THE APPLICATION OF QNDEXING PROCEDURES EN THE CERTEFECATEON QF STONE FRUET NURSERY STOCK Thesis for flu Degree of M. S. MICHIGAN STATE COLLEGE Dean Frank Lavifl' W49 THESIS This is to certify that the thesis entitled THE APPLICATION OF INDEXING PROCEDURES IN THE CERTIFICATION OF STONE FRUIT NURSLR! STOCK. presented bl] Dean P. Lovitt ‘ has been accepted towards fulfillment of the requirements for MASTER OF SCIENCE— degree in _BLANI_PATHOLO}Y ”WW @aZZm Major professor Date “A! ’27, 1949 0-169 n—r-m—.._ .. ._ __ ' 561W ‘ .l 4‘ *{Kw’f ‘5 H; .. \\\.‘\ :F' S“)!- ‘3?“ 7. fix? \gI‘ *‘e 3. .3 IQ)???” «£3: " k M If." “a 3"! L; 'fiJ'é \) :35) £{-' ”..~I';;:; :.:.' 12% WW a; ’yifil'k-s . if ‘ ,.» -. "J." '4‘ ' 7:. ‘L I t; . ‘JHIH . I""\"*'. < :1“ a ‘ ' ' ‘0 15.9" h . L. 3 § ‘ - lat-‘1?" 'IM“. .' ‘- ' 3“, ' ‘r1\";1.".:|‘u ."",'15."nt . * . \ ’(' n ' I I. a' ‘ ‘.‘\\."u 1 ~.‘ ‘01-?1‘.\' '_ a 1,, ’ f‘ - .. . . 355‘ THE APPLICATION or INDEXING PROJEDURE‘S IN THE CERTIFICATION OF STONE FRUIT NURSERY STOCK By Dean Frank Lovitt A THESIS Submitted to the School or Greduete Studies of Michigan State College e: Agriculture and Applied Science in partial fulfillment of the reqnirements fior the degree of IMASTER.OF SCIENCE Department of Botany ' 191.9 MWIENI‘S The writer wishes to express his sincere thanks to Mr. C. A. Boyer and Professor Donald Cation for their help in making this thesis possible. Thanks are due to Mr. Boyer for his cooperation in permitting the writer to conduct these investigations as a part of his regular duties with the Bureau of Plant Industry, Mich- igan Department of Agriculture. Professor Cationis advice and criticism was invaluable throughout the course of these investi- gations . Thanks are also due to Dr. John Vaughn for his advice and criticism during the preparation or the nanuscript. 217808 TABLE OF CONTENTS Introduction Page 1 . Peach.yellows complex. 2 Rosett mosaic 2 Xsdisease 2 Peach necrotic spot 3 Green Ring yellows 3 Line pattern ‘ 3 Prune dwarf 4 Cherry yellows and necrotic ring spot 4 Indexing with sour cherry'as a host 7 Indexing with peach as a host 7 Indexing with flowering cherry as a host 8 Objectives 10 Materials and Methods: Experiment 1. 10 Results: Experiment 1. 10 Table l. 12 Photographs 13-18 Table 2. 18 DiscuSsion of results: Experiment 1. l9 llisterials and Methods: Experiment 2. 20 Mbntmorency selections indexed 20 Results: Experiment 2. 22 Table 3 24 Table A 38 Discussion of results: Experiment 2. 39 Summary and conclusions 42 Bibliography #4 INTRODUCTION The virus diseases of stone fruits in Michigan are more or less easily transmitted by budding or grafting and it is reasonable to assume that the nurseryman could unwittingly propagate the diseases in the nur- sery through the use of infected scionwood. The rather long incubation periods of most of the viruses within the hosts, seasonal variations in symptom expression and complete masking of symptoms in some hosts or the lack of symptoms in others at the time sciomvood is selected make unin- tentional propagation of these diseases possible. In an attempt to pre- vent the introduction of the diseases into nursery stock through the use of contammated scionwood, a plan for the certification of stone fruits in Michigan was established in 191.0. This plan has been under the ad- ministration of the State Department of Agriculture in cOOperation with Michigan State College. Briefly, this program provided for the inspec- tion of parent scionwood trees and surrounding trees and orchards during that season of the year most favorable for the expression of symptoms. The first inspection was to be made during the season that budwood was cut from the parent tree and inspections made during two successive sea- sons thereafter. Then, provided the parent tree did not express symptoms during that time, the nurseryman was given a special certificate under which the progexw could be marketed. A number of virus diseases are known to be present in the stone fruit orchards of the state. Those diseases of peach found in the state 111ch rosette mosaic, X—disease, necrotic spot and the diseases of the peach yelJm complex (peach yellows, little peach and red suture). Swoet and sour cherry trees are known to be infected with the viruses causing cherry yellows, necrotic ring spot, green ring yellows and I—disease. Plum may be hosts for cherry yellows and necrotic ring spot (28) and those diseases of the peach yellows complex. The line pattern virus is also known to be present in plums. Prune dwarf, although not reported in the state may be present in latent form in sour cherry, sweet cherry and pllm. Those virus diseases of the peach yellows complex have been known and studied for many years (1) (ll) (12) as serious diseases of peach. At the present time the peachflyellews portion of the complex is almost non-existent in the state, being far outranked in importance by little peach and red suture. 0f the latter, little peach is the more preva- lent. Commercial plantings of plum may be serving as hosts of the yellows complex in some areas . The visual inspection system for certi- fication is thought to be adequate for these diseases but the applica- tion of heat treatment of sciomvood as described by Kunkel (25) is under consideration as a more positive means of control. P If Rosette mosaic of peach, first described by Cation(9) in 1933 is found occasionally in the orchards of Southwestern Michigan and although, a very damaging disease to the individual tree, spread from tree to tree appears to be negligible. Since the disease is not widespread and be- cause symptoms are strikingly evident at the time of the selection of scionwood, visual inspection is thought to suffice for certification purposes. ’ I-disease is found on peach and chokecherry in Macomb and Iapeer counties and on chokecherry in other sections of the state but has not been found on peach in the major peach producing counties. The disease was first described by Stch (37) in Connecticut in 1931. and later was found to be a morpeach disease in Massachusetts (5) and New York 3 (22). Cation (8) reported the disease in Michigan on peach and choke- cherry in 191.1. rRecently, sour cherry was found to be a host of 1:- disease in New York (30). The disease causes a serious wilt and decline of sour cherry and was found on Montmorency in one orctnrd in Macomb County during 1948 by Cation and Levitt (14). A similar disease of peach (Western X-disease) (31s) is found in Western United States and British Columbia, and is-assOCiated with the cherry diseases Idiom as Buckskin (33), little cherry (35) and wilt and decline (36). The x- disease in Michigan does not present a serious problem of Certification in that the virus is difficult to transmit by budding and its effect upon sour cherry would not make such trees choices for the selection of propagation wood. If necessary the sciomtood could be subjected to heat treatments (38) similar to that for peach yellows. Peach necrotic spot reported by Cation (7) in 1942 was discovered as a latent virus in Windsor cherry. Subsequent experiments resulted in isolation of the virus from symptomless Windsor, Napoleon and Bing cherry trees. The disease has not been observed as occuring in the commercial peach orchards of the state. Green ring yellows was first observed by Rasmussen (31) in 1937. The disease has been found to be of ndnor importance in several orchards of the state. Innoculations to peach and sour cherry have produced a shock reaction similar to that of necrotic ring spot, although chronic symptoms on sour cherry do not suggest the presence of ring spot. Certi- fication of cherry scionwood for this disease could be accomplished by indeadng on peach but because lmotm cases of infection are confined to a few orChards, visual impaction has been depended upon as a means of certification. The line pattern virus known to be present in plums is apparently l, of minor importance. The disease has been reported by several investi- gators and variously named. Valleau (1.1) was probably the first to ob— serve the line pattern virus, and a similar disease was described later by Thomas and Rawlins as Vacaville plum msaic (1+0). Cation (6) de- scribed the disease on peach as isolated from Red June and Abundance plums in Michigan, while Berkley (2) and Willison (1.3) have investigated a Shiro plum line pattern mosaic in Ontario. Zeller and Milbrath (1.1,) have de- scribed a similar virus from flowering cherry. Cation, Berkley, Milbrath and Zeller (13) now believe they were dealing with an identical virus. Prune dwarf (39) is known only to affect Italian prune seriously. The disease may be latent in most varieties of plums grown commercially and has been found on sour and sweet cherries in New York (20) and on sweet cherry in Ontario (42). Because the disease has been found on these various hosts in New York and Ontario, it is likely that it may occur in latent form in Michigan. Certification of cherry and plum for freedom from this disease would involve innoculations to Italian prune. Although it produces syniptoms on peach, reaction to some strains are unreliable. Cherry yellowe and necrotic ring spot are a challenge to the method of certification by visual inspection. Cherry yellows was demonstrated to be a bud-transmissible disease of sour cherry by Keitt and Clayton (23) in 1939. Since that time the disease has become recOgnized as one of the, most serious virus diseases of sour cherry. Necrotic ring spot was first reported from peach by Cochran and Hutchins (17) whose investigations in— dicated the disease to be widelyr dispersed in the stone fruits of Califor- nia. Chamberlain, Willison, and Berkley (15) described ring spot from sour cherry after its discovery in Ontario in 1939. At about the same time, the disease was described on sour cherry by other investigators (21). There are certain known factors concerning these two virus diseases that an. certification by visual inspection inadequate. The more pertinent of these factors are given below. ‘ 1. Cherry yellows as described by Berkley and Willison (A) has two phases of symptom expression following infection. An acutephase first occurs as growth is initiated in the spling and is characterised by a delay in foliation followed by the appearance of etched rings on the newly formed leaves. Later these areas become chlorotic , necrotic, and drop out , giving the leaves a ragged or shot-holed appearance. This phase is not repeated during the remainder of the life of the tree except for the occasional appearance of some translucent rings on leaves early in the season. The acute phase is followed by a chronic phase of yellowing of the leaf tissue, generally between the veins, and the casting of those leaves. Sometimes green leaves are cast. This phase persists season after season depending upon the spring temperatures. The acute phase is believed to be the necrotic ring spot entity of a virus complex as the reaction of host plants to necrotic ring spot virus alone is apparently indistinguishable from this phase. Since, the acute phase of cherry yellows or necrotic ring spot in itself become latent after the "shock" symptoms described above it can be readily realised that visual inspec-l tion is not an adequate means of diagnosis. 2. Both cherry yellows and necrotic ring spot viruses have been shown to be latent in sweet cherry and in most varieties of plums (28). Therefore, it is impossible to adequately certify these species by visual inspection. 3. Kcitt and Moore (21.) have shown that night temperatures above 60°F. are not conducive to symptom expression of cherry yellows on sour ca cherry. Mills (27) has noted that mean temperatures above 61°F. follow- ing petal fall On Montmorency cherry reduced the amount of symptom ex- pression in a given season. Thus, during seasons of warm spring tempera- tures following the petal fall period, accurate diagnosis for cherry yellows by visual examination is not possible. It. Symptom expression of cherry yellows is of short duration even in favorable seasons. These symptoms occur four to six weeks after petal fall and may last for two weeks. Some symptoms have been observed later in the season but these are unreliable diagnostically. Such a short period of symptom expression has created a problem in completing all necessary inspections in the state. 5. Originally budwood from parent trees was increased by budding from the progeny, or row to row propagation. However, Cochran (16) has proved that necrotic ring spot could be transmitted through Mazzard seeds and Cation (10) has shown that both the cherry yellows virus and the ne- crotic ring! spot virus may be transmitted through the seed Of m M understocks. Field observation has pointed to possible contamina- tionin conmercial sources of these rootstocks. Thus, the practice of row to row budding is considered unsafe for maintaining disease-free parent scionwood. 6. The incubation period of most of the stone fruit viruses pre— sent in Michigan may be as long as two years. This makes certification by visual inspection an unwieldy procedure and any process to accelerate the investigation of scionwood sources would be of value. The above factors make adequate diagnosis of these diseases of sour cherry, sweet cherry and plums the most pressing problem in scionwood certification at present. As a result, investigators in several sec- 7 tions of this country and Canada have been directing their efforts to- ward the development of reliable and expedient means of diagnosis for the cherry yellows and necrotic ring spot viruses. These efforts have generally been focused on the development of indexing techniques on several host plants, and have crystalized into indexing systems using peach, sour cherry, and Japanese flowering cherry as hosts. A brief smmnary of these three systems follows: Indexing with Peach as a Host (3) (19) (32): 1 Seedling peach or most clonslvarieties innoculated in August with buds carrying the necrotic ring spot virus show symptoms Of infection the following spring. First, a delay in foliation is evident, and as growth is initiated necrosis of new shoots occurs. Innoculation with the cherry yellows virus first produces these acute symptoms followed by a shortening of the internodes of new growth, in many cases to the point of forming loose rosettes. The leaves of this growth are usually darker green than normal. Symptoms are followed by recovery as the growing season progresses and those of ring spot do not recur during successive seasons; however, chronic symptoms of cherry yellows do reappear. Indexing with Sour Cherry as a Host (29): I Sour cherry is being used as an indexing host in the greenhouse where controlled temperatures my be maintained. Two techniques have been developed, namely, a direct method and an indirect method. Direct Method: Disease-free trees are innoculated at bud-break with buds from those trees under investigation. The budded trees are then held at a temperature of 70°F. for a period of three to four weeks. Should the innoculated trees fail to show symptoms of necrotic ring spot within a reasonable time the trees in question are considered to be free of ring spot and of cherry yellows as the former virus always appears to occur with the latter. To determine whether the trees in ques- tion are actually'infected‘with cherry yellows it is necessary to maintain the host trees at temperatures below 60°F. fer six to seven weeks. If positive symptoms of cherry yellows are not noted the first year the trees I should be carried over and observed the succeeding year. Indirect Method: This method has been developed on the premise that the necrotic ring spot virus and the necrotic ring spot component of cherry yellows immunizes against itself, i.e. once the virus becomes latent in the host, reinoculation will not produce the acute shock symptoms of in, itial infection. The trees under investigation are brought into the green, house and while dormant all desireable scionwood is removed and placed in cold storage to be used for propagating should the parent tree prove to be disease-free. The parent tree is then inoculated at budébreak'with a known source of ring spot and held at a temperature of 70°F. The develop- ment of ring spot symptoms on the inoculated tree indicates that the tree ‘was not infected.with ring spot or cherry yellows previously. The dorb mant budwood that had been removed and stored is then used to increase the disease-free strain. The resulting trees will be disease-free pro- vided that uncontaminated understocks are used. Indexing with Flowering Cherry as a Host (26): The varieties, Kwanzan and Shirofugan are sensitive reactors to many Of the latent viruses including the necrotic ring spot virus. Several trees may be indexed on Shirofugan at one time and individual readings obtained because it is characteristic that these viruses be- come systemic very slowly in trees of the variety. Infected buds fail to unite, soon die and within a period of two to three months gumosis occurs in the vicinity of the inserted bud. For mass-indexings, buds may be placed as close as four inches apart on a limb. After readings are made the limb may be cut off below the buds and the tree used again as a host. Trees indexing negative on Shirofugan are usually reindexed on Kwanzan since there apparently are latent viruses that react on the latter that do not react well on Shirofugan and vice-versa. The viruses become rapidly systemic in Kwanzan and therefore a tree my be used for only a single indexing. . . All three of these hosts or techniques have their advantages and disadvantages, the choice for practical application in the certification of nursery stock depending upon the facilities and materials available, and upon climate if indexing is to be done out-of—doors. The techniques using sour cherry as a host require suitable greenhouse facilities which have not been available for the mass indeidngs necessary to certify scion- wood on a commercial scale in Michigan. Japanese flowering cherry trees have not been available in quantity and the difficulties encountered in propagating them makes this host somewhat expensive as compared with peach seedlings or even propagated clonal varieties. Both seedling peach and some clonal varieties, of peach have been previously tested for re- action to the viruses herein concerned. The varieties Elberta and Ro- chester (3) have been recommended as outstanding varieties and seedling peach has reacted favorably. Since seedling peach trees are less expen- sive than clones they seem the natural choice for certification purposes in Michigan. Greenhouse facilities are not necessary, in fact reactions of peach to cherry yellows and ring spot are more favorable out-of-doors than in the greenhouse. 10 OBJECTIVES The experiments described in this thesis were designed to test the application of peach indexing to the certification of cherry and plum nursery stock in Michigan. Experiment 1. is concerned with testing eleven representative clonal varieties of peach for comparative reaction to cherry yellows to determine whether any were so superior as indexing hosts as to warrant the additional expense of using prepagated trees in- stead of seedlings. Experiment 2. was conducted to test peach seedlings as possible hosts for mass indeedngs of cherries and plums under typical nursery management conditions. This experiment was carried out at a comercial nursery for the purpose of selecting parent scionwood trees free from cherry yellows; the selected trees to be planted out in an orchard and maintained as a scionwood source. MATERIAIS AND METIDDS: EXPERIMENT 1. Five one year nursery trees. ofeach of the varieties Halehaven, Red Haven, 0!Boy, Elberta, Rochester, Red Elberta, Fertile Hale, J. H. Hale, Golden Jubilee, Amber Gem, and Nectarine Special. were inoculated with a strain of cherry yellows on September 12, 191.6. These trees were dug later in the fall along with five adjacent uninoculated trees of each variety and all were heeled in out-of-doors over winter. Early in the following spring of 1947 the trees were set out in a single row with the ten trees of each variety planted as a separate unit and the inoculated trees alternated in the row. RESULTS: EXPERDENI‘ l. Readings for varietal reaction to the cherry yellows virus were made on June 8 and June 21, 191.7, and on June 11., 191.8. The photographs following Table l. were taken on these dates and show representative in- ll. oculated and uninoculated trees of each variety (except for a photo- graph of Elmerta, June 8, 191.8). In the majority of cases the same trees were photographed on all three ”occasions. The severity of symptoms expressed during 1947 were very similar within each variety, thus data concerning varietal reaction to the virus was confined to rating the individual varieties on the basis of severity of symptom expression in the following three arbitrary categories: 1. A severe reaction to both phases of the virus shown by a considerable amount of die-back, delayed foliation and particularly, extreme rosett- ing of new growth. 9 2. A less severe reaction, moderate die-back, delayed foliation and less severe shortening of internodes of the new growth. 3. A mild reaction particularly, as to the chronic phase of the virus; the inoculated trees somewhat difficult to distinguish from those not inoculated. As can be seen from the photographs there were gradations between three such categories but more specific. analysis did not appear to be within the scape of this investigation. Readings of second year symptoms (June 11., 19148), were made of each individual tree of the varieties as symptoms were not so striking and inoculated trees of some varieties showed few or no symptoms. Thus cate- gorical evaluation similar to the previous season! s readings was thought to be more accurately made by comparisons based on readings of indivi- dual trees rather than by field observation alone. The individual read- ings are given in Table 1. It should be remembered that five of the trees represented in each variety as being symptomless are uninoculated. Table 2 shows the ratings given each variety for each of the three readings-made. 12. .Usov 095 A3 «mafia mlpoppfinm 65 «983?? now. offinmo: Alv «95994.er .Hom o>3wmom A5 eeopwadoogfiod A3 «Umpdgoozcfl A5 "magma 93 Qfi USoom one maoaim mrnfisoaag one aux. .mde «j 05:. some mwfiwsofi .x. :ADVQHH unavmoa uApvooa :Apvoma ufisvoma oszoeH umavaefl ufipvona umavaaa [szwwa unavona sAHVmHH xAvaoH :Avao -AvamH _xAvamH :AvarH umsvaoa oAvamH nAvaaH -AvamH uavaoH :Apvaaa :Asvooa ufipvem ufipvmma aasvama nlavana xAvaoH umavmma unpveaa ufipvooa Inpvoom omHveHH mfiHveoa sanveo .sxAHVemH.sxAHvrmH :AHverH sxAanoH anvrma sxfiHvraH xAanwH sAHVan [Anvoma uAavaoa unavmo ufisvama ulpvooa IADVOmH unsvmoa uAsvomH nfiavmaa umavmma [Amvooa xAHVmHH omvaoa uavam onvaNH sAvamH uAvanH :AHVMbH_sxAHmeH.axAvaaH _xava H sxAvaoH , _ IADVOHH sfisvmoa -Aavao uApvomH unavmma oApvmnH uApvoaa ufiavoma unavoaa unavoma unavaoa oAvaHH oAHVmoH IAvao xAHvNNH.sxAvamH :AvasH oAvaoH _xAvamH.sxAvaaH sxfivamH uavaoH umavmaa ofiavoaa sfisvoo [Asvomfl -ADVomH nasvoea ufiavoca nasvama unavoua [Aavama rapvmea xAHVHHH :AHVHOH oAHvHo .axAHvHNH sxAHvaH eAHVHnH :AHVHrH _xAHvaH_sxAHVHaH umHvaaermAHvaa dfionam Hofloogm QmKESmHflH can: Lnommo moafladw odd: dfionfim E00 993930.: Cobfll mcnmhwpooz .: .H. GOUHOU magnetism mom 908$ mom .x. .99? misoaaow nflsoso 0....3. op cogoson pow moflpoflpwir meson gonads Ho moopp ngflimvmfl 93 .Ho owned mmmflveox .H «Vanda. 13. ma 3% as s see .15 .1' V .v. ....r .5 . . .. .c , a «. . I .. than: .. . .. . h»; . . r Me). a ‘ O gasses SS :8 SE. . . .. . \O .x, q. . .I...fl.yb.r. . sh... gang . .32 .H ssh ea. .1) ‘ -e . .lW‘. dung 580mm Sm." «AN on: e«‘ '- 2833 E . ER.” «AN 33. -.. -. 4.38%“... .. 16. -, . e ..1 fidcrq‘e'fe mmnHm=e_zanuoo puma tam ones _ a. .. .- sunfish.- a area .m ones y . 3m...” :2” 08:. 4m h ....-a. ._ in: r . - _ So 0 .u.v.dfle\1).ll- .w .... ..’...x'~. . I! . a, . . «E a SS .5“ 8.6 g .m .e. H .Hu 8.... t‘l Table 2. Categorical ratings as to severity of symptoms expressed by . each of eleven clonal varieties of peach tested for use as indexing hosts for the cherry yellows virus. Red Haven 2* 3 3 Rochester l l 3 Amber Gem 2 3 3 Red Elberta 2 2 1 Fertile Hale 3 3 3 Golden Jubilee 3 3 3 Way 3 3 3 J .. H. Hale 2 l 2 Halehaven 2 2 3 Nectarine Special 1 l 1 Elberta 2 2 1 * The categories represented by numbers are as follows: 1. severe reaction, 2. moderately severe reaction, 3 . mild reaction to the point of being indiscernible. e .2. “Eu E‘I O ' ' ’ y . 1,. o . 19. DISCUSSION OF RESUIES: EXPERDENI‘ l. The observations made of the indeccings in this eacperiment and the data presented reveal that for all practical purposes six of the clonal varieties of peach showed little variation in reaction to the cherry yellows virus. From averaging the category ratings in Table 2, the varieties may be grouped in the following manner beginning with the most desirable variety for symptom expression. 1. Nectarine Special 2. Rochester, Elberta, J. H. Hale, Red Elberta, Amber Gem. 3. Red Haven, O!Boy. I... Halehafiren. A 5. Fertile Hale, Golden Jubilee. The variety Nectarine Special was observed to be most outstanding for symptom expression, while Rochester was a close second. The varie- ties Elberta, Red Elberta, J. H. Hale, and Amber Gem were satisfactory host varieties but not as outstanding as the first two mentioned. Red Haven, O!Boy and Halehaven probably would react favorably in most cases encountered in indenting. Golden Jubilee did not prove to be of value as an indexing host and the use of the variety is not recommended. The Fertile Hale variety failed to show severe symptoms of the chronic phase of cherry yellows but striking symptoms of the necrotic ring spot phase were expressed in the manner of very obvious chlorotic rings on the leaves of this variety. Such symptoms were few or lacking in the other varieties. Thus the variety may have some value as an indexing host for the ring spot virus. However, it is possible that this attribute would not remain constant under all seasonal variations that occur from year to year. 20. MATERIALS AND METHODS: EXPERIMENT 2. In this experiment 500 one year Montmorency trees propagated from eleven different selections of budwood were indexed as well as 1.25 one year plum trees of the varieties Stanley Prune, Shr0pshire Damson and Blue Damson. In addition, one year trees of two met cherry varieties were indexed, namely, 125 trees of the Windsor var- iety and 121. trees of the Schmidt variety. An attempt was made to select trees of uniform size, or trees approximately seven-sixteenths inches in caliper. Therefore, successive trees in the nursery row were not indexed. Each of these trees was indexed in triplicate on three seedling peach trees in the nursery!s propagation block. The buds were inserted in late August and early September of 191.7. Both inoculated and uninoculated check trees were left throughout the in- dancing for convenience in making a comparative diagnosis and also to allow for any difference in reaction due to variable vigor of the seed- lings. The conventional shield bud was used, the buds being held in place by. wrapping with the rubber bands commonly used in commercial propagation. MDNTEORENCY SELECTIONS INDEEED ' Considerable attention is being given by the research and regula- tory personnel of several states to the develOpment of sources of disease-free Montmorency budwood, particularly because of the serious reduction in fruit yield as the result of infection with cherry yellows. The following comments on the various selections of Montmorency cherry used in experiment 2 are presented as a background for evaluating the data obtained. Montmorency Knuth: Eight trees were selected from an old cherry 21. orchard on the Robert Knuth farm in Berrien County in 191.6. Although, some 30% of the trees were showing visual symptoms of cherry yellows at the time of selection, symptoms were not observed on these eight trees. The nursery firm wished to attempt selection of bud-wood free of cherry yellows from this orchard because of its past production record. Therefore, it was nnrtually agreed that after the visual in- spection each tree would be indezmd on seedling peach and further, that the progeny of any tree indenting positive for cherry yellows would be destroyed. Readings made of these indexings during the spring of 191.7 revealed thathvro of the selected trees were infected with cherry yellows 3 however, through an oversight the progetv of one of these trees (Knuth 2-1) were not rogued from the nursery row. Eleven trees of Knuth 2-l'were included in the 191.8 indexing for the purpose of rechecking the previous years reading. Montnnrency 1.65, 297, 261.: These numbers represent individual Montmorency strains selected by the Horticulture Department at Michigan State College (18). Budwood for prOpagating the indexed trees was obtained directly‘from the college scionwood orchard. Despite known infection with necrotic ring spot these strains are outstanding in pro- duction and the scionwood source trees free of cherry yellows. Montmorency Wendzel: The indend tmes were the progeny of three year trees located in the Henry Wendsel orchard, Berrien County. This orchard was visually inspected in 191.6 and approved as a source of. Montmorency budwood. The source of the orchard trees has not been determined. Visual observations in this orchard were impossible dur- ing 191:7 because of a severe infection of cherry leaf spot but obser— vations during 191.8 revealed one tree showing positive symptoms of 22. cherry yellows and two trees which were questionable. However, these trees were not in the area of the orchard from which budwood was selected in 191.6. ms: WENT 2. Three readings of the. indexings were made during the spring of 191.8. As indicated by Table 3 these readings were made on May 13, May 20, and June 2. The May. 13 reading was made specifically for necrotic ring spot as growth. had not developed sufficiently to diagnose symptoms of cherry yellowe accurately. The succeeding readings were made for diagnosis of the chronic phase of cherry yellows. The determinations of May 13 were made under some difficulty because of an excessive amount of killing- back by winter temperatures. Winter injury plus die-back due to the ring spot factor killed peach seedlings back to within four to six inches of the ground line in many cases. The presence of inoculated and un- inoculated checks mde it possible to differentiate between ring spot dieme and winter injury. Table 3 presents the data obtained for the three readings of each individual indexing . Following this a summary has been prepared (Table I.) to show the total number of trees of each selection indexing positive for necrotic ring spot and cherry yellows. Data is also presented in this table as to the number of cases where buds from a selected tree produced the same effect on all three of the peach seedlings budded. In all readings a dead seedling was regarded with suspicion except in instances where the other two seedlings of the triplicate did not show symptoms. Readings of individual seedlings are noted by a question mark where a slight deviation from normal growth was observed but de- finite symptoms were not demonstrated. In final analysis, indexings 23. showing one or more seedlings with positive symptoms of cherry yellows on May 20 or June 2 and those indexings regarded as questionable were considered sufficient evidence for discarding the cherry or plum trees represented. .--Percentages of infection of necrotic ring spot and of cherry yellows given in Table 1. include the trees indexing "quest- ionable" . 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DISCUSSION OF RESULTS: EKPERDIENI‘ 2. The severalobjectives of this eccperiment were as a whole concerned with testing the application of the peach indexing technique to the com- mercial certification of cherry and plum scionwood. The principal ob- jectives were; (1) to determine whether seedling peach would react con- sistently when used as an indexing host; (2) to develop a system for in- dexing that would not seriously interfere with conventional nursery practices; (3) to find improvements for the certification of sour cherry, sweet cherry and plum; (II) to initiate the idea of a certified scionwood orchard, thereby eliminating possible perpetuation of rootstock contamina- tion through row to row budding. The reaction of the peach seedlings to ring spot and cherry yellows or the absence of reaction in cases of negative indexings was constant. This may be noted by comparison of the number of trees indexed with the total number of cases where trees of a triplicate reacted alike. Totals in Table 3 indicate that of 874 indexings the seedlings of a triplicate reacted alike in 850 on May 13, in 865 on May 20, and in 867 on June 2. On a percentage basis the seedlings of a triplicate reacted alike in 98.5% of the indeflngs. This would indicate that resistance to these vims diseases is not often found in seedling peach and that indexing in triplicate would make accurate conclusions possible. Duplicate indesc- ings do not seem as satisfactory since the death or subnormal growth of one seedling would cause the validity of an apparently symptomless com- panion seedling to be doubted. There were some instances in this ex- periment, mwever, that lead to speclfletion concerning possible re- sistance in some seedlings. For instance, 35 of the 36 peach seedlings inoculated from the progerw of Montmorency Knuth 2-1 gave reactions for 40. both viruses, the one exception occuring with one seedling in the indexr ing of tree No. 365. This one seedling was outstanding in its freedom from visual symptoms. is all. three inoculation buds of No. 365 came from the same budstick there seems to be but two possible explanations for the failure of the one seedling to show symptoms. One, the bud might possibly have failed to unite and transmit the disease or two, the seed, ling may have been resistant to infection. To check these possibilities the peach seedling was removed from.the field in the fall of 1948, potted and heeled in out-ofbdoors until January'1949. At that time in- oculations were made on peach in the greenhouse. The results were inp conclusive. Positive symptoms of transmission of a virus have not been observed to date on the inoculated trees nor on several control trees inr oculated with known ring spot, however, the seedling itself has failed to grow normally. In fact, terminal growth has been subnormal and the leaves have a dwarfed chlorotic appearance. A more thorough investi- gation is planned to determine the status of this seedling. There were no other cases where three seedlings of an indexing gave such a differen- tial reaction.throughout the three readings. An.accurate explanation cannot be given.for two apparently disease-free trees among the progeny of Knuth 2-21 since the other 55 trees of this selection indexed, "posi- II..», for ring spot. It may be possible that the ring spot virus‘had not become completely systemic in the parent tree at the time of bud selection or that the same might have been true in the two progeny trees indexed. Reindexing of these two trees will be necessary for more accue rate determination. There were a number of cases where the triplicate failed to give a constant reaction for the individual readings. Meet of these cases occured in the readings for ring spot symptoms and as pre- Al. viously mentioned, severe winter die-back made this reading somewhat difficult. The two readings for cherry yellows conflicted in some cases but if either or both readings indicated at least one diseased seedling in the triplicate or two dead seedlings or two questionable reactors, the cherry or plum tree represented was discarded as undesir- able for‘ scionwood. Those trees indexing, ”positive", for necrotic ring spot alone, were not discriminated against as conclusive evidence has not yet been presented to show effects from the virus to be of serious commercial significance. However, the nurseryman is urged to select scionwood from those trees indexing free of the virus insofar as practicable. The Montmorency selections rather consistently indexed positive for ring spot except for the Wendzel selection of 220 trees. Thirty six of these trees gave positive reactions for ring spot infection, four of which were also shown to have cherry yellows, or on a percent- age basis a cherry yellows infection of 1.8% and a total virus infection of 16.1%. Successive trees were not indexed, thus, these percentages should not be considered accurate representations of the actual virus contamination of this selection nor should the other percentages given in Table 2 be regarded as such for the other selections. The Schmidt variety, showed considerably less virus contamination than Windsor, especially in the amount of ring spot infection. The three plum varie- ties were shown to be entirely free of infection of either virus. Although data is not presented, it was interesting to note the variation in mortality of inserted cherry buds on peach. Since it was known that cherry and peach are not satisfactorily compatible species, a rather high mortality of buds was expected. However, it was defini- 1+2. tely demonstrated that a measure of incompatability is brought about by the presence of the ring spot virus. It could be readily observed that an appreciably greater number of buds from ring spot-free selec- tions initiated growth on peach than did those from diseased selections. SUMMARY AND COMLIISIONS Two experiments have been conducted, the first, a test of varietal reactions of peach clones to the cherry yellows virus. The second. experiment was a test of the use of seedling peach as a host for com- mercial certification of sour cherry, sweet cherry and plum. The in- deodngs were made for the purpose of obtaining foundation scionwood stock free of the cherry yellows virus. A test of eleven peach varieties was made to determine their re- lative values as indexing hosts for the cherry yellows virus. The results obtained may be best summarized by the following list of the varieties in order of decreasing desireability as indexing hosts. 1. Nectarine Special 2. Rochester, ‘Elberta, J. H. Hale, Red Elberta and Amber Gem 3. Red Haven, 'O!Boy ' ' 1.. Halehaven J' 5. Fertile Hale, Golden Jubilee The last named varieties, Fertile Hale and Golden Jubilee were found to be undesireable as indexing hosts. Nectarine Special proved to be the better of the eleven varieties but Rochester would be nearly as good a choice. Peach seedlings were used in indexing 87h one year trees of Mont- morency, Schmidt and Windsor cherries and Stanley Prune, Blue Damson #3- _and Shropshire Damson plums. The trees were selected from the stock- trees of a Michigan nursery. The indexing was carried out on the nur- sery premises for the purpose of; l. selecting foundation scionwood trees free of the cherry yellows virus; 2. determining whether index- ing proceedures would have such application; 3. observing whether seed- ling peach would give a constant reaction to various sources of the virus; and A. establishing a scionwood orchard as a source of certified budwood for use by the mlrsery, thereby eliminating perpetuation of possible rootstock borne viruses which was not possible under the con- ventional row to row propagation system. Although the necrotic ring spot virus is not being discriminated against in the present certification of stone fruit nursery stock in Michigan, readings were made of the indexings for this virus as well as for the cherry yellows virus. Each selected tree was indexed in triplicate on three peach seed- lings. and the results show that in 98.5% of the indexings the seedlings of the triplicate gave the same readings. The majority of the trees indexed were Montmorency because of its commercial importance and be- cause of the serious reduction of crop in this variety when infected with cherry yellows. The results of these experiments indicate that either seedling peach or clonal varieties of peach are useful tools for the certifica- tion of sour cherry, sweet cherry and plums as far as necrotic ring spot and cherry yellows are concerned. Indexing on this host can be adapted to certification on a commercial scale with a minimum of inter- ference with nursery management practices and at a nominal cost. Tests of reactions of clonal varieties show a varietal difference in severity id engllwg‘hifi alum! AA. of symptom expression. The varieties, Golden Jubilee and Fertile Hale were shown to be unreliable as indefing hosts. Those varieties ”verily effected by the viruses such as Nectarine Special, should be useful in experimental indexings but as seedlings have been shown to be satisfactory hosts, the matter of cost would mks the latter a choice for commercial certification ventures. The indexing of one or two year trees of the particular strains and varieties of sour cherry, sweet cherry and plum grown by the individual nursery is entirely feasible as a means of providing foundation stock to be used as a source of disease-free budwood. The maintainence of this indexed stock in a permanent bud orchard would avoid increasing any virus incident in understocks. Occassional reindexing of this foundation stock would be desireable to eliminate trees which might possibly be- come infected through insect transmission. Frequent spraying with BK), DDT, Parathion or similar insecticides would also be desireable. The bud orchard should be given the best isolation practicable from uncertified nursery stock, orchards and possibly wild prunus species. BIBLIOGRAPHY 1. Bennett, C. W. Peach yellows and little peach situation in Michi- gang. Ann. Rept. State Hort. Soc. , Michigan. 56: 187-191;. 192 2. Berkley, G. H. Prune dwarf and Shire line pattern Mosaic: two new ‘ virus diseases in Ontario. Publ. No. 679 of the Div. of Bot. and Plant Path. Sci. Serv. Dom. Dept. of Agr., Ottawa, Canada. (Repr. for Canadian Hort. and Home Mag. , Oshawa, Ontario) 2 pp. illus. l9hl. 3. , Cherry yellows and necrotic ring spot of sour cherry in . Ontario. No. l The value of Prunus persica and E. domestics \(rar. Italian prune as index hosts. Phytopath. 37: 2-3, 191.7. Abst. A5. I... , and R. S. Willison. Yellows and necrotic ring spot of sour cherries in Ontario: Inoculation experiments. Phyto- path. 38: 509-518. 191.8. 5. Boyd, 0. C. X-disease of peach found in Massachusetts. P1. Dis. _ Reptr. 25: 406-407. 1941. 6. Cation, D. The line pattern virosis of the genus Prunus. Phyto- path. 31: 1004-1010. 1914.1. 7. , Necrotic spot, a peach disease transmissible by budding. . Phytopath. 32: 2. 191.2 (Abst.) 8. , “X"-disease of peach and chokecherry found in Michigan. ., . Pl. Dis. Reptr. 25: h06-hO7.-1941. 9. , An infectious rosette of peach trees. Mich. Agr. Expt. ~Sta. Quart. Bul. 16: 79-84. 1933. 10. , Transmission of cherry yellows virus complex through seeds. Phytopath. 39: 37-AO. 1949. ll. , Three virus diseases of the peach in Michigan. Mich. 12. , A. B. Groves, T. F. Manna, H. H. Thornberry. Peach ' yellows. pp. 7-9..In E. M. Hildebrand, G. H. Belliey, and D. Cation, handbook of virus diseases of stone fruits in North America. Mich. Agr. Expt. Sta. Misc. Puhl. 191.2. 13. , G. H. Berkley, J. A. Milbrath, S. M. Zeller. Line Pattern. - In handbook of virus diseases of stone fruits in North Am— erica. 2nd. Ed. - In press. 14. _ , and D. F. Lovitt. Unpublished data. 15. Chamberlain, G. C, R. S. Willison and G. H. Berkley. Necrotic ring spot (Canadian Type). pp. 63—61.. In Hildebrand,E.M., and G. H. Berkley, and D. Cation. Handbook of virus diseases of stone fruits in North America. Mich. Agr. Expt. Sta. Misc. Publ. 19h2. 16. Cochran, L. C. , Passage of the ring spot virus through "Mazzard cherry seeds”. Science (N.S.) 104: 269-270. 191.6. 17. , and L. M.‘Htrtchins. A severe ring spot virosis of peach. ...... Phytopath. 31: 860. 191.1. (Abst.) 18. Gardner, V. R. and W. Toenjes - Strain differences in the Mont- morency cherry. Mich. Agr. Expt. Sta. Quart. Bul. 31 (1): 83-90. l9h8. l9. Hildebrand, E. M. Indexing cherry yellows on peach. Phytopath. 32: 712-719. 191:2- 20. 25. 26. 27. 29. 300 310 32. 33- 31.. 35. A6. ,Prune dwarf and the cherry virus complex. Science ~(N.S. ) 100:1A7-148.19l.1.. , G. W. Keitt, J. D. Moore and H. J. Miller. Necrotic ring spot. pp. 62-63. In E. M. Hildebrand, G. H. Berkley and D. Cation., Handbook of virus diseases of stone fruits in North America. Mich. Agr. Expt. Sta. Misc. Publ. 191:2. , and D. H. Palmiter. Yellow-red virosis (Ii-disease) of peach and ohokecherry. Pl. Dis. Reptr. 22: 394-396. 1938. Keitt, G. W. and C. N. Clayton. A destructive bud—transmissible disease of sour cherry in Wisconsin. Phytopath. 29: 821- 822. 1939. , and J. D. Moore. Masking of leaf symptoms of sour cherry - yellows by temperature effects. Phytopath. 33: 1213-1215. 191:3. Kunkel, L. 0. Heat treatments for the cure of yellows and other virus diseases of peach. Phytopath. 26: 809. 1936. Milbrath, J. A. and S. M. Zeller. Indexing fruit trees for virus. American Nurseryman, Sept. 1,.19h8. Mills, W. D. Temperature effects on the expression of the yellows virus in sour cherries. Phytopath. 36: 353-358. 191:6. Moore, J. D. and G. W. Keitt. Host range studies of necrotic m spot and yellowsof sour cherry. Phytopath. 34: 1009. 191.1: Abst) . and _, An indexing method for necrotic ring spot and yellows of sour cherry. Phytopath. 39:15-16.19h9. (Abst.) Palmiter, D. H. and K. G. Parker. Peach X-disease on sour cherry. Phytopath. 38: 20. 1948. (Abst.) Rasmussen, E. J. Green ring mettle. p. 61. In Hildebrand, E. M., G. H. Berkley and D. Cation, handbook of virus diseases of stone fruits in North America. Mich. Agr. Expt. Sta. Misc. Publ. 1942. , D. Cation. Experiments on the yellows disease of sour cherry'. (m us cerasus). Phytopath. 32: 15. l9h2. (Abst.) ...... Rawlins, T. E. and H. E. Thomas. The buckskin disease of cherry and other stone fruits. Phytopath. 31: 916-925. 1941. Reeves, E. L. and L.M. Hutchins. A progress report on Western X- disease, a virus. disease of peaches. Wash. State Hort. Proc. 37: 27-29. 191:1. Richards, E.L., LcM. Hutchins and E.L. Reeves. The Western’x" virus a cause of "littlecherry" in Utah. Phytopath. 39: 19. l9h9.(AbstJ Ila .... In ..- iii 36. #1. 1+2. #7. Richards, B. L., E. L. Reeves, and L. M. Hutchins. Wilt and decline, a virus of sweet and sour cherries in Utah. Phytopath 36: 1.09. 191.6. (Abst.). Stoddard, E. M. fiogress reports of investigations on new peach trouble. Proc. Conn. Pomol. Soc. 13: llS-ll7. 1931.. , The X-disease of peach and its chemotherapy. Conn. ‘Agre Ekpt. Sta. Bus 506. 1947. Thomas, H. E. and E. M. Hildebrand. A virus disease of prune. Phytopath. 26: nus-131.8. 1936. , and Rawlins, T. E. Some mosaic diseases of Prumls . - . species. Hilgardia 12: 623-644. 1939. ----- Valeau, W. D. A virus disease of plum and peach. Ky. Agr. Willison, R. 5. Strains of prune dwarf. Phytopath. 3t: 1037- 101090 19440 , A line pattern virosis of Shiro plum. PhytOpath. .... .35: 991-1001. 191:5. Zeller, S. M. and J. A. Milbrath. Banded chlorosis, a trans- missible disease of cherry. Phytopath. 32: 634—635. 191.2. .. so, #5 52‘ I“: 5.3 a i "i 3 \ n! i ‘r r. i; M'clfiifififlfifllfiflilMUflfliifliflWflflflilTVs