured on the two

most recently expanded leaves using one of two ﬂuorescence measurement systems (P.K.
Morgan Instruments, Inc., Andover, MA, model CF—lOOO; Opti-Sciences, Inc.,
Tewksbury, MA, model 0S-500). Dark adaptation cuvettes were attached to leaves on
trees of each treatment 20-30 min. prior to measurement. The optical cable of the
ﬂuorescence unit was then attached to the cuvette and the leaf irradiated with a pulse of
1000 tunol-m'zsl 680 nm actinic light for 60 s. The following ﬂuorescence parameters
were measured and recorded: initial ﬂuorescence, Fo; maximal ﬂuorescence, Fm; variable
ﬂuorescence, Fv = Fm-Fo; terminal ﬂuorescence, Ft; time to one—half ﬂuorescence
quenching, q; time to ﬂuorescence quenching, t; and calculation of the ratio Fv/Fm. The
Fv/Fm ratio is signiﬁcant in that it is closely correlated with photochemical efﬁciency
(Krause and Weis, 1991). The Opti-Sciences unit also measured "photosynthetic yield,"
which is an indicator of quantum yield of photosynthesis, via pulse modulation and
measurement of secondary maximal and variable ﬂuorescences (Opti—Sciences, Inc.,
1994).

Chlorophyll. Chlorophyll determinations were made by collecting 4 disc samples
from the interveinal lamina proximate to the base of 4 different leaves on comparable
canopy positions from each tree. The discs were collected using a paper hole punch
yielding leaf discs with an area of 0.32 cm2 and immediately placed in 20 ml glass
scintillation vials containing 7 ml of N ,N-dimethylfonnamide and placed on ice. The
samples were stored at +5°C for 48 hours and then the absorbance of the N,N-
dirnethylforrnamide solution measured at 623, 647 and 664 nm relative to a N,N-
dirnethylforrnarnide control. The levels of chlorophyll a, chlorophyll b, protochlorophyll
and total chlorophyll were then calculated using the formulas from Moran (1982). The
chlorophyll levels were converted to ug-cm'2 of leaf tissue.

Carbohydrate Status. Tissue samples for carbohydrate determinations were

collected from the 1993 growing season's shoot and root tissue on November 10, 1993

15
and April 14, 1994. Current season shoots of 50-80 cm and 10g of freshly dug root tissue

were collected from orchard trees and wrapped in polyethylene bags and transported on ice
to the laboratory and then 1 cm sections placed in envelopes and frozen. Samples were
frozen at —80°C and held for 3-4 weeks and then lyophilized and ground (40 mesh) . To
100 mg samples, 3.5 ml of 80% ethanol was added, vortexed and allowed to stand for 30
min. The samples were then centrifuged for 5 min at l879xg and the supernatant collected
for carbohydrate extraction and analysis. The pellet was extracted twice as above,
suspended in 80% ethanol, vortexed, and centrifuged. The pellets from the samples were
then frozen and dried (Speedvac SC200). Two ml of neutralized acetic acid (0.1 M, pH
5.0) was added to the dried pellet, vortexed gently and incubated at 100°C for l h.
Samples were cooled to room temperature for 10 min and then 10011.1 of arnyloglucosidase
solution (0.0166 g'ml'l) was added, the samples gently mixed and incubated at 55°C for 16
h. The samples were then centrifuged for 5 min at 1500xg and 201.11 removed from each
sample and combined with 1 ml double distilled H20. 250 [.11 of these sample solutions
were combined with 2.5 ml of Sigma color reagent solution and incubated to develop color
for 40 min. Absorbance at 440 nm was then measured and the percent starch calculated
according to standards of known concentrations.

Cold Hardiness. Cold hardiness of stem cambium tissues was determined on eight
dates from October to May based on a method described by Flore et a1. (1983). On each
collection date, six 50-80 cm stems of the most recent seasons growth were cut from the
top of the canopy of each tree. Within 3 hours, the stems were prepared for cold hardiness
tested as described below. The basal and apical thirds of each stem were discarded, and the
middle third, 20-25 cm, was cut into 1.5-2.0 cm sections. These stem sections were taped
together, wrapped in wetted gauze, and then rolled inside aluminum foil. Each roll of
aluminum contained one stem section from each treatment, and a 26-gauge copper-
constantan thermocouple inserted into one sample of each roll to monitor tissue

temperature. Sufﬁcient samples were prepared to allow freezing tests at 6 temperatures and

16
a control held continuously at 5°C. Three replicates for each temperature were prepared.

The samples were placed inside a Beckmann freezer modiﬁed for precise temperature
regulation and cold hardiness studies. The temperature was maintained at -1°C for 8-12 h
and then the temperature lowered at the rate of 3°C per hour. Stem temperatures were
monitored and 3 replicates removed when predesignated temperatures were achieved. The
samples were thawed slowly at 5°C for 20-24 h and then the stem sections removed from
the aluminum foil and gauze. The stem samples were then placed in plastic containers and
air bubbled through a small volume of water to maximize humidity for 7-10 days. Each of
the samples was examined and the cambium visually assessed as alive or dead .
Calculations were then made to determine the T50 of the cambium for each treatment using
the Spearman—Karber method (Bittenbender and Howell, 1974). The T50 was calculated

as

T,O =T100 -O.5d+‘—i):—bi
n

where
T50 = temperature at which 50% of the stems are killed,
T100 = temperature at which 100% of the stems are killed,

(1 = temperature interval between treatments

b; = number of dead stems in im temperature

n = number of replicates sampled at each treatment temperature.
Cumulative cold hardiness was calculated as the average T50 value over the dormant period
for each treatment.

Mite—Feeding Studies. Treatments consisted of control trees to which a miticide
was routinely applied and mite-infested trees to which no miticides were applied. The
treatments were assigned in the orchard utilizing a randomized complete block design with
4 blocks. Each plot consisted of four trees in a row with the two center trees used as data
trees. Bean plants infested with two—spotted spider mites were attached to the trees on July

21, 22 and 23. Naturally occurring populations of European red mite were also present

l7
and counted. Fifty leaves from each tree were collected biweekly and brushed to determine

mite populations. Mite-days were used in an effort to express the duration of the mite
population levels and calculated as the average number of mites for two consecutive
counting dates multiplied by the number of days between the counting dates (Sances, et al.,
198 1).

Container trees were infested with two—spotted spider mites by attaching infested
bean leaves to the stems of the trees. The number of MPL on three representative leaves
from each tree was counted weekly. An additional application of infested bean leaves was
required after it was determined that there were 0 mites in the trees on 13 July. Trees were
blocked by height and a randomized complete block design was used with 11 replications.

Vegetative and Reproductive Growth. In the orchard study, 5 one-year-old shoots
were randomly selected from each treahnent tree and initial measurements of stem
diameter, stem length, number of leaves, fruit diameter, fruit weight and fruit number made
on June 3, 1993. The fruit weights were determined by harvesting 30 developing fruit.
Fruit diameter, weight and number were measured again on July 22 just prior to harvest by
collecting 30 fruit. Stem diameter, stem length and number of leaves were measured again
on September 29, 1993.

Miticide Study. After 8 weeks of growth, container trees were blocked according
to shoot length and treatments were randomly assigned. The initial greenhouse experiment
utilized 6 repetitions and a subsequent experiment utilized 4 repetitions. The treatments,
applied by dipping leaves into 1L of solution, were a water control containing 1 ppm
proprietary surfactant: alkylarylpolyoxyethylene, gycol, free fatty acids and isopropanol
(X-77), Omite applied at the labeled rate of 6 pounds per 400 gallons (1.8 g0L'1) with 1
ppm X-77, and Vendex applied at the labeled rate of 8 ﬂuid ounces per 100 gallons (0.6
ml-L°1) with 1 ppm X-77. All experiments were designed and analyzed as randomized

complete blocks. Treatment differences were considered signiﬁcant at P2005 using

analysis of variance. Photosynthetic parameters were measured at 0,1,3,5,7 and 10 days

18
after treatment. Shoot length was measured on the day prior to treatment and 30 days after

treatment. Chlorophyll ﬂuorescence parameters were measured at 0,1,3,5,7 and 14 days
after treatment. Chlorophyll levels were measured 15 days after ueatments were applied.

Terbacil Studies. A preliminary experiment was undertaken to determine how
different rates of terbacil herbicide reduced photosynthesis in sour cherry. A sour cherry
(Prunus cerasus L. 'Montrnorency') orchard established in 1982 in East Lansing Michigan
was utilized for the concentration study. The treatments were randomly assigned to a
single leaves of orchard trees in a randomized complete block design with four repetitions.
The experimental design for the greenhouse concentration study was 4 repetitions of each
treatment. In both the orchard and greenhouse experiments, single leaves were dipped in 1
L solutions of 0, 25, 50, 100, 200, 400 and 800 ppm terbacil each containing 1 ppm X-77.
Chlorophyll ﬂuorescence and A were monitored at days 0, 1, 2, 3, 4, 6, 7 and 14 after
treatment.

Four different times of terbacil application and a control were randomly assigned in
the orchard in a randomized complete block design utilizing 4 blocks. Each experimental
unit consisted of a four tree plot row, with the two center trees used as a data trees. Sixty-
three ppm terbacil with 1 ppm X-77 was sprayed to drip, approximately 20L per tree, on
the designated plots on the following dates in 1993: June 1 coinciding with stage I of
cherry fruit development, June 15 coinciding with pit hardening or stage II, August 4 at
harvest, and September 17, 6 weeks prior to leaf fall. The terbacil was applied using a
handgun sprayer at mid—moming of each date.

Vegetative and Reproductive Growth. In the orchard study, 5 one-year-old shoots
were randomly selected from each treatment tree and initial measurements made on June 3,
1993 of stem diameter, stem length, number of leaves, fruit diameter, fruit weight and fruit
number. The fruit weights were determined by harvesting 10 developing fruit. Fruit
diameter, weight and number were measured again on July 22 just prior to harvest. Stem

diameter, stem length and number of leaves were measured again on September 29, 1993.

19
Statistical Protocol. All experiments were a randomized complete block design and

statistical signiﬁcance was determined using analysis of variance and F test. Treatment
effects were considered signiﬁcant at the 5% level or the 10% level where early detection of
thresholds were sought.

Results

Mite—Feeding Studies. Orchard Study. Mite Populations. The average CMD in
the orchard trees through the 1993 season are shown in Figure 2A. The CMD averages
were similar in the two treatments until a Carzol miticide spray was applied to controls on 2
August which resulted in higher mite populations in MIT relative to controls. Mite
numbers then ﬂuctuated slightly with the treatment trees consistently having more mites
than the miticide—treated controls. By the end of the 1993 season the average CMD was
1764 for the treatment trees and 1093 for controls, an increase of 61%.

Gas Exchange and Fluorescence. Single leaf A for 4 dates during the 1993
growing season along with corresponding CMD are given in Table 1. The treatment trees
had increasing mite populations over these dates, but no Change in single leaf A was
observed. E and gs were also not affected by mite—feeding: on 19 August, E was 2.38
mmol0m'203'1 for control trees and 2.28 mmolom'ZOS‘1 for MIT, and gs was 148 mmol-m'
2’8'1 for control trees and 140 mmolem'203‘l for MIT. Likewise, chlorophyll ﬂuorescence
was not altered by the treatments on 5 dates, speciﬁcally the Fv/Fm ratios (Table 2). The
Fv/Fm ratio declined slightly over the growing season but this can be attributed to the aging
of the leaves. Measurements of whole tree A taken on 17 August, 8.5 mmol-m‘Q'S'l for
controls and 7.4 mmol-m'ZOS'1 for MIT, were not different among the two treatments. It
was hypothesized that a regression of A or chlorophyll ﬂuorescence against the mite

population levels would give a better indication of mite—feeding effects. There was no

20
signiﬁcant conelation for either tree or leaf A or ﬂuorescence when compared to MPL or

CMD (Appendix A).

Chlorophyll. Total chlorophyll levels in the MIT were not signiﬁcantly different
relative to control trees with increasing mite-days. 0n 8 October total chlorophyll for MIT
was 59.8 and 64.0 p.g°cm'2 for controls. The MIT did have reduced levels of chlorophyll a
on 23 September, 46.8 rtg°cm'2 for MIT and 50.6 ttg°cni'2 for controls, but on 8 October
the MIT chlorophyll a had recovered to the level of controls. 0n 8 October, the level of
chlorophyll b was reduced in the MIT, 14.3 rrg°cm'2 relative to 15.3 rtg'cm'2 for controls.
There were no differences on any date for the ratio of chlorophyll a to chlorophyll b.

Vegetative and Reproductive Growth. There were no treatrrrent differences for
stem diameter, stem length, and leaf number for the 1993 growing season, nor in
subsequent year (1994) ﬂowering, fruit set and yield. Stem length averaged 22.3 cm for
controls and 22.2 for MIT. Return yield was 0.174 kg'cm'2 of trunk cross section for
controls and 0.149 kg°cm'2 for MIT. Additionally, there were no treatment differences in
the subsequent year for fruit diameter or fnrit weight: fruit diameter averaged 20 mm for
controls and 19 mm for mites, whereas fruit weight averaged 4.1g for both controls and
MIT.

Carbohydrate Status. Starch levels measured from shoot tissues collected on 1
November 1993 and on 14 April 1994 showed no effects of the mite treatment. MIT
shoots had 7.8% and 13.8% starch on 1 November and 14 April, respectively, and
controls had 7.6% and 10.1% starch on 1 November and 14 April, respectively. Starch
levels in root tissues collected on 14 April 1994 were slightly higher (signiﬁcant at p26%)
in MIT, 7.0% compared to 5.8% for controls.

Cold Hardiness. Measurements of one-year old stem cambium cold hardiness of
over the dormant period subsequent to the mite-feeding are shown in Appendix B. Actual
cold hardiness depends on the environmental conditions the trees are exposed to in the

orchard and the T50 values determined on each date were a reﬂection of actual minimum

21
temperatures recorded at the orchard. The mite populations had no effect the T50 values

relative to control.

Container Study. Mite Populations. Mite populations were markedly different in
the 2 treatments due to greater control of MPL on container grown trees. After initial
establishment of a high number of MPL on the treatmenttrees, there was a decline to 0
MPL after which a second establishment was required and mite numbers then remained
stable. The number of CMD in the mite treatment increased gradually over the season to a
ﬁnal maximum of 917 (Figure ZB), 10 times the number for control trees .

Gas Exchange and Fluorescence. Single leaf A for 4 dates over the season are
shown in Table 3 along with the conesponding CMD. As CMD increased, there was no
difference in A until 2 August when the mite treatment trees had accumulated 900
mite'days. All photosynthetic data measured on 2 August is given in Table 4. Whole tree
A, single leaf A, photosynthetic yield from chlorophyll ﬂuorescence, and WUE all reﬂect
an effect of mite—feeding on the trees. However, Fv/Fm ratio from chlorophyll
ﬂuorescence, E and gs do not reﬂect any effects of mite feeding.

Effects of Miticides. The application of the two miticides had no measurable effect
on any of the photosynthetic parameters measured when compared to control or to each
other. Assimilation levels remained constant over the period of measurement and there was
no effect of the miticides on A, gs, E, or Fv/Fm ratios (Appendix C) compared to the
water-treated controls. As Fv/Fm ratio is an indicator of the physiological status of the
photosynthetic apparatus, there is no indication of any adverse effects of the miticide
applications.

There was no reduction in shoot growth, 10.9 cm for controls, 11.8 cm for Omite
and 11.8 cm for Vendex, or total chlorophyll levels, 10.9 ug'cm'2 for controls, 11.8
ug'cm'2 for Omite and 11.8 rrg°cm'2 for Vendex, as a result of miticide applications.

The absence of alteration in shoot growth could be an indication that miticide—

treated plants were not being affected by the chemicals. However, as photosynthetic

22
compensation in response to injury has been demonstrated in cherry (Layne and Flore,

1992), apple (Flore and Irwin, 1983), and other plants (Hodgkinson, 1974; Poston, et al.,
197 6), shoot growth could remain unchanged as the tree compensates for any possible
damage from the nriticide. The miticide application resulted in no decrease in A and,
therefore, no apparent photosynthetic damage. Also, there was no increase in A, and thus
no photosynthetic compensation. The observation that there was no effect of the miticide
chemicals on photosynthesis or the photosynthetic apparatus is conﬁrmed by the
chlorophyll ﬂuorescence measurements. The Fv/Fm ratios remained constant and there
was no change relative to control treatment. The E and gs levels varied with the exact
environmental conditions of the greenhouse at the time of measurement, but there was no
increase or decrease relative to control. The miticide study was carried out under
greenhouse conditions in which the leaf surface tends to have less waxes and therefore
would be more susceptible to injurious chemical applications, whereas in the orchard the
leaf has a much thicker cuticle. Thus, the effect of Omite and Vendex under orchard
conditions would be expected to be less than any effects observed under the greenhouse
conditions. As mite infestation and miticide applications occur late in the growing season,
there would be no effect on the current season's fruit growth and yield, and presumably no
effect on fruit growth and yield of the subsequent season.

Terbacil Studies. The results of the preliminary experiment to determine the effect
of different terbacil concentrations on tart cheny photosynthesis are given in Appendix D.
The Fv/Fm ratio from chlorophyll ﬂuorescence was the best indicator of photosynthetic
injury among the chlorophyll ﬂuorescence parameters in this study. A of the terbacil-
treated trees was initially reduced more than Fv/Fm, but A recovered more quickly than the
Fv/Fm ratio. The 50 ppm terbacil—treated trees were reduced to 32% A and 43% Fv/Fm of
controls one day after treaunent, and both parameters returned to the level of the controls by
7 days after treatment. The 100 ppm terbacil treated trees were reduced to 5% A and 34%

Fv/Fm ratio, but by day 7 the A was 85% of controls while Fv/Fm was only 72%. Higher

23
terbacil concentrations exhibited increasing reductions and longer periods of recovery. The

400 and 800 ppm terbacil—treated trees exhibited visual symptoms of phytotoxicity and did
not recover full photosynthetic capacity.

The A and Fv/Fm ratios from the orchard trees for each of the spray dates is given
in Figure 3. Again, A was initially reduced more than the Fv/Fm ratio but recovered
quicker. Notably, the Fv/Fm for the spray on 23 September (Figure 3D) did not
completely recover to the level of control trees. Whole tree photosynthesis for trees
sprayed at stage II of fruit development is given in Figure 4. The reduction in whole tree A
and recovery essentially mirrored single leaf A (Figure 3B).

Yield and fnrit quality were not adversely affected from the photosynthetic
reduction caused by the terbacil treatment (Table 2). Similarly, terbacil application and the
resulting photosynthetic reduction did not reduce yield in 1994, the season following
treatment. Fruit set was increased as a result of terbacil application at stage II of fruit
development, but this increased fruit set did not lead to any increase in yield.

There was no effect of the terbacil applications on shoot extension or total
chlorophyll levels (appendix E). Chlorophyll a and b levels ﬂuctuated slightly over the
season but this was not considered signiﬁcant.

Percent starch levels were altered by application of terbacil at the different stages
(Table 5). Starch samples from shoot tissue collected on 1 November 1993 showed
reduced levels for terbacil application at stage I of fruit development only, Terbacil #1. The
following spring, shoot starch levels were no different in Terbacil #1 than the control, yet
starch levels were increased for the other terbacil application dates. Conversely, root starch
levels were decreased for Terbacil #2 (stage II) and Terbacil #3 (harvest) compared to
controls, but not affected for Terbacil #1 and Terbacil #4.

Shoot cambium cold hardiness was not affected at any time during the dormant
season by terbacil application at any of the spray dates (Appendix F). Also, the average

cold hardiness for the entire dormant season was also not affected by terbacil application.

24

Discussion

Mite—Feeding Studies. As there was no reduction in any photosynthetic
parameters, but there was a reduction in chlorophyll levels, one may conclude that the
orchard tree leaves increased their photosynthetic production of remaining chlorophyll to
compensate for the loss of chlorophyll. This photosynthetic compensation for leaf damage
has been described in sour cherry (Layne and Flore, 1992) as well as other tree species
(Bassman and Dickmann, 1982; Proctor, et al., 1982; Satoh, et al., 1977). We were
unable to detect a measurable increase. The chlorophyll loss was slight and therefore any
compensation would have been small and difﬁcult to detect even with careful and precise
measurements. The trees were able to maintain full photosynthetic capacity even under the
stress of continually increasing mite damage. Another possibility may be that the high mite
populations in the control trees reduced A photosynthesis in those trees comparable to the
MIT.

The higher mite populations and reduction in chlorophyll levels in MIT had no
effect on fall starch levels (Table 6) in the shoots and no effect on shoot cold hardiness
(Table 7). This suggests that either any photosynthetic compensation was sufﬁcient, or
mite damage was insufﬁcient, to allow MIT shoots to maintain starch levels comparable to
controls levels sufﬁcient to maintain shoot cold hardiness. In the spring of the subsequent
year, shoot starch levels were again not affected in MIT relative to controls, however, root
starch levels were increased over controls. This increase could be due to a number of
factors. The MIT could be maintaining high root starch levels at the expense of
carbohydrate transport. That was not the case as there was no alteration in the shoot starch
levels. The photosynthetic compensation of MIT over the season could be high enough to

result in increased starch levels in the plant resulting in high levels transported and stored in

25
the roots. In effect, an over-compensation in response to the mite-feeding and subsequent

reduction in chlorophyll. If this were the case, one would expect a measurable increase in
photosynthesis, but no such increase was observed. Layne and Flore (1992) have
suggested that photosynthetic compensation is most likely due to enhancement of both
carboxylation efﬁciency and ribulose, l-5-bisphosphate regeneration. This enhancement
may account for sufﬁcient starch being stored in the roots over the winter and present in the
spring.

In the container study, mite populations were more easily managed due to the
smaller tree size and the smaller number of total mites involved. The mite-free control trees
were maintained consistently at less than 5 MPL (Figure 4) and often they had 0 MPL.
Additionally, mites populations could be maintained in MIT by addition of mites added as
needed. Consequently, MIT accumulated ten-fold the number of mites as the controls.

The large difference in CMD at the last measurement date in the container study
coincided with signiﬁcant reductions in several photosynthetic parameters. Reductions
were observed in single leaf A, whole tree A, photosynthetic yield from chlorophyll
ﬂuorescence, and WUE (Table 4) when MIT had 900+ CMD. N o reductions were evident
14 days earlier when there was again a ten-fold difference in CMD between the two
treatments, but MIT had only 700 CMD. Thus, during the time period when mite
populations went from 700 to 915 CMD, MIT reached the photosynthetic threshold for
mite-feeding and damage. Effective control of mite populations below that photosynthetic
threshold might have resulted in no decrease in any of the parameters. As these trees were
relatively small and grown in containers, it is not known how that reduction might effect
yield, or the yield threshold for mite damage. That photosynthetic threshold occuned after
the 1994 harvest date and any effects on yield would not be observed until the following
year. As the orchard trees tolerated mite damage with no effect on photosynthesis,
similarly, trees could tolerate some postharvest photosynthetic inhibition with no effect on

yield or tree hardiness.

26
In the orchard study, there was no decrease in photosynthesis, tree hardiness, and

more importantly subsequent year's yield as a result of high mite populations, which
suggests that control measures were unwarranted in 1993 at this orchard. There could be
an effect of the measured mite population on photosynthetic parameters of controls, which
accumulated 1093 CMD. If one were able to compare the controls to truly mite-free trees,
there may indeed be an effect on photosynthesis as the controls in our study may have been
above a biological threshold. Since completely mite—free trees cannot be achieved, and
when comparing mite control with the absence of mite control, there was no beneﬁt of the
miticide applications. Much of the high CMD in controls was due to high numbers of MPL
prior to any miticide application, suggesting that an earlier application of miticide would
further reduce mite populations. Many miticides are not labeled for preharvest applications,
and as demonstrated in this study, the miticide applications that were made did not reduce
mite populations enough to enhance tree productivity. The container study involved trees
that were much closer to being mite free and a photosynthetic threshold was reached at only
900 CMD, much lower than the 1093 CMD in the orchard control trees. That difference is
most likely due to differences in tree size, age, and vigor that create an ability in the orchard
trees to buffer stresses such as the mite feeding.

The 1764 CMD seen in the ﬁeld MIT fall short of a photosynthetic threshold
(biological threshold) as measured in this study. The photosynthetic threshold, and other
biological thresholds are important for determining an action threshold and ultimately the
economic threshold. As we detected no reduction in photosynthesis and no reduction in
yield in either 1993 or 1994, it is unclear where the economic threshold lies, and thus
where the action threshold should be. Of particular interest to growers is where the
economic threshold lies with respect to the photosynthetic or other measurable biological
thresholds. One possibility is that a particular biological threshold cOincides with the action
economic thresholds, which would require the grower to employ pest controls actions as

soon as the biological threshold is reached. Another possibility, and more likely, is that the

27
action and economic thresholds come at a point beyond the biological threshold and

therefore are difﬁcult to determine. This study has demonstrated that the economic
threshold is not reached prior to measurable losses in photosynthetic activity. Yield
reduction likely occurs only after photosynthesis has been reduced by a speciﬁc level or for
a speciﬁc period of time. The grower must then identify the period between the
photosynthetic threshold and the economic threshold and utilize it for control measures.
With respect to current season's yield, the mite damage in the year of this study occurred
after harvest. The only effect possible on cunent seasons yield would be if mite
populations were high enough early in the season to reduce the leaf to fruit ratio prior to
harvest or reduce A signiﬁcantly during the ﬁnal stage of fruit growth. This is rarely the
casein Michigan sour cherry orchards. Howell, et a1. (1973) showed that signiﬁcant levels
of defoliation and photosynthetic reductions are required to result in fruiting losses in the
subsequent year and thus reach the economic threshold. Also, the detrimental effect of mite
feeding may take a number of years before it results in an measurable effect, particularly
with regard to yield and hardiness. As we did not reach a biological threshold in the ﬁeld
mite studies, we attempted to identify an economic threshold using a herbicide for
photosynthetic reduction.

Direct comparisons of the effects of CMD between the orchard and the container
studies are difﬁcult. While the rateof increase in cumulative mite days over time was
comparable for the orchard and the container studies, there are numerous, obvious
physiological and environmental differences that prevent direct comparisons. Of particular
importance is that the orchard trees are older and more vigorous than the container trees and
therefore have an increased buffering capacity to mite damage. Therefore, the orchard trees
may have a larger overall photosynthetic compensation effect, another mechanism of
increased tolerance to mite damage. Mobley and Marini (1990) have also demonstrated a
greater tolerance of orchard peach trees to mite-feeding compared to greenhouse-grown

container trees. Additionally, the tolerance of trees to mite-feeding can be different

28
depending on the exact timing of mite damage (Candolﬁ, et al., 1992), and thus CMD

numbers can result in different thresholds depending on the time of injury.

An additional objective of this study was to determine if chlorophyll ﬂuorescence,
relatively easy to measure, could be used as an early indicator of mite damage to the
photosynthetic apparatus, prior to any visual damage. There were no reductions in
ﬂuorescence in the orchard study even though mite damage was detected via chlorophyll
levels and damage was evident visually. Quantum yield of photosynthesis as determined
with the modulating ﬂuorescence system was reduced in the container study but coincided
with detected reductions in A. Other ﬂuorescence parameters, in particular the Fv/Fm ratio,
did not detect mite damage in the container study even though A had been reduced (Table
9). It is surprising that photosynthesis was reduced without affecting the Fv/Fm ratio
which has been shown to be a sensitive indicator of the photosynthetic apparatus. The
senior author noted while conducting the container study that initially Fv/Fm values may
have actually increased with increasing mite damage, however, there was no such increase
statistically. Visual damage was the best indicator of mite damage as determined from this
study.

Sour chenies exhibit a considerably higher tolerance to mite-feeding than either
apple or peach. This tolerance is in part due to the fact that mite damage occuned late in the
growing season when photosynthetic demand in sour cherry is relatively low. Also, sour
chenies are able to compensate for photosynthetic losses due to mite-feeding to the point
that cold hardiness and return yield are not affected. This ability of tart cherries to
withstand mite—feeding late in the season means that routine miticide applications can be
reconsidered in Michigan orchards. Mite control measures that signiﬁcantly reduce mite
populations relative to the uncontrolled alternative had no signiﬁcance on tree hardiness and
yield in this study.

Eﬁects of Miticides. The miticides used for sour cherry production in Michigan

can be used with no concern that the chemicals themselves will have any adverse effects on

29
tree productivity. Repeated applications or applications of other chemicals may have an

adverse effect on the crop (Ayers and Barden, 1975; Andersen et al., 1986).

Terbacil Studies. By selecting the exact concentration of terbacil, photosynthesis,
particularly A and Fv/Fm ratio, could be reduced by a predetermined amount for a
predetermined amount of time. The degree and time of recovery of both A and Fv/Frn were
concentration dependent. We purposefully chose a concentration that would reduce A by
approximately 50% for 7—10 days. Presumably, naturally occuning stresses in the orchard
would be detected and control measures employed that would allow the trees to recover in
this time period.

A was initially reduced more than Fv/Fm ratio but recovered more quickly. The
slow recovery of the Fv/Fm ratio suggested to us that that parameter was detecting
photosynthetic damage that the A measurement was not. _ As mentioned previously, the
Fv/Fm ratio was not effective as an early indicator of mite damage, but may still be an early
indicator of other types of stresses. Photosynthetic recovery to the application of terbacil
was similar on all applications dates except on spray date 4. Full recovery of Fv/Fm was
not achieved prior to leaf fall, and full recovery of A took 14 days, twice the time for
recovery for the other applications dates. leaf senescence had been initiated at this point
and the effect of terbacil was compounded by leaf senescence.

The effects of terbacil spray on whole tree A and recovery were not signiﬁcantly
different than single leaf measurements of A. It might be assumed that tree as a whole
would be quicker to recover net photosynthesis due to photosynthetic compensation as
more and more leaf area recovers. No compensation was observed, and the inhibition was
likely not enough to induce measurable photosynthetic compensation in such a short time
period.

The lack of any adverse effects of the terbacil treatments on fruit yield or quality
indicates that sour cherry trees can tolerate substantial reductions in photosynthesis -

during critical periods of photosynthate demand - with no effect on yield. Byers et al.

30
(1990) have shown an effect of terbacil on yield of peach and used photosynthetic

inhibitors as fruit thinners. It is important to note that the leaf to fruit ratios in the orchard
used in this study were well above the threshold for fruit production and the trees therefore
had signiﬁcant buffering capacity to the effects of the terbacil. Additionally, sour cherry
trees appear to have signiﬁcantly greater abilities to withstand both mite damage and
photosynthetic reduction. Inhibition of photosynthesis can occur at one of several times
during the growing season, including postharvest and still have no effect on yield or
quality. Additionally, multiple reductions in photosynthesis that require up to 14 days for
100% recovery had no adverse effects on current season's yield or quality (Appendix 0)

As the mode of action of terbacil is on electron transport of photosystem II
(Gardiner, 1981), such low concentrations as used in this study should not have affected
chlorophyll levels and did not. Yellowing of leaves due to chlorophyll loss is a signal of
terbacil toxicity and concentrations that cause such damage were purposefully avoided to
allow 100% recovery of leaves.

The treatment of trees with terbacil which led to the reduction in photosynthesis did
have an effect on other parameters in this study. Shoot extension was reduced in the trees
sprayed with terbacil immediately prior to harvest The inhibition of photosynthesis during
the ﬁnal days of fruit carbohydrate assimilation and growth resulted in reduced production
of photosynthate, apparently at the expense of stem tissue growth. The effect may have
been to slow shoot growth or to accelerate terminal bud set, the timing of which was not
measured in this study. There was no measurable increase in A after recovery to
compensate for the reduced photosynthesis at such a critical stage. The application of
terbacil and subsequent photosynthetic reduction at other stages of development had no
effect on shoot extension. At those stages of growth, the trees apparently have the ability
to withstand photosynthetic reduction due either to reduced demand during these periods.

or to undetected photosynthetic compensation. The period of time immediately before

3 1
harvest is critical for maintaining maximum photosynthesis for fruit development and shoot

growth.

While shoot extension was affected by photosynthetic reduction at harvest, shoot
starch levels at the end of the growing season were not affected by photosynthetic reduction
at harvest. Reduced autumn starch levels of trees treated at stage 1 indicates that fruit
development at this stage has priority over assimilate partitioning to the point that fruit
development was unaffected but storage carbohydrates were reduced. However, by the
spring of the following year, the shoots had recovered such that they had starch levels
comparable to controls. There was sufﬁcient time between that early June photosynthetic
reduction and leaf fall to produce sufﬁcient starch that could be transported to the shoots in
the early spring. The spring starch levels of the other treatment dates were increased over
controls, even for the treatment date of 17 September. This indicates that the time of
application relative to leaf fall had little to do with the amount of starch stored in the shoots.
The response of the trees to photosynthetic inhibition was to preferentially transport starch
to the shoots in the spring but not in the fall. That preferential transport may have been at
the expense of root starch as levels were reduced for treatment date 2. The effect may have
been for the treated trees, presumably lower in overall starch levels, to transport starch to
the shoots to maximize shoot cold hardiness, which was not affected. The starch levels
were consequently reduced in the roots, signiﬁcantly for treatment date 2. The terbacil-
induced photosynthetic reductions at stages 1 and 2 had conu'asting responses with respect
to starch storage between the roots and shoots. Inhibition of photosynthesis during stage I
of fruit growth, a period of rapid and proliﬁc cell division, resulted in less starch being
stored in the shoots but no effect on the roots. This indicates that root starches are possibly
produced or transported later in the season. Inhibition 14 days later resulted in no change
in sheet starch levels but considerably less in the roots. Later dates of inhibition had no
effect on root starch levels. This indicates that the period of pit hardening, when fruit

development pauses, may be a period during which starch is produced for or transported to

32
the roots. Assirnilate is still transported to the fruit during this time, and if starch is

destined for the roots during this stage, signiﬁcant levels‘photosynthate required during this
brief period. This would be in direct contrast to the partitioning that occurred only 14 days
earlier during stage 1. Starch storage at stage II is obviously markedly different than the
storage at stage I particularly under conditions of reduced photosynthesis.

These affects on starch levels may have been inconsequential as there was no effect
on shoot cambium cold hardiness or subsequent season's yield. Starch concentrations
have been correlated with cold hardiness of peach (Lasheen et al., 1970; Flore and Howell,
1987), but there was no effect in our study of reduced starch levels on cold hardiness of
tart cherry. Greater photosynthetic reduction would have been required to alter cold
hardiness (Flore, et al., 1983) or subsequent season's yield (Howell and Stackhouse,
1973).

The results of this study indicate that sour cherry trees can tolerate substantial losses
in photosynthesis without any adverse effects on overall orchard production. While the
levels of photosynthetic reductions in this study did not lower total yields or fruit quality,
other factors were altered dependent upon the time of photosynthetic reduction. Shoot
extension and root and shoot starch levels were altered as a result of photosynthetic
reductions. The lack of changes in the yield components as well as cold hardiness indicate
that the sour cherry trees were able to compensate for lost photosynthate with no cost to
reproductive growth or tissue hardiness. Therefore, photosynthetic reductions that occur in
the natural life of the orchard, whether those reductions be due to mites, insects, diseases,
drought or ﬂooding suess, or otherwise, can have little effect on the economically
important factors of yield and tree vigor. Granted there are many other substantial
differences between the terbacil induced photosynthetic reduction of this study and typical
environmental stresses an orchard will encounter. When considering the effect of
photosynthate loss alone, which the terbacil ueatrnent allowed, photosynthetic reductions

of up to 10 days can have little detrimental effect.

33
This data can be incorporated into integrated pest management programs for sour

cherry. The demonstrated tolerance of orchard sour cherry trees to mite feeding and
photosynthetic reductions will be useful information for growers as they make decisions
concerning when to utilize mite and other pest control measures. It is important to note that

the long term effects of mite feeding are still uncertain.

34
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applications of pesticides on leaf gas exchange of peach. HortScience 21(3):508-
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Anderson, PC. and R.F. Mizell, III, 1987. Impact of the peach silver mite, Aculus
comutus (Acari: Eriophyidae), on leaf gas exchange of 'Flordaking' and 'June
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Avery, DJ. and J.B. Briggs. 1968. Damage to leaves caused by fruit tree red spider mite,
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Ayers, Jr., J .C. and J.A. Barden, 1975. Net photosynthesis and dark respiration of apple
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Bailey, P. 1979. Effect of late season populations of twospotted mite on yield of peach
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Bassman, J. H. and D. I. Dickmann, 1982. Effects of defoliation in the developing leaf
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Beers, EB. and L.H. Hull. 1987. Effect of European red mite (Acari: Tetranychidae)
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Bittenbender, BC. and 0.8. Howell, Jr., 1974. Adaptation of the Spearman—Kalrber

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Byers, R.E., J.A. Barden and DH. Carbaugh, 1990. Thinning of spur 'Delicious' apples
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Corelli-Grappadelli and E. Magnanini, 1993. A whole-tree system for gas exchange
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Flore, J.A., G.S. Howell and CE. Sams, 1983. The effect of artiﬁcial shading on cold
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Flore, J.A. and C. Irwin, 1983. The inﬂuence of defoliation and leaf injury on leaf

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39

Figure 1. Whole plant photosynthetic C02 assimilation chamber.

Figure 2. (A): Calculated cumulative mite'days on control and mite—infested orchard sour
cherry trees on speciﬁed dates during the 1993 season at Clarksville Michigan. Arrow
indicates miticide application to control treatment trees. Each point is the mean of 8 trees.
Vertical bar represents LSD at 5%. (B): Calculated cumulative mite-days on control and
mite—infested container sour cherry trees on speciﬁed dates during the 1994 at East Lansing
Michigan. Each point is the mean of 22 trees. Vertical bar represents LSD at 5%.

Figure 3. The effect of spraying 63 ppm terbacil on CO2 assimilation and Fv/Fm ratio of
orchard sour cheny leaves at Clarksville Michigan in 1993. Treatment dates were (A): 1
June, (B): 15 June, (C): 4 August and (D): 17 September. Each point is the mean of 4

trees. Vertical bar represents LSD at 5%.

Figure 4. The effect of spraying 63 ppm terbacil on whole tree C02 assimilation of orchard
grown sour cherry trees at Clarksville Michigan in 1993. Each point is the mean of 2 trees.

Vertical bar represents LSD at 5%.

 

Figure 1.

Cumulative MiteeDays

Cumulative MiteoDays

41

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

 

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Table 1. The effect of mite-infestation on single leaf C02 assimilation of sour cherry leaves

from Clarksville Michigan orchard trees on speciﬁed dates in 1993.

 

 

 

 

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Table 5. The effect of application of 63 ppm terbacil on shoot starch levels on 1 November

1993 and shoot and root starch levels on 14 April 1994 of orchard sour cherry trees.

Terbacil #1 sprayed on 1 J une, Terbacil #2 sprayed on 15 June, Terbacil #3 sprayed on 4

August and Terbacil #4 sprayed on 17 September.

 

.Ll‘ioxembeuﬂﬂl __M_Anril_129_4__

 

 

___In:atmsmt Shoot Shoot Root
Control 8.38% b 8.11% b 5.77% ab
Terbacil #1 6.13% a 11.49% ab 6.98% a
Terbacil #2 7.73% b 12.13% a 2.17% c
Terbacil #3 8.99% b 12.23% a 4.58% bc
Terbacil #4 8.46% b 12.64% a 5.26% abc

p<5% p<8% p<10%

Treatment differences are not signiﬁcant, NS, or signiﬁcant at the probability level

indicated.

 

SUMMARY AND CONCLUSIONS

It was demonstrated that moderate levels of mite populations that occur in the
absence of chemical miticide applications can result in no signiﬁcant decrease in tree yield
or hardiness compared to typical control measures taken. Compared to an absolutely mite—
free tree, the effects may be very different. In this study, 1764 CMD did not reduce
photosynthesis, yield or hardiness compared to 1080 CMD in the orchard study. In the
container study, 900 CMD reduced a number of photosynthetic parameters when compared
to only 90 CMD of controls. Therefore, while orchard trees not treated with miticides were
shown not to be adversely affected, there may have been adverse affects if they had been
compared to truly mite—free trees. The latter consideration may be inconsequential since
absolutely mite—free orchards are not attainable. Additionally, the orchard trees may have a
significantly greater capacity to withstand the effects of mite feeding. It may be possible
that during certain years sufﬁcient control measures can be taken that would reduce
explosive mite populations enough to achieve a benefit, although this was not the case in
our orchard in 1993.

The question of whether the miticide sprays themselves have a detrimental effect on
photosynthesis and therefore tree yield and hardiness was addressed also as part of this
study. The two major miticides labeled for sour chenies and used in Michigan
demonstrated no inhibitory effects on photosynthesis.

As the mite populations in the orchard did not affect photosynthesis and the mite
populations in the container study did reduce photosynthesis, it is unclear as to how much
photosynthetic reduction the tree and the orchard can tolerate before yield and hardiness are
affected. It was demonstrated that reductions in photosynthesis that required 10 or more
days to fully recover had no effect on current season or subsequent season yields. These

were photosynthetic reductions of 50% or more that gradually recovered to 100% in 5- 14

50
days. Therefore, plant stresses that reduce photosynthesis to these levels for similar time

periods may be insigniﬁcant in regard to sour cherry yield and vigor.

All of these data indicate that in some seasons, applications of miticides may be
unnecessary even when reductions in photosynthesis and growth can be detected. Mite
and insect population thresholds for sour cherry may be much higher than originally
believed, particularly with regard to thresholds established for apple and peach.
Additionally, economic and action thresholds could be pushed back as well to reﬂect the
tolerance to photosynthetic reduction.

While reduction of chemical inputs into the orchard can be of beneﬁt to the grower,
the loss of these management tools must be replaced by other pest control measures.
However, it was demonstrated that for our orchard in 1993, a mite population threshold
was not reached that reduced photosynthesis, yield or hardiness. Likewise, a
photosynthetic reduction threshold was not reached that reduced yield or hardiness.
Therefore, mite control measures were not necessary in that year. Additionally, measures to
alleviate other stresses to photosynthesis may not be as necessary as well in similar
situations. Thus the tart cherry grower could effectively produce a crop with reduced

chemical inputs.

51

APPENDD( A

Regression Figures of 1993 Orchard Single Leaf C02 Assimilation, Whole Tree C02
Assimilation, and Chlorophyll Fluorescence Against Mites Per Leaf and Cumulative
Mite°Days.

52

Figure 5. Single leaf C02 assimilation (umol-m'ZOS'l) for orchard sour cherry trees in
1993 at Clarksville Michigan plotted against (A) mites per leaf (r2 = 0.03, n=44), and (B)

cumulative miteOdays (r2 = 0.12, n=44).

Figure 6. Whole tree C02 assimilation (umokm‘z-sl) for orchard sour cherry trees in
1993 at Clarksville Michigan plotted against (A) mites per leaf (r2 = 0.12, n=8), and (B)

cumulative mite'days (r2 = 0.0001, n=8).

Figure 7. Fv/Fm ratio from chlorophyll ﬂuorescence for orchard sour cherry trees in 1993
at Clarksville Michigan plotted against (A) mites per leaf (r2 = 0.13, n=74), and (B)

cumulative mite'days (r2 = 0.33, n=74).

Figure 8. Effect of cumulative mite day populations for orchard sour cherry trees in 1993
at Clarksville Michigan on (A) 1994 yield (kg per tree) (r2 = 0.01, n=16), and (B) cold
hardiness (T 50) of 1993-94 dormant season (r2 = 0.004, n=16).

Figure 9. Effect of cumulative mite day populations for orchard sour cherry trees in 1993
at Clarksville Michigan on (A) 1993 fall shoot starch concentrations (percent by weight) (r2
= 0.004, n=16), (B) 1994 spring shoot starch concentrations (percent by weight) (r2 =
0.27, n=16), and (C) 1994 spring root starch concentrations (percent by weight) (r2 =

0.26, n=16).

53

9. Fa. 8.883 gouge—m3 NOD

 

mm mim...m.

15 20 25 30
Mites Per Leaf

10

 

 

_ _
[1 1 . 4

m. m. whim

8 6 4 2. 0.

3- m. N- 8.3.53 :33ng ~00

1500 2000

1000

Cumulative Mite-Days

500

Figure 5

54

A

h p h —
a q _ . — . _

_ _ p

q a a
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15 20 25 30
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1 1

3. 9n- 8.383 8088834 NOD

 

1250 1500

1000

750
Cumulative Mite-Days

500

250

Figure 6

Fv/Fm Ratio

55

0.8

 

 

 

 

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o 0. o 09 ,
If: ‘ 21”.. 9
0.6
, o
o
. O
0.4 —~
0.2 -~
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0 500 1000 1500 2000
Cumulative Mite-Days

Figure 7

Cold Hardiness — T60) (°C)

 

56

 

 

 

70 q—
4 o
60 —- . o
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z 30 db . o .0
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-20 -_
- -_ 0 ’ o
30 0 ~, ' . ’ . .
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0 500 1000 1500 2000
Cumulative Mite-Days

Figure 8

 

57

 

 

 

 

 

0.12 T A
'5 01 3» .
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r: J ’
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0 500 1000 1500 2000
1993 Cumulative Mite-Days

Figure 9

58

APPENDIX B

Cold Hardiness of Orchard Sour Cherry Trees Infested with Mites in 1993.

59

Figure 10. Stem tissue cold hardiness as determined by T(5()) values for controls and

mites-infested trees on the speciﬁced dates in the 1993-94 dormant season at Clarksville

Michigan (each bar is the average of 8 trees). Treatment means were not signiﬁcantly

different analyzed using ANOVA and F test at 5%.

lS-Oct l9-Nov 21-Dec l4-Jan lS-Feb 15-Mar 12-Apr Average

,1 L l l 7
1 on. I I'll I III-l

 

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[:J Control

 

 

 

 

 

 

 

 

 

 

 

ESEMites

 

 

 

 

 

 

 

Figure 10

61

APPENDIX C

Effects of Miticides on C02 Assimilation, Stomatal Conductance, Transpiration and

Chlorophyll Fluorescence of Greenhouse Grown Sour Cherry Trees.

62

Figure 11. The effect of miticide application on (A): C02 assimilation (A), (B): stomatal

conductance (gs), (C): transpiration (E), and (D): Fv/Frn of expanded sour cherry leaves

in East Lansing Michigan greenhouse in 1994. Means i SE (n: 10).

63

 

 

 

 

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Table 6. The effect of miticide application on shoot length increase and chlorophyll of

greenhouse grown sour cherry trees after application.

 

Treatment Lgngth (cm) Total Chlorophyll (ugOQm'Zl

 

 

Control 10.92 92.4

Orrrite 11.75 92.3

Vendex 11.75 95.2
NS NS

Treatment differences are not signiﬁcant at 5% probability level, NS, or signiﬁcant at the

probability level indicated.

 

65

APPENDIX D

Effects of Differing Concentrations of Terbacil on C02 Assimilation and Chlorophyll

Fluorescence of Sour Cherry Trees.

66

Figure 12. The effect of terbacil concentration on (A): C02 assimilation (A) and (B):

Fv/Fm ratio of expanded leaves of container grown sour cherry trees. Means :t SE (n=6).

67

 

 

 

 

90.850 8.3 c .«o 88
S- m. N.8883 83388.8(. Nov

 

 

 

 

 

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5

3
Days after treatment

4

2

1

Figure 12

68

APPENDIX E

Shoot Extension and Chlorophyll Levels of Orchard Sour Cherry Trees Treated with 63
ppm Terbacil on Speciﬁced Dates in 1993.

69

 

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70

APPENDIX F

Cold Hardiness of Orchard Sour Cherry Trees Treated with 63 ppm Terbacil on Speciﬁced
Dates in 1993.

71

Figure 13. Stem tissue cold hardiness as determined by T(50) values for controls and trees
treated with 63 ppm terbacil on the speciﬁced dates in the 1993-94 dormant season at
Clarksville Michigan. Cont: unsprayed controls, Terbl: sprayed on June 1, Terb2: sprayed
on June 15, Terb3: sprayed on Aug 4, Terb4: sprayed on Sept 23 (each bar is the average
of 4 trees). Treatment means were not signiﬁcantly different analyzed using ANOVA and

F test at 5%

19-Nov 21-Dec l4-J an 15-Feb 15-Mar 12-Apr Average

lS-Oct

72

 
 

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73

APPENDIX G

1994 Terbacil Spray Methods and Results

In 1994, trees at the Horticultural Research and Teaching Center at East Lansing
Michigan were sprayed one or more times with 63 ppm terbacil with 1 ppm X-77, as a
study of repeated photosynthetic reduction. Trees were sprayed as one of 5 treatments:
controls received no sprays; single sprays at stage I of fruit development (30 May); multiple
sprays at stage I and stage H (30 May and 14 June); single spray at stage II (14 June); or
multiple sprays at stage II and late stage H (14 and 27 June). Spray applications were made
prior to 9:00 EDT using a backpack sprayer with foliage sprayed to drip, approximately 8
Utree.

The effects of multiple terbacil applications during fruit development on A and
Fv/Fm ratios are shown in Figure 14. Figure 14A shows that A recovered within 7 days
even for multiple applications. Fv/Fm ratios (Figure 14B) took longer to recover after the
second application but fully recovered by 14 days. Signiﬁcantly, this additional reduction
in photosynthesis and extended reduction in photochemical efﬁciency (Fv/Fm) did not

reduce fruit quality (Table 8) or fruit yield per limb cross sectional area.

74

Figure 14 . The effect of single and multiple spray applications of 63 ppm terbacil on (A):
C02 assimilation and (B): Fv/Fm ratio of expanded sour cherry leaves from orchard trees
in 1994. Terbacil #1 was treated on 30 May, Terbacil #1' x 2 was treated on 30 May and 14
June, Terbacil #2 was treated on 14 June and Terbacil #2 X 2 was treated on 14 June and

27 June. Arrows indicate dates of terbacil application. Means 1 SE (n=4).

% of Control

Fv/Fm ratio

75

 

 

 

   

 

 

 

 

 

_°— Control
0'4 ‘ —-—- Terbacil #1
‘ —*- Terbacil #1 X2
0.2 ‘ —°_ Terbacil #2
+ Terbacil #2 X2
0.0 I I r 1

6/1/94 6/6/94 6/15/94 6/22/94 6/28/94 7/5/94

 

 

 

 

 

 

 

 

0.8
0.7 ‘
0.6 ‘
0.5 ‘
.4. T
t T —'°— Control
03‘. —*— Terbacil #1
0.2 - —*—' Terbacil #1 X2
: —°— Terbacil #2
0‘1 . —-— Terbacil #2 x2
0.0 I I l

 

 

1 Jun 6 Jun 15 Jun 22 Jun 28 Jun 5 Jul 12 Jul

Figure 14

76
Table 8. The effect of application of 63 ppm terbacil on fruit weight, soluble solids and

removal force for orchard sour cherry trees in 1994. Terbacil #1 sprayed on 30 May,
Terbacil #1 X 2 sprayed on 30 May and 14 June, Terbacil #2 sprayed on 14 June and
Terbacil #2 X 2 sprayed on 14 June and 27 June.

 

 

 

Treatment Fruit Weight Soluble Solids Fruit Removal Force
(g) (%) (g)
Control 4.50 1 1.50 376
Terbacil #1 4.21 12.13 311
Terbacil #1 X 2 4.08 11.67 354
Terbacil #2 4.31 11.58 430
Terbacil #2 X 2 4.75 10.92 387
N S N S N S

Treatment differences are not signiﬁcant, NS, or signiﬁcant at the probability level

indicated.

 

77

APPENDIX H

Pesticide Residue Study

PESTICIDE RESIDUES IN PEACH (PRUNUS PERSICA L.
'NEWHAVEN') FRUIT GROWN UNDER IPM AND CONVENTIONAL
PEST CONTROL

Mark A. Hubbardl, James A. Florez, E. Hanson3, W. Shane3, J. Johnsonz, J. Wisel, M.
Whalonz, G. Bird2 and A. J ones2

Department of Horticulture, Michigan State University, East lansing, MI 48824-1325

Received for publication

1Graduate Research Assistant

2Professor

3Associate Professor

78
Cellular and Whole Plant Physiology

Pesticide Residues In Peach (Prunus persica L. 'Newhaven') Fruit Grown Under IPM And

Conventional Pest Control

Additional index words. IPM

Abbreviations: EPA, Environmental Protection Agency; GC, gas chromatography; IPM,
integrated pest management; CONV, conventional level of chemical input; MOD, moderate

level of chemical input; LOW, low level of chemical input

Abstract. Six peach (Prunus persica L.) orchards were established in 1990 and three
distinct management strategies were employed that compared different levels of synthetic
chemical input: a conventional system (CONV) that utilized standard chemical control
measures, a moderate level of chemical input (MOD) that monitored pest populations for
pesticide use and integration of nonchemical controls, and a low level of chemical input
(LOW) that further reduced chemical inputs by employing additional IPM strategies and
applying pesticides only when absolutely necessary as determined by entomologist.
Selected pesticide residues in the fruit and soil were determined for 1992 and 1993. For
fruit harvested in 1992, there were no signiﬁcant differences between the pesticide residues
from the different orchards, although there had been a distinct difference in the number of
pesticide sprays each orchard received. In 1993, there was again a distinct difference in the
pesticide sprays for each orchard with only minor difference in residues. The CONV
orchards did have higher residues of the fungicide Captan than the MOD or the LOW
orchards. Also, the conventional orchards had residues of chlorpyrifos of 0.08 ppm which
is above the Environmental Protection Agency (EPA) tolerance of 0.05 ppm. The
chlorpyrifos residues in the MOD and LOW orchards were below the EPA tolerance, 0.03

79
ppm and 0.04 ppm respectively. For the two management years of this study, few

differences were found in the pesticide residues among the different treatments, however,

CONV orchards did have chlorpyrifos residues above EPA tolerances in 1993.

There is growing concern about the use of synthetic pesticides in the production of
food crops. This has sparked considerable interest and study in the use of management
practices that reduce chemical control measures and utilization of nonchemical strategies.
Integrated pest management (IPM) incorporates the technologies of horticulture,
entomology, plant pathology and other ﬁelds to maintain quality crop production using
innovative pest management strategies to minimize environmental impact to the site as well
as the crop itself.

Peach growers are encountering pressure to produce quality fresh fruit and at the
same time meet the increasing consumer demand for fruit produced with a minimum of
chemical input to the orchard environment. The orchard manager must control a number of
persistent insects, mites, and diseases to produce a crop and maintain crop quality. With
the advent of IPM strategies and new biological controls, there exist the possibility of
producing the quality and quantity of fruit with reduced use of synthetic chemicals.
However, these techniques require greater skill in orchard management and the production
results of exclusively using these strategies on a commercial scale in unknown. Also, the
effects of reduced synthetic inputs on pesticide residues in the fruit is also uncertain.
Residues have become a major health concern, but not without controversy. The debate
ranges from dramatic press headlines (Blume, 1987) to scientiﬁc review (CAST, 1990).
The EPA has established pesticide residue tolerances for all synthetic chemicals registered
for use in the US. (Code of Federal Regulations, 1994). The methodology by which the
EPA establishes health risks has also been called into question (Gold et al. 1992; Arnes and
Gold, 1990).

80
This study was initiated to compare three levels of chemical input and IPM

strategies to determine if reducing synthetic chemical input into the orchards will reduce the

detectable pesticide residues in the fruit and soil.

Materials and Methods

Orchard Management Strategies. The establishment and management strategies of
the orchards has been previously discussed in detail (Flore, et al. 1994) and will be brieﬂy
described here. Six 'Newhaven' peach orchards were established in 1990, and the
treatments applied were conventional chemical input. moderate level of chemical input and
low level of chemical input. The conventional treatment consisted of production practices
typical of those used by peach producers in southwest Michigan: preplant fumigation,
clean cultivation of the soil, broadcast application of fertilizer, scheduled insecticide sprays
and herbicide sprays of paraquat and simazine, and dormant pruning. Pesticide sprays
were utilized according to spray guidelines issued by Michigan State University
Cooperative Extension Service. The moderate level of chemical input included a fescue
ground cover, fertilizer application through drip irrigation lines, insect scouting for spray
scheduling, application of sulfur in place of synthetic fungicide sprays, conventional
herbicide sprays of sirnazine and paraquat, and dormant pruning. Scouting included
monitoring the presence of Oriental fruit moth, tarnished plant bug, and peach tree borers
with sticky boards and traps. Once treatment thresholds were exceeded a spray was made
to control populations. The low level of chemical input included an endophytic rye ground
cover for tarnished plant bug control, utilization of novel insect controls, insect scouting for
spray scheduling, control of nematodes with Pseudomonas predation, nitrogen fertilizer
applied in the form of horse manure, application of sulfur in place of synthetic fungicide

sprays, straw mulch in tree rows for weed control, no synthetic fertilizer application, and

8 1
summer pruning. Insect controls included pheromone disruption for control of Oriental

fruit moth by placing pheromone ties in the trees to effectively saturate the orchard
environment and thus prevent mating. The amount of horse manure applied gave the
equivalent amount of actual nitrogen per ha as the other fertilization methods.

Pesticide Extraction and Analysis. Fifty kg of fruit were randomly collected from
each orchard and the samples for each orchard combined. Twenty fruit samples were then
cut into 4-5 g sections, retaining the epidermis but removing the pit, and stored at -30° until
extracted. For extraction, fruit was thawed for 10 min at room temperature to allow for
additional sectioning and blending. Soil samples were collected from 4 predesignated
locations on the perimeter as well as from the center of each orchard and combined and the
200 g samples stored at -30° until extraction. All pesticides were extracted according to
procedures obtained from M. Zabik of the Pesticide Research Center at Michigan State
University and were in accordance With EPA methodology: sirnazine was extracted from
the soil using the procedure of Smith (1981) and polyclonal antibody kits obtained from
Millipore Corp. (Bedford, MA); chlorpyrifos and azinphos methyl were extracted from a
procedure originally obtained from Shell (Modesto, CA); fenvalerate was extracted based
on a procedure originally obtained from DuPont (Wilmington, DL); iprodione,
chlorothalonil and captan were extracted based upon the procedure of Liao, et al. (1991);
elemental sulfur was extracted using a procedure originally obtained from the EPA
(Washington, DC). Triplicate samples were extracted from the 1992 fruit and duplicate
samples extracted from the 1993 fruit. Samples spiked with each pesticide were extracted
every six samples.

Extracted samples were analyzed on a Hewlett Packard gas chromatograph (model
5890 series 11) with a 25m capillary DB-5 column and an electron capture detector.
Injection was via a Hewlett Packard automatic injector (model 7673 series H). GC
parameters were as follows: split injection mode, inlet temperature 240°; helium carrier gas

with a column head pressure of 12.9 psi, nitrogen makeup gas, initial oven temperature of

8?.
180° for 20 min and then increasing at 10° per min to 275° and holding at 275° for 5 min;

and detector temperature was 300°. Duplicates were run of each sample along with the
spiked samples of pesticide standards obtained from Chem Service (Chester, PA). A
standard was delivered no less than every six samples. Standards that were run included
the ﬁrst degradation products of chlorpyrifos, chlorothalonil and iprodione as these
compounds are included in the EPA tolerance levels (Code of Federal Regulations, 1994).
Retention times for the samples were compared to retention times for the standards and the
amount of pesticide in each sample calculated from the area of the detector response as
measured by integration (Hewlett Packard model 3396 series 11 Integrator). Recoveries of
the extraction procedures were calculated based on the amount of pesticide found in the
spiked samples.

Statistical Procedure. Treatment means were analyzed for differences using
ANOVA and F test. Differences were considered signiﬁcant at the 5% level. Residues
were statistically analyzed separawa for 1992 and 1993 as pest control treatments were

different for each year and residue values were markedly different each year.
Results

Orchard Management Strategies. The number of pesticide spray applications to
each treatment orchard are reported in Table 9 for 1992 and Table 10 for 1993. In 1992,
the CONV orchards received a total of 14 synthetic chemical sprays compared to 9 for the
MOD and 2 for the LOW. In addition the LOW orchards received 3 pesticide sprays that
were applied only to the perimeter of the orchard, thus minimizing the amount of pesticide
applied to harvested fruit. Also, the MOD orchard was treated with 6 sprays of elemental
sulfur and the LOW was treated with 2 sulfur sprays. In 1993, the CONV orchard

received 16 spray applications of synthetic chemicals,- the MOD 6 spray applications and

. 83
the LOW 2 applications. Again, the MOD orchard received 6‘ sprays of elemental sulfur

while the LOW received 2.

Pesticide Extraction and Analysis. The results from the pesticide extraction and
analysis of the fruit are given in Tables 11 and 12. In 1992, Iprodione was detected in the
CONV but not in the MOD or the LOW as it was not applied to these orchards. No
iprodione was detected in any of the 1993 fruit samples. Chlorothalonil, applied only to
the CONV was not detected in any of the samples except for detection of trace levels in
1993 in the CONV orchards. No captan was detected in 1992, but was detected in the two
orchards to which it was applied to in 1993. Chlorpyrifos was not detected in 1992, but
was detected in fruit from each. treatment orchard in 1993,. In the case of the CONV,
chlorpyrifos exceeded the EPA tolerance of 0.05 ppm. No azinphos methyl was detected
in any of the samples analyzed for either year. Fenvalerate was detected in equal amounts
in each treatment for both 1992 and 1993. Sulfur residues were highest in the MOD as
those orchards received the most sulfur spray applications.

Sirnazine residues in the soil were not statistically different between treatments in
1992, the CONV had 8.0 ppb, the MOD 22.3 ppb and the LOW 12.2 ppb.

Recoveries for the extraction and analysis procedures ranged from 80-160% as

determined from analysis of the spiked samples.
Discussion

Distinctly different levels of pesticides were applied to each of the different
treatment orchards as a result of the distinct management strategies employed for each. The
MOD orchards utilized a number of IPM techniques that eliminated the need for several of
the synthetic chemical sprays. In particular, the use of sulfur as a fungicide markedly
reduced the chemical input into those orchards. The fruit quantity and quality was
comparable to the CONV (Flore et al., 1994) indicating that IPM strategies can effectively

84
control pest populations and reduce chemical input into the orchard. The LOW orchards

utilized additional IPM strategies and used synthetic chemicals only if absolutely necessary
to prevent damage to the entire crop. However, the LOW orchards had a lower level of
fruit free of insect and disease damage, 79% compared to the 95% for the CONV (Flore et
al., 1994). Thus, a signiﬁcant crop can be produced by drastically reducing chemical
inputs, but at present there is a tradeoff in quality and quantity.

There are obvious beneﬁts to the producer and the consumer from producing fruit
crops with reduced levels of synthetic chemicals. The producer may realize a net cost
savings as a result of lower pesticide expenses, and the cumulative effect of pesticide
application to the orchard site is reduced. The consumer beneﬁts from knowing that fewer
pesticides have been added to their food and the environment as a whole.

There is considerable controversy over the health effects of reduced pesticide input
in fruit production. It is assumed that lower chemical inputs will mean more beneﬁcial fruit
as a result of lower pesticide residues. This study demonstrates that all chemicals inputs
are not clearly reﬂected in the fruit residues, but reducing late season chemical inputs can
reduce pesticide residues. The levels of fungicides applied were most noticeably different
each year, but only in 1992 were any residue differences detected. The CONV orchards
received the most fungicide applications and consequently had the higher iprodione residue
in 1992. In 1993, captan was detected at 6.3 ppm in the CONV, yet there was no
signiﬁcant difference due to the high variability in the captan residues in the CONV '
orchards. Additionally, while fruit from the LOW and MOD orchards were free from
synthetic fungicide residues, sulfur residues from those orchards was highest, although not
statistically different. The insecticide fenvalerate was detected in each orchard each year,
but only at higher levels in the 1993 in the CONV and MOD orchard fruit, In 1993, the
level of chlorpyrifos in the CONV orchards was higher than the EPA tolerance for peach
fruit, and the residues detected in the other treatment orchards were below the tolerance

limit and different from the CONV residue level. As seen in this study, conventional pest

85
management can lead to fruit with excessive levels of pesticide residues as can poorly timed

applications of pesticides in an IPM scheme. Implementation of IPM strategies is
becoming a necessary pest management tool due to increased restriction on pesticide use,
producer concern for the orchard environment and consumer preference. Reduced
synthetic chemical pesticide input does not necessarily mean lower residues in the fruit.
The manager must take into consideration the timing of chemical application with respect to

harvest as well as alternative pest control measures.

Literature Cited

Ames, B.N. and LS. Gold, 1990. Too many rodent carcinogens: Mitogenesis increases

mutagenesis. Science 249:970-971.

Blume, E., 1987. Poisoned peaches, toxic tomatoes: reckoning pesticide risks. Nutrition

Action Newsletter, October pp. 8-9.

Code of Federal Regulations, 1994. Title 40, Part 180. Tolerances and exemptions from
tolerances for pesticide chemicals in or on raw commodities. US. Government

Printing Ofﬁce, Washington, DC.

Flore, J.A., M. Hubbard, E. Hanson, J. Johnson, J. Wise, M. Whalon, G. Bird, and A.
Jones, 1994. Low input production of peach (Prunus persica cv. 'Newhaven').

Acta Horticultura, in press.

Gold, L.S., T.H. Slone, B.R. Stern, N .B. Manley and B.N. Arnes, 1992. Rodent

carcinogens: setting priorities. Science 258:261-265.

86

Liao, W., T. Joe, and W.G. Cusick, 1991. Multiresidue screening method for fresh fruits
and vegetables with gas chromatography/ mass spectrometric determination. J.

Assoc. Off. Anal. Chem. 74(3):554~564.

Smith, A.E., 1981. Comparison of solvent systems for the extraction of atrazine,
benzoylprop, flamprop, and trifluralin from weathered ﬁeld soils. J. Agric. Food
Chem. 29:111-115.

87

 

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91

APPENDD( I

Pesticide Residues Extraction Protocols

Protocol for azinphos methyl and chlorpyrifos:

1.
2.

Add 65 mL acetone to 20.0g of peach fruit and blend for 3 min. at medium speed.
Filter through Whatrnann #4 paper using suction; rinse the blending apparatus
thoroughly and also ﬁlter.

. Extract with equal volume of methylene chloride in separatory funnel by shaking for 1

min. and allowing the phases to separate for 3+ min.

. Percolate the lower, acetone—methylene chloride layer through a 2x5cm plug of

anhydrous sodium sulfate.

Extract the upper aqueous phase with 25 mL methylene chloride via separatory funnel ,
shaking 30 sec. and allowing 3+ min. separation, percolate lower phase through
anhydrous sodium sulfate.

Evaporate the acetone—methylene chloride eluate to dryness with a jet of air and 50°C

water bath.

. Dissolve residue in 50 mL hexane and transfer to 250 mL separatory funnel, using

additional 50 mL of hexane to aid transfer.
Partition the hexane with three 25 mL portions of acetonitrile shaking 1 min. each time.

Combine acetonitrile extracts and discard hexane layer.

. Evaporate acetonitrile extracts to dryness with a jet of air and 50°C water bath, dissolve

residue in 2 mL acetone.

92

Protocol for fenvalerate:

1.

Add 200 mL of hexane:isopropanol (3:1) to 20.0g of peach fruit and blend for 3 min at
medium speed.

Filter using vacuum into a 250 mL separatory funnel.

Add 100 mL of water, shake carefully for l min. and discard lower, aqueous phase.

Wash hexane with 2 additional 100 mL volumes of water to remove all isopropanol,

measure and record volume of hexane.

. Column preparation: cover 6.0g of activated Florisil (overnight heating at 145°C) with

hexane, and add to a glass column using hexane to complete transfer. Tap column to
settle Florisil and add a 1cm layer of anhydrous sodium sulfate to the top of the
column and drain excess hexane keeping anhydrous sodium sulfate covered in
hexane.

Add 2.0 mL of the hexane extract to the column and drain, adding 5 mL additional

hexane to wash.

. Add 50 mL of hexane to column and discard hexane fraction.

. Add 50 mL of 5% ethyl acetate in hexane and save this fraction for analysis, evaporate

to <5 mL.

Protocol for chlorothalonil, iprodione and captan:

1.

Add 50 mL of acetonitrile to 20.0g of peach fruit and blend for 5 min at medium speed.

2. Filter using vacuum, add 10g of NaCl and shake vigorously for 1 min.
3.
4
5

Allow phases to separate; if no separation occurs, centrifuge at low g for 3 min.

. Aliquot 40mL of upper, acetonitrile phase to a ﬂask (recording exact volume of aliquot).

. Add 0.5 mL of toluene and percolate through a column containing 5g of anhydrous

sodium sulfate.

. Rotoevaporate to 0.5-1.0 mL — DO NOT DRY COMPLETELY!

. Filter into a vial and rinse ﬁlter twice with 0.5 mL acetone to complete transfer.

93

Protocol for benomyl:

l.

$99999.“

Add 25 mL of methanolzethyl acetate (1:1) to 20.0g of peach fruit and blend for 1 min at
low speed.

Filter using vacuum and wash the cake with additional 25 mL of methanolzethyl acetate.

Transfer the ﬁltrate to a 1L ﬂask containing 1.0 mL of 0.5N HCl.

Extract with 10 ml. of hexane and discard the upper hexane layer.

Adjust this solution to pH 7 .5—8.0 by addtion of 1N NaOH.

Extract twice with 100 mL portions of ethyl acetate and save the ethyl acetate extract.

Dry the ethyl acetate extracts by percolating through anhydrous sodium sulfate and then
wash the sodium sulfate with 50 mL of additional ethyl acetate.

. Rotoevaporate at 35°C to 4—5mL.

Protocol for sulfur:

. Prepare a dry, clean beaker and weigh accurately!
. Add 10 mL of carbon disulﬁde to 20.0g of blended peach fruit in a ﬁlter and mix

gently, catching the ﬁltrate in the weighed beaker..

. Wash with 3 additional 5 mL portions of carbon disulﬁde.
. Evaporate the carbon disulﬁde in an exhaust hood at room temperature.

. When carbon disulﬁde is completely evaporated, heat the beaker and residue for 15-20

min. at loo-105°C and weigh.

Subtract to determine the weight of elemental sulfur.

94
Protocol for sirnazine:

1. Weigh 20.0g wet soil into a 33x94mm Soxhlet extraction thimble and cover with glass
wool. Make a moisture content determination of a seaparate soil sample.

2. Place thimble of soil in Sohlet extractor with 150mL of 10% water in methanol and
reﬂux for 23 hours and then collect waterzmethanol int0250 mL ﬂask and
rotoevaporate to dryness.

3. Add 50 mL methylene chloride to dissolve residues and transfer to a 250 mL separatory
funnel.

4. Rinse ﬂask with 1015 mL of methylene chloride and 50 mL of water and add to
seaparatory funnel. Rinse a second time with 25 mL methylene chloride but keep
second rinse separate.

5. Shake separatory funnel vigorously for for l min., allow phases to separate completely
and drain methylene chloride into a 250 L ﬂask. Add second rinse to separatory
funnel, shake for 30 sec., collect and combine methylene chloride fractions and
rotoevaporate to dryness.

6. Redissolve in 2 mL isooctane and transfer through a glass funnel containing isooctane-
saturated glass wool to a preweighed 13x100mm glass, teﬂon-lined screw cap culture

tube. Repeat twice bringing total volume to 6-7 mL.

95

APPENDIX I

Data for 1993 Mite Population Counts

96

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97

APPENDIX K

Data for 1993 Chlorophyll Fluorescence Measurements of Field Study on Mites

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APPENDIX L

Effect of Mites on Shoot Growth and Flowering

Table 15. The effect of mites on 1993 shoot growth and 1994 ﬂowering and fruit set for

orchard trees infested in 1993.

 

 

 

Shoot Growth Flowers Fruit Set
ﬁgment (cm) (#) (%)
Control 22.85 62.0 16.0
Mites 22.12 60.1 17.3

NS NS NS

Treatment differences are not signiﬁcant at 10% probability level, NS, or signiﬁcant at the

probability level indicated.

100

APPENDIX M

Effect of terbacil on fruit yield and quality in 1993 and 1994

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102

APPENDIX N

Chlorophyll Data Collected in 1993 on Mite Treated Trees

103

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104

APPENDIX 0

Yield Data Collected in 1993 and 1994

Table 18. Yield Data collected in 1993 and 1994 from orchard sour cherry trees.

1993 Yield

I 11
84.43 50.91
1 14.48 30.37

III Average
101.72 79.02
90.31 78.38

Control
Terb l

Terb2

87.65

1994 Yield

Control
Mites

Control
Terb 1
Terb2
Terb3
Terb4

I
41.65
61.72

30.94
43.10

42.58
55.18

72.34

II
15.03
12.38

6.48
16.98
14.94
8.72
17.74

76.62

III
29.93
23.79

31.64
45.04
21.02
24.18
18.92

78.87

IV
50.34
35.55

42.32
24.66
62.96
46.64
28.08

Average

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33.36

27.85
32.45
32.97
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29.98

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Starch Data Collected in 1993 and 1994

105

APPENDIX P

Table 19. Starch data collected from orchard sour cherry trees in 1993 and 1994.

Fall Shoot Starch

1mm
'13
T1
'13
M1
M1
Tl

M2
T4
T3
C
T4

M3
M1
M2
T1
M1
M2
T4
M2
T2
M3
T1

C
T4
T2
M3
T2

0330

iﬁN: mu
9M% 1
3ﬂ% 2
8.95% 3
8%% 4
6m% 1
6.98% 2
5x% 3
8ﬂ% 4
8M% 1
6%% 2
9$% 3
7%% 4
6m% 1
9M% 2
7m% 3
6M% 4
8H% 1
95% 2
7M% 3
93% 4
7M% 2
6w% 3
8m% 4
6.93% 1
7%% 2
85% 3
6m% 4
8m% 1
9m% 2
8%% 3
10.07% 4

Spring Shoot Starch

mm

8.20%
6.95%
9.79%
7.50%
16.57%
10.50%
10.89%
9.68%
17.62%
8.92%
13.34%
9.83%
12.81%
14.56%
20.46%
13.26%
16.39%
7.59%
10.07%
1 1.91%
13.68%
7.46%
15.25%
11.50%
8.25%
13.27%
15.90%
15.37%
10.81%
14.67%
9.70%

Spring Shoot Starch
32m 22m

AmNHAwNH-hww#wNHAwNwamrdthv—‘th-HE

ﬁﬁﬁﬁddddﬁﬁﬁ

7.26%
3.51%
5.49%
6.81%
5.04%
5.71%
4.71%
3.67%
3.28%
2.18%
0.63%
2.01%
7.41%
0.82%
0.79%
6.07%
11.02%
5.11%
7.40%
4.40%
1.27%
1.51%
3.74%
7.01%
2.89%
6.27%
2.15%
10.85%
3.31%
3.22%
3.64%

106

APPENDIX Q

Data for Container Grown Mite Study

107

Table 20. Data collected from container grown control and mite trees in 1994.

RE ML
' 7'.

 

108

Table 20 (cont’d)

.4. o o .5 0"”

l 6

11111111

. i

1111

1

. 5.

l

7
5

1

1

 

109

Table 20 (cont’d)

 

110

Table 20 (cont’d)

 

111

APPENDIX R

Daily Minimum and Maximum Temperatures for 1993-1994 Dormant Period at Clarksville
Michigan

112

Figure 15. Daily Minimum and Maximum Temperatures for 1993-1994 Dormant Period at
Clarksville Michigan

113

vaBS.

_
_

 

vaRdxm

_
_

 

vakxd

_
‘

vath

_
d

M.M..
22......“

 

 

 

waxed: vaBE

_
d A

 

 

 

 

 

 

 

da\:\d_

d

‘i..__

 

daRdx: makx: maxw =2

_ _ _
_ _ A

38. 52
9:3. 582

 

 

 

 

 

 

od-

9.

9

Gd

cm

Figure 15

114

APPENDIX S

Air ﬂow generating fan and inlet setup for whole plant photosynthetic C02 assimilation

chambers

115

Figure 16. Air ﬂow generating fan and inlet setup for whole plant photosynthetic C02

assimilation chambers.

116

 

Figure 16

117

APPENDIX T

Schematic drawing of whole plant photosynthetic C02 assimilation chambers.

"11111111111111111111“