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[f]!!! u. ‘ 4 . ‘l\.\‘fi\‘ 1’11"] - ‘Ill’ I “\lyby' ‘i'.‘-((.Ia.¢ '0.» I w , ‘ .- (ll 0 J u If: j, DJII‘IHI'IIIIIII'DI‘I. l . \1 1 ll 0.. I I? ’u w‘ . fl.ul'll l-IJI’III‘. (ION ‘I'llnir‘ln I |1ll| I’l-lr'l."l IHHWIHHIIHHHWIHMI“WNWIJLHIHZIHHHI _3_ 1293 1051 This is to certify that the dissertation entitled fax; y{6/77/&w(,m14 figéé fW Sam/mi MW ///Ut/of'n:1 cl/fl/4' C/Vfl‘ét/uj ({wa DOK/(A) /)/qu fl'L/L presented by N7 6" 7 "-CL A2 W has been accepted towards fulfillment of the requirements for _f’AD degreein #fl/fféu/ae JWMIQWWQQ Major professor DateW W7) ’36” MS U is an Affirmative Action/Equal Opportunity Institution 0-12771 MSU LIBRARIES “ RETURNING MATERIALS: Place in back drop to remove this checkout from ‘your record. FINES will be charged if book is returned after the date stamped below. [:2 gm ROLE OF ETHYLENE AND THE EFFECT OF SU.PLEMENTAL HAND POLLINATION IN APPLE (MALUS DOMESTICA BORKH.) FRUIT SET By Majid Rahemi A DISSERTATION Submitted to Wichigan State University in partial fulfillment of the requirements for the degree of DOCTOR OF PHILOSOPHY Department of Horticulture H KO CI) p.) ABSTRACT ROLE OF ETHYLENE AND THE EFFECT OF SUPPLEMENTAL HAND POLLINATION IN APPLE (MALUS DOMESTICA BORKH.) FRUIT SET By Majid Rahemi To determine the role of ethylene in apple fruit set, aminoethoxyvinylglycine (AVG), an inhibitor of ethylene synthesis, silver thiosulfate (STS), an inhibitor of ethylene action, and (2-chloroethyl) phosphonic acid (ethe- phon), an ethylene generating compound, were applied in 1980 and 1981 to branch units of 'McIntosh', 'Delicious', and 'Golden Delicious' apple trees at full bloom and/or at 18 to 21 days after full bloom. Additional branches were scored with a knife. When applied at full bloom AVG (200 ppm) significantly increased set in all cultivars, but effects of 100 ppm were generally non-significant. Although both concen- trations reduced ethylene evolution from flowers excised 1 to 10 days after treatment, the effects were small and non-significant. Ethephon (A0 to 100 ppm) did not affect fruit set significantly, yet markedly increased ethylene Majid Rahemi production. Fruit set was much better correlated with ethylene evolution in 'Delicious'than in 'McIntosh' or 'Golden Delicious', but r values were non-significant. Therefore the effects of AVG on fruit set appear to be in- dependent of its effects on ethylene synthesis. 2 Application of AVG (200 ppm) prior to "June" drOp had no significant effect on ethylene evolution in any of the three cultivars. The chemical significantly reduced fruit retention in 'McIntosh' and increased it in 'Delicious' in 1980, but had no significant effects in 1981. Neither STS nor scoring had consistent effects on either set or ethylene evolution. Ethephon (200 ppm) significantly increased both fruit drop and ethylene evolution in all three cultivars, but 100 ppm was effective only in 'McIntosh' and 'Golden Delicious'. Ethylene evolution prior to and during "June" drop was measured in two populations of fruits selected on the basis of diameter. Small fruits with a higher abscission potential generally produced more ethylene per unit weight than large fruits sampled at the same time. However, when the rate of ethylene production of non-treated fruits was plotted against fruit weight, rather than against date of sampling, abscission potential appeared to be in- dependent of ethylene production. These data suggest that differences in the rate of ethylene evolution were due to the differences in size, rather than to differences in ab- scission potential, and that ethylene is not the primary factor responsible for "June" drop. Majid Rahemi Aqueous sprays of AVG were applied at full bloom to flowers on bagged limbs of 'McIntosh' and 'Delicious' to determine its effect on the effective pollination period. AVG-treated and control flowers were hand-pollinated with 'Empire' pollen at l to 3 day intervals beginning at an- thesis. AVG (200 ppm) increased fruit set in 'Delicious' but not in 'McIntosh'. Fruit set decreased as the time of pollination was delayed and reSponse of AVG-treated flowers paralleled that of control flowers. The data obtained in this study indicate that AVG has little or no effect on the effective pollination period. To test the hypothesis that "basal gaps" between the stamens of 'Delicious' flowers limit fruit set by permitting bees to obtain nectar without transferring pollen to the. stigmata, open-pollinated flowers of 'Delicious' and 'Mc- Intosh' were hand-pollinated at anthesis. Supplemental hand pollination increased initial and final fruit set in 'De- 1icious' but not in 'McIntosh', supporting the hypothesis. However, final set of open pollinated flowers was no greater in 'McIntosh' than in 'Delicious'. The results are there- fore inconclusive. Staining with aniline blue made pollen tubes visible in the entire length of 'McIntosh' styles, but only in the upper half of 'Delicious' styles. Many pollen tubes in open- and open-plus hand-pollinated flowers reached the base of 'McIntosh' styles within “-6 days, but very few reached the base in self-pollinated (bagged) flowers and most terminated in swollen tips and highly cal- loused plugs. Such flowers set almost no fruits. Dedicated to The Martyrs of the Islamic Revolution of Iran 11 ACKNOWLEDGMENTS I would like to express my sincere appreciation to Dr. Frank G. Dennis for his invaluable suggestions, guidance, all he taught me, and for his friendship throughout my research and preparation of this thesis. I am grateful to Dr. Robert L. Andersen for his advice and encouragement during my graduate study and this research, and to Dr. James A. Flore, Dr. David A. Reicosky, and Dr. Roger A. Hoopingar- ner for their helpful suggestions during my study. I thank Dr. Robert C. Herner, Dr. Kenneth C. Sink, and Dr. David R. Dilley for suggestions and for use of their laboratory facilities, and to all my fellow graduate students for their help and respectful relationships during my study. I especially thank my wife, Zahra, for her patience, understanding and support, and my parents, sisters, and brothers for their love and support during my graduate studies. iii Chapter TABLE OF CONTENTS LIST OF TABLES. LIST OF FIGURES INTRODUCTION. REVIEW OF LITERATURE. I. Delicious Apple Production in the United States . . . II. Factors Affecting Fruit Set of Delicious A. Environmental Factors B. Pollination 1. Flower Structure. 2 Pollinizer Cultivar 3. Pollinizing Agents. A Supplementary Pollination C. Ovule Longevity D. Ethylene. III. Physiology and Biochemistry of Ethylene. . . . . . . A. Ethylene as a Plant Hormone B. Ethylene Biosynthesis 1. Biochemical Pathway 2. Factors Affecting Rate of Ethylene Biosynthesis C. Ethylene Generators iv Page viii xi U‘lU'l OZ! 13 1A 16 18 19 21 21 22 28 Chapter Page D. Inhibitors of Ethylene Action. . . . . . . . . . . . . . . . . 29 IV. Evidence for the Role of Ethylene in Fruit Set. . . . . . . . . . . . . . . 31 A. Ethylene Production by Flowers and Fruits. . . . . . . . . . . . . . . 31 1. Effect of Pollination . . . . . . . 31 2. Effects of Auxins . . . . . . . . . 33 B. Effects of Ethylene-Generating Compounds . . . . . . . . . . . . . . . 3A 1. Effects on Initial Set. . . . . . . 3A 2. Effect on "June" Drop . . . . . . . 36 C. Effects of Inhibitors of Ethylene Biosynthesis. . . . . . . . . . 39 D. Effects of Inhibitors of Ethylene Action. . . . . . . . . . . . . . A1 SUMMARY . . . . . . . . . . . . . . . . . . . . . . A2 LITERATURE CITED. . . . . . . . . . . . . . . . . . AA SECTION I THE ROLE OF ETHYLENE IN FRUIT SET OF APPLE. . . . . 55 Abstract. . . . . . . . . . . .‘. . . . . . . . . . 56 Materials and Methods . . . . . . . . . . . . . . . 59 Experimental procedure, 1980. . . . . . . . . . 59 Experimental procedure, 1981. . . . . . . . . . 61 Results . . . . . . . . . . . . . . . . . . . . . . 62 Effects on fruit set and characteristics, 1980. . . . . . . . . . . . . . . . . . . . . . 62 Effects on ethylene evolution, 1980. . . . . . . . . . . . . . . . . . . . . . 66 Effects on fruit set and charac- teristics, 1981 . . . . . . . . . . . . . . . . 68 V Chapter Effects on ethylene evolution, 1981. . Discussion. Literature Review . . . . . . . . . . . SECTION II RELATIONSHIP BETWEEN ENDOGENOUS ETHYLENE EVOLUTION AND APPLE FRUIT ABSCISSION DURING "JUNE" DROP . . . . . . . . . . . . Abstract. Materials and Methods Results Discussion. Literature Cited. SECTION III EFFECT OF AMINOETHOXYVINYLGLYCINE ON THE EFFECTIVE POLLINATION PERIOD OF APPLE Abstract. Materials and Methods Experimental Procedure. Results Discussion. Literature Cited. SECTION IV THE EFFECT OF SUPPLEMENTARY HAND POLLINATION ON FRUIT SET AND POLLEN TUBE GROWTH IN APPLE. Abstract. vi Page 73 76 814 86 87 9“ 114 119 122 123 125 125 126 134 136 138 139 Chapter Page Materials and Methods . . . . . . . . . . . . . . . 1H2 Results . . . . . . . . . . . . . . . . . . . . . . 1A3 Effect of supplementary pollination on fruit set. . . . . . . . . . . . . . . 143 Germination of pollen and growth of pollen tubes in the style . . . . . . . . . . . 1A5 Discussion. . . . . . . . . . . . . . . . . . . . . 1&5 Literature Cited. . . . . . . . . . . . . . . . . . 152 SUMMARY AND CONCLUSIONS . . . . . . . . . . . . . . ISA LITERATURE CITED. . . . . . . . . . . . . . . . . . 158 vii LIST OF TABLES Table 1. Average annual production (million Kg) of 'Delicious' apple in l97h-l977 and pro- Jecged production (Ricks and Pierson, 197 ). 2. Effects of ethephon on apple fruit set when applied prior to or during bloom . 3. Effects of ethephon on apple fruitlet ab- scission when applied at various times after full bloom Section I 1. Mean of maximum and minimum temperatures (°C) during sampling for C2Hu measure- ment from apple flowers and young fruits at the Horticultural Research Center, East Lansing, MI 1980-81. 2. Effects of AVG and ethephon on fruit set, fruit retention, fruit per cm limb cir- cumference and fruit characteristics at harvest of apple, E. Lansing, MI 1980. 3. Effects of AVG and ethephon on ethylene evolution from apple flowers and young fruits l to 10 days after treatment, E. Lansing, MI, 1980. . . . . . . . . 4. Effects of AVG and ethephon on mean ethylene production (nz/g f.w./hr) by apple flowers during 3 days follow- ing treatment. E. Lansing, MI 1980. 5. Effects of AVG and ethephon on fruit set, fruit retention, fruits per cm limb circumference and fruit characteristics at harvest of apple, E. Lansing, MI 1981 . . . . . . . . . viii Page 35 37 63 6M 67 69 7O Table 6. Effects of AVG and ethephon on ethylene production of apple flowers and young fruits as a function of days after treatment, E. Lansing, MI 1981 Effects of AVG and ethephon on mean ethylene production (nA/g f.w./hr) by apple flowers during 3 days following treatment. E. Lansing, MI, 1981 Relationship between initial fruit set of apple and mean ethylene evolution from flowers during 3 days following chemical treatment . Effect of petal removal on ethylene productionbby apple flowers, 1981. Section II Mean of maximum and minimum temperatures (°C) during time of sample collection for ethylene determination in apple fruits at the Horticultural Research Center, East Lansing, MI Effects of AVG, ethephon and double scoring on fruit retention and fruit diameter of apple 1980. Sprays applied June 3 ('McIntosh'), 8 ('Delicious’ ) or 9 (' Golden Delicious' ) . . . Effects of aminoethoxyvinylglycine (AVG), ethephon and double scoring on ethylene evolution (nA/g/hr) from 'McIntosh' apple fruits prior to "June" drop 1980. Treat- ments applied June 3. . . . . . . . . . Effects of aminoethoxyvinylglycine (AVG), ethephon and double scoring on ethylene evolution (nA/g/hr) from 'Red Prince Delicious' and 'Golden Delicious' apple fruits prior to "June" drOp 1980. Treat- ments applied June 5 ('Delicious' ) or 9 (' Golden Delicious' ) . . . . ix Page 74 75 79 82 95 96 97 98 Table 5. Effects of AVG, ethephon and silver thio- sulfate (STS) on fruit abscission and fruit characteristics at harvest of 'McIntosh' and 'Red Prince Delicious' apple, 1981. Treatments applied May 28 ('McIntosh'), or 30 ('Delicious') Effects of aminoethoxyvinylglycine (AVG), ethephon and silver thiosulfate on ethylene evolution (11?. /g/hr) from 'Mc- Intosh' and 'Red Prince Delicious' apple fruits prior to "June" drop 1981. Treat- ments applied May 28 ('McIntosh'), or 30 ('Red Prince Delicious'). . . Average fruit weight, diameter, abscis- sion potential and ethylene production of untreated control apple fruits prior to and during "June" drop, 1981. Summary of effects of treatments on apple fruit retention and ethylene evolution 1980- 81 . Section III Effect of AVG on effective pollination period and fruit characteristics at harvest of 'McIntosh', Leslie, MI 1981. Flowers on bagged limbs treated with AVG (200 ppm) at full bloom (May 6). Effect of AVG on effective pollination period and fruit characteristics at harvest of 'Starking Delicious', Leslie, MI 1981. Flowers on bagged limbs treated gith AVG (200 ppm) at full bloom (May ) . . . . . . . . . . . . . . . . . . Section IV Effect of supplemental hand pollination on fruit set of apple at Leslie, MI 1981. Effects of self— (8), open- (O), and open-plus hand- (O+H) pollination on pollen germination and tube growth in apple flowers, 1981, as determined by staining with aniline. . . . . . . Page 101 102 109 115 127 131 IAN 150 LIST OF FIGURES Figure Section I 1. Relationship between mean ethylene evolution from flowers the first three days after treatment and initial fruit set of apple Section II 1. Effect of cultivar on relationship between fruit diameter on June 3 (’McIntosh'), 8 ('Delicious') or 9 ('Golden Delicious') and abscission during "June" drop 1980. Surviving fruits counted June 15 ('Mc- Intosh'), 20 (' Delicious' ) or 21 ('Golden Delicious‘ ). 2. Relationship between diameter and weight of 'McIntosh' fruits in 1980 and 1981. 3. Relationship between fruit diameter at various sampling times and fruit ab- scission as of June 12 (McIntosh) or IR (Delicious) 1981. N. Ethylene production by large vs small 'McIntosh' (A + B) and 'Red Prince Delicious' (C + D) apple fruits as a function of day of sampling (A + C) or fruit weight (B + D). Day 0 was May 27 ('McIntosh' ) or 30 ('Delicious' ), 1981 . . . . . . Section III 1. Effect of AVG on effective pollination period (E.P.P.) of 'McIntosh' and 'Starking Delicious' apple 1981. AVG (200 ppm) applied May 6 ('Mc- Intosh') or 8 ('Starking Delicious') to bagged limbs. Flowers hand-pol- linated with 'Empire' pollen at xi Page 78 106 108 111 113 Figure indicated times. Broken lines indi- cate initial set of AVG-treated (a) and control (b) flowers, solid lines final set of AVG- treated (c) and con- trol (d) flowers . . . . . 2. Fruit set in response to AVG expressed as percent of the control as a func- tion of time of pollination. Section IV 1. Appearance of pollen tubes in 'McIntosh' apple styles under ultraviolet light after staining with 0.1% aniline blue in 0.1 N K3P0u. a and b self-pollinated; c and d Open-pollinated; e and f open- and hand—pollinated. Flowers collected 2 (A,C,E) or 6 (B,D,F) days after hand- pollinating at full bloom. Abrupt ter- mination of fluorescence in (0) indicates point at which style was severed 2. Appearance of pollen tubes in 'Starking Delicious'_apple under ultraviolet light after staining with 0.1% aniline in 0.1 N K POu. a and b self-pollinated; c and d Open pollinated; e and f open- pollinated; e and f open- plus hand- pollinated. Flowers collected 3 (A, C ,E) or 6 (B,D, F) days after hand- pollinating at full bloom. . . . . xii Page 129 133 1“? 1H9 Guidance Committee: The Journal-article format was adopted for this dissertation in accordance with departmental and university requirements. Four sections were prepared and styled for publication in the Journal of the American Society for Horticultural Science. INTRODUCTION Although “,000 to 5,000 cultivars of apple had been" described by 1938 (Hedrick 1938) only two dozen cultivars accounted for 95% of the total commercial crop in the United States in 1969 (Henderson 33 a1. 1969). The leading cultivar in the United States, 'Delicious', represented 30% of the total 0.8. production in 1969 and 35% in 1975 (USDA, 1977). 'Golden Delicious" ranked second and ac- counted for 13% in 1975. Other leading varieties were 'McIntosh', 10%; 'Rome Beauty', 8%; 'Jonathan', 6%; and 'York Imperial', 5%. 'Delicious' is now the world's most popular variety. Its high color and characteristic shape make it a favorite of both grower and consumer. According to Maas (1970) no variety of apple of American origin was ever accepted more readily by the American public. Despite being the leading apple variety and having a higher market demand,'Delicious' is recognized as a light bearer. In 1928 Howlett stated .... "that the 'Delicious' apple under certain growth conditions tends to be a rela- tively light yielding variety is gradually becoming recog- nized. In Ohio 'Delicious' has shown a tendency to set light even in mixed plantings". Young Delicious trees often bear light crops for several years after coming into production. Greene and Lord (1978) stated that lack of flower bud initiation and/or fruit set often limits the productivity of young 'Delicious' apple trees. The light yields are due at least in part to light fruit setting rather than to irregular and light fruit bud formation (Howlett, 1928). 'Delicious' performs better in Washington State, presumably because of higher solar radiation (Way 1973). In the eastern states production of 'Delicious' remains av continuing challenge because of erratic and low yields per acre. Currently research is being directed toward methods for increasing yields of 'Red Delicious' in eastern regions. The purpose of this study was to determine the role of ethylene and the effect of supplemental hand pollination in controlling fruit set and development of apples under orchard conditions. REVIEW OF LITERATURE I. Delicious Apple Production in the United States The 'Delicious' apple is one of the most desirable of all the varieties grown commercially in the United States (Forshey, 1953). Of 17 leading varieties,production of only 'Delicious', 'McIntosh', 'Golden Delicious', and 'Cortland' has increased appreciably in the U.S. since 1942 (Childers, 1969), with 'Delicious' and 'Golden De- licious' increasing spectacularly. In Washington State 'Delicious' yields heavy crops. Tukey (1978) stated that good management, skill and hard work are important elements to bring trees into production. Since about 1950 'Delicious' and other varieties have been yielding better under Eastern U.S. conditions, apparently due to the use of milder fungi- cides and better cultural practices and strains (Childers, 1969;-Way, 1973). Over 150 red strains of 'Delicious' have been named but many have not been fully evaluated for yield and/or fruit quality. 'Starking' and 'Richared' are two strains which are widely planted (Childers 1969). Dennis (1979) summarized published yield differences of several strains of 'Delicious' in the United States, Italy and Poland and concluded that "strain has a marked influence on yield, although the spur strains appear to be less variable than non-spur strains". U.S. 'Delicious' production more than doubled in the last twenty years, increasing from an average of 500 mil- lion (26 million bushels) in the early 1950's to 1090.1 million Kg (58 million bushels) in 197A-l977 (Ricks and Pierson 1978). Planting of 'Delicious' increased in most regions during the last two decades, particularly in Wash- ington State. Washington production increased from an average of approximately AOA.06 million Kg (21 million bushels) during the early 1970's to 590.2 Kg in 1977, an increase of “7% within just five years. Washington now produces about 55% of the national 'Delicious' crop (Ricks and Pierson, 1978). Production of 'Delicious' in eastern states has risen onlygradually, in part because of con- tinuing erratic and low yield per acre. II. Factors Affecting Fruit Set of Delicious A. Environmental Factors Environmental conditions during bloom and early post bloom affect the performance of pollinating insects as well as the blossoms themselves. Bee activity is reduced by low temperature. Low temperature following pollination may reduce fruit set either by injury to pollen or pistil (Boyd and Latimer 1933). Differences in frost susceptibility exist among cultivars and 'Delicious' flowers are more susceptible to frost injury than flowers of many other cultivars (Hartman and Howlett, 1954; Roberts, 1946, Meader and Blasberg, 1946, Westwood gt a1,,l976). Wilson and Williams (1970) and Forshey (1978) suggested that sub- lethal injury may reduce the fruit-setting capacity of flowers which survive a freeze, but no data sum: available to support this suggestion. Temperature may have important effects on pollination, pollen tube growth and bee activity. Low temperature following pollination may reduce the set of fruit by slowing down pollen tube growth so that the sperm nuclei do not reach the embryo sac before disintegration com- mences (Boyd and Latimer 1933). Gardner gt a; (1949) col- lected data from 41 'Delicious' orchards across the United States and in Nova Scotia, and concluded that tempera- ture was one of the critical factors controlling fruit set in 'Delicious' with high temperature favoring and low tem- perature reducing set. On the other hand Roberts (1947) believed that warm nights decreased fruit set in 'Delicious'. Lu and Roberts (1952) hand pollinated flowers on apple trees held in greenhouses at 21 to 24°C or a minimum of 13. Better fruit set occurred in 'Delicious', 'McIntosh' and 'Wealthy' under cool than under warm conditions. They also reported that 'Delicious' blossoms dropped heavily at temperatures above 21°. Laping and Arndt (1974) reported that optimum temperature for pollen tube growth in 'De- licious' flowers was between 7.2°_12.8° and that pollen grains failed to germinate below 4.4°. They also report- ed that pollen tubes couldreach the base of the style in 3 days at an average day-night temperature of 12.8° and in 5 days at an average temperature of 7.8. Forshey (1977) re- ported that apple pollen tube growth practically ceased at 7.2°C. Cool temperatures reduce bee activity during bloom and may affect fruit set in apples. Lu and Roberts (1952) at- tributed poor setting of 'Delicious' during cool blossom periods to poor bee activity. Brittain (1933) studied bee activity in Nova Scotia apple orchards and found a rise in activity from 10 to 18°C followed by a gradual decrease to a low level at 30°. Although limited honey bee flight oc- curs at 15° to 18°, full flight requires temperatures ex- ceeding about 21° (Morse 1975). Reports on the effects of sunshine and cloudiness on fruit set during bloom are conflicting. Boyd and Latimer (1933) reported that lack of sunshine and cloudy weather wererun:detrimenta1 to fruit setting in the apple. They obtained a heavy set of fruit when hand pollination was performed during humid, cloudy or rainy weather with mild temperature. They attributed the better results obtained from pollination performed on cloudy days to high humidity. HoWever, Gardner,et a1.(l949) associated high fruit set of 'Delicious' with high radiant energy during the week after full bloom. Subsequently Dennis (1979) analyzed their data and found correlation coefficients were low and non- significant. Rain has no effect on fruit set unless, of course, it continues throughout bloom and prevents bee flight (Griggs 1958). Boyd and Latimer (1933) found that heavy rain did not wash pollen from the stigmas in sufficient quantity to be detrimental to obtaining a satisfactory set of fruit. They reported that set was as good following a heavy rain as following cloudy weather and high humidity without rain- fall. B. Pollination Delicious is totally self-unfruitful (Howlett, 1928; Overholser and Overly, 1931; Roberts, 1945) and cross pollination is essential for consistent heavy production. Howlett (1928) concluded that a considerable part of light fruit setting of 'Delicious' is due to inadequate pollina— tion. However, even with adequate cross pollination 'Delicious' was less productive than such heavy setting varieties as 'Jonathan', 'Grimes', 'Baldwin', 'Wealthy' and 'Yellow Transparent' (Howlett 1928). Several factors may contribute to unsatisfactory pollination of 'Delicious'. 1. Flower Structure Most apple flower clusters, which contain 5-6 flowers, are produced on spurs on 2- to 3-year-old wood, although some are borne laterally on l-year-old wood. The apple flower produces both nectar and pollen in greater abundance than most other deciduous fruit trees (Smith and Bradt, 1967). Roberts (1945) concluded that one reason for the poor set of 'Delicious' was the relative length of the pistils and stamens; the pistils were so short that bees which collected only pollen did not always touch the stigma. Forshey (1953) measured the length of pistils and stamens in several cultivars, and reported that they were of equal length in 'Delicious', 'Jonathan' and 'Rome Beauty'. He concluded that the structure of blossom is not responsible for poor set of 'Delicious'. However, Roberts (1945) also reported that the structure of 'Delicious' blossoms per- mits honey bees to remove nectar without pollination of the stigma; only about 20 percent of the bees visiting the flowers crawled over the stigma. Robinson (1980) reported similar "sideworking" of honey bees on 'Delicious' blossoms as a result of "basal gaps" or spaces between stamens. The maximum width of the tongue (glossa) of a honey bee is about 180u. Robinson measured the percent of basal gaps larger than 180 u between stamens in several cul- tivars of apple. He found the width of these gaps to be greater in 7 'Delicious' sports than in 11 other cultivars. 10 Therefore, nectar collection through basal gaps was more difficult in cultivars such as 'Golden Delicious', 'Mc- Intosh', 'Idared', 'Cortland', 'Jonathan' and 'Rhode Is- land Greening' than in several 'Delicious' strains or in 'Northern Spy', which also exhibits large basal gaps. He believed that the behavior of honey bees on apple blossoms is determined by the presence or absence of these basal gaps, which reduce the rate of cross-pollination. 2. Pollinizer Cultivar Investigators have used techniques such as hand pollina- tion and bagging or emasculating blossoms to determine the effectiveness of various commercially important cultivars as pollinizers for 'Delicious'. Overholser and Overly (1931) reported that 'Blackjon', 'King John', 'Red Rome', 'Jonathan' and 'Rome Beauty' were satisfactory pollinizers. All red sports of 'Delicious' tested, with the exception of 'Van Buren', have proven to be incompatible with the parental variety and thus should not be used as pollinizers. (Wellington, 1947). All of the triploids such as 'Baldwin', 'Rhode Island Greening', 'Stayman' and 'Winesap' have 51 chromosomes, an uneven number. They therefore produce pollen of low germination and consequently are not reliable pollinators (Overholser and Overly, 1931; Wellington,l947). The diploid cultivars have 34 chromosomes in each somatic 11 cell and produce good or at least a fair amount of viable pollen (Wellington,1947). Roberts (1947) made a survey to compare fruit set in 166 blocks of 'Delicious' from Arkansas and Minnesota to Virginia and Nova Scotia and around the Great Lakes. He found a consistent record of good set of 'Delicious' when pollinizers set well. He noted that the best pollinizers, based upon yield of 'Delicious', were 'Rome Beauty' and 'Northern Spy'. 'McIntosh' and'Winesap' (triploid) were intermediate. 'Winter Banana' and 'Baldwin' (triploid) were relatively poor and 'Rhode Island Greening' (triploid), 'Duchess' and 'Stayman' (triploid sport of 'Winesap') very poor. The value of 'Golden Delicious' as a pollinizer for the 'Delicious' is uncertain. Whitehouse and Auchter (1927) noted that 5.8 to 9.8% of flowers of 'Delicious' hand pollinated with 'Golden Delicious' pollen set fruit whereas open pollinated blossoms set 16.7%. Knowlton (1929), however, reported that 'Golden Delicious' was a good pollinizer of 'Delicious' in West Virginia. Overholser and Overly (1931) found that the pollen of 'Golden Delicious' gave an average set of 11.6% vs. 14.1% for 'Jonathan' pollen when used on 'Delicious' over a 3-year period (1928-31). Tukey (1978) considered 'Winter Banana' to be superior to 'Golden Delicious' as a pollinizer for 'Delicious' in Washington State because it blooms early, overlapping the king flower of 'Delicious'. Hull (1978) stated that 'Jonathan', 'Empire' and 'Idared' have performed 12 well in Michigan as pollinizer cultivars for 'Delicious'. In planting solid blocks of commercial apple cultivars, use of top grafts of pollinizers or interplanting small trees of flowering crabapple may correct the cross pollina- tion problem (Hoffman, 1966; Williams,1972). Artificial pollination techniques may be required to increase the crop in rows far from pollinizers even if natural pollination is optimal (Williams,1970). Lapins and Arndt (1974) reported that inadequate numbersenuipoor distribution of pollinizers was one of the main causes of poor fruit set. A ratio of four rows of 'Delicious' to one of pollinizer should be satisfactory in most situations but in colder locations with generally poor pollination conditions two rows of 'Delicious' and two rows of another variety could be a better arrangement. Roberts (1947) found that 'Delicious' trees at the edge of an orchard regularly had somewhat better set, as incoming bees brought pollen from a distance. He concluded that greater bee visitation is needed to set 'Delicious' than is required for most other varieties. He also suggested that 'Delicious' should be planted no more than one row away from a good compatible pollen source. Pollen tube growth differs following cross- vs.self- pollination. Modlibowska (1945) observed 3 types of pollen tubes in apple and pear styles. 1) Incompatible tubes - these grow slowly and are inhibited earlier at 25 to 30°C than at 10-20°C;~ 2) Semi-compatible tubes - these tubes l3 grow slowly but unlike the completely incompatible tubes they do not stop growing completelys 3) Compatible tubes — the growth rate of tubes at 10-15°C is similar to that of those in classes 1 and 2, but is accelerated by a rise in temperature. Temperature affects the growth of incompat- ible tubes in the opposite way. 3. Pollenizing Agents Pollination agents are important during bloom of apple. The chance of pollen transfer by wind is very low and fruit set is highly dependent upon insect activity and sources of viable pollen (Free,l964). Apple pollen grains are sticky and dense and adhere tenaciously to one another, in contrast to the wind borne pollen of some plants. Robinson (1980) confirmed that wind played an insignificant role in pollina— tion of apples by caging apple trees to exclude insects but permit wind transfer of pollen. Flowers did not set fruit even if bouquets of a pollinizing cultivar were suspended among the branches. Another obvious problem with wind pollination is that it is undirected. The "target area" (stigmatic surface) is very small on apple blossoms, and pollen must be transferred directly to the stigmas to be effective. These facts leave insects as the main vector, par- ticularly honey bees (Hoffman,l966, Wilson and Williams 1970). The number of honey bees per colony (about 30,000 14 at apple bloom) is huge in comparison with the number of other bees and they are easily moved in large numbers by commercial bee keepers. The colonies should be placed at points where they receive maximum sunlight and the sites should be well drained and protected from winds. One colony per acre is recommended (Robinson,l980). 4. Supplementary Pollination Supplemental hand pollination is a potential method for enhancing fruit set, but the cost would probably be prohibitive. Williams (1970) investigated the effects of natural and supplementary hand pollination on fruit set of 'Cox's Orange Pippin' apple in England. He found that the average final set per 100 flower clusters for 21 'Cox's' orchards was 33.1 for branches receiving supplementary pol- lination (one flower hand pollinated per 4 clusters) vs. 25.6 for controls with natural pollination only. Supple- mentary pollination increased yield by an average of 29%. He concluded that orchards in which hand pollination is effective are deficient in bees, assuming suitable pol- linizers and spacing. Kondrate'ev,et a1.(l972) hand- pollinated apples, cv. Golden Winter Pearmain, and plum, cv. Tulew Gras, repeatedly at 6 to 72 hour intervals, and concluded that supplementary pollination speeded up fer- tilization and improved fruit set. However, they did not present data to support their conclusion. Increased fruit 15 set may have been the result of higher numbers of pollen grains per stigma. Forshey (1978) reported that the amount and timing of cross-pollination are critical. Theoretically only a few pollen grains per flower are required for ferti- lization. However, 'Delicious' flowers must be saturated with pollen because a relatively low percentage of pollen tubes actually reach the embryo sac. When post bloom weather is cool, slow pollen tube growth in combination with early degeneration of embryo sacs results in poor set. He demonstrated that receptivity of 'Delicious' flowers is not uniform throughout the bloom period and the effective pol- lination period may be limited to one or two days. How- ever, he failed to factor out flower age. Lapins and Arndt (1974) found a close relationship be- tween the amount of pollen on the stigmas and final set. A low amount of pollen was found in cool areas and in dense orchards. They rated crop production from 1 to 10. In or- chards with a high proportion of pollinizer, the rating was 6.0 whereas in orchards with insufficient pollinizers the rating was 1.5. They reported that less than about 50 pol- len grains per stigma usually resulted in poor germination of pollen, slow growth of pollen tubes and low set of fruit. However, they did not present data to support this observa- tion. Another factor which may control pollen tube growth is double pollination. In an early report Cooper (1928) 16 indicated that when 'Delicious' pistils were pollinated with pollen from various cultivars, 'Ben Davis' pollen tubes were the longest, followed by 'Transparent', 'Jonathan', 'Stayman' (triploid) and 'Delicious'. Knight (1917) re- ported that pollen tubes make their way through the tissue along a more or less well-defined path which is accompanied by the decomposition of cells or extrusion of material from them. Visser and Verhaegh (1980) investigated the effect of two consecutive hand pollinations of 'Golden Delicious' flowers, as well as the separate effect of each, with the aid of mildew- or scab-resistant pollen donors. The flowers were either pollinated once or twice at intervals of one or two days, the seeds were harvested from the fruits obtained, and the seedlings evaluated for disease resistance. They found that double pollination with the same pollen had a similar effect on fruit and/or seed development as a double pollination with pollen from two different cultivars. In all trials an average of 37% of the seeds originated from the first and 63% from the second pollination. They con- cluded that pollen tubes grew more rapidly in styles which had been previously penetrated by pollen tubes. C. Ovule Longevity Ovule longevity may also be an important factor in 'Delicious' fruit set. Hough (1947) reported that in ovules of 'Delicious' the most frequent abnormality was 17 either a tardy initiation of the megaspore mother cell or a slower rate of development of the megaspores and embryo sac. Such retarded embryo sacs would seldom be expected to develOp into fully differentiated eight nucleate embryo sacs in time for fertilization, especially if their develop- ment continued to be at a slower than normal rate. Other apparently normal embryo sacs broke down soon after the flower opened, even though the flowers had been pollinated with compatible pollen. Hartman and Howlett (1954) found that a considerable percentage of 'Delicious' embryo sacs were delayed in development or showed signs of premature' degeneration and the percentage increased markedly after 72 hours. When pollination was delayed until 48 hours after anthesis fertilization was greatly reduced. They attributed this largely to the early ovule degeneration. Forshey (1978) stated that a significant prOportion of 'Delicious' flowers are not viable; however, he did not present support- ing data. In some nonviable flowers the embryo sac is im- mature at bloom, while in others early degeneration of the embryo sac excludes the possibility of fertilization. Rootstock may have an effect on embryo sac degeneration in 'Delicious'. Marro (1976) made a comparison of fruit set cfi'PRichared Delicious' apple trees on seedling vs. M9 rootstock. Flowers were hand pollinated at petal opening, full bloom and petal fall. Embryo sac degeneration was observed in a certain percentage of flowers pollinated at 18 full bloom but was greater at petal fall. On both dates embryo sac degeneration was greater on the seedling root- stock. Williams (1970) studied the relationship between flower rfertility and fruit set in 'Cox's Orange Pippin'. He termed the period during which pollination results in fertilization the "effective pollination period" (E.P.P.). E.P.P. is a function of ovule longevity and rate of pollen tube growth. Some flower clusters were pollinated on the day the flower opened (day 0), others 2, 4, or 6 days later. The results of this study indicated that the effectiveness of pollination decreased over the period investigated. The E.P.P. of various apple cultivars ranged from 2 to 3 days to 8 to 10 days after anthesis (Williams,1965a). Tem- perature during anthesis influences the length of the E.P.P. because of its effect on both tube growth and ovule longevity. D. Ethylene Greene (1980) reported that application of aminoethoxy- vinylglycine (AVG), an inhibitor of ethylene synthesis, increased set and reduced ethylene evolution in apple flowers. He suggested that because exogenous ethylene induces abscission, endogenous levels of ethylene in apple flowers may reduce fruit set. Williams (1980) applied AVG to spur 'Delicious' trees at 450 ppm prior to harvest in 1979. In the spring of 1980, ethylene evolution from buds 19 treated with AVG was lower than for control buds. This supports Greene's suggestion. However, neither investigator applied ethylene generating compounds such as ethe- phon at full bloom to determine their effects on fruit set. The role of ethylene in fruit set is discussed more fully in the following sections on the biosynthesis and physiological effects of this hormone. III. Physiology and Biochemistrygof Ethylene A. Ethylene as a Plant Hormone Evidence for the role of ethylene as a naturally-oc- curring plant hormone has accumulated over several decades (Leopold and Kriedemann, 1975). Neljubow (1901) was first to report that ethylene regulated the growth and development of plants. He observed that pea seedlings germinated in the lab grew in a horizontal direction. However, plants grown in air drawn from outside the lab grew normally in a verti- cal fashion. By adding illuminating gas to outside air he obtained the same growth phenomenon observed with laboratory air. He suspected that the hydrocarbon content of illumin- ating gas was the active factor. Laboratory air passed through a CuO ignition tube lost its ability to alter the growth of pea seedlings. Neljubow examined a number of the compounds of coal gas for their effects on plants. 802, C82, benzol, xylol, and naphthalene caused injury and in- hibited growth, but only acetylene and ethylene induced 2O horizontal growth. The initial suggestion that plants produced ethylene came from the report of Cousins in 1910. He observed that oranges produced a gas that promoted the ripening of bananas. Denny (1924) found ethylene to be the active component in combustion fumes which induced fruit ripening. Gane (1934) proved chemically that apple fruits produced ethylene. Denny and Miller (1935) exploited this idea and presented evidence for ethylene production not only by ripening fruits but also by flowers, seeds, leaves and even roots. Ethylene is a natural plant hormone because it is a product of plant metabolism, acts in trace amounts, and is neither a substrate nor cofactor in reactions associated with major plant developmental processes (Lieberman, 1979). Using very sensitive instruments and very careful tech- niques, it is possible to show that ethylene is an endo- genous growth regulator in plants, and that it is present in fruits from the earliest stages of development (Pratt and Goeschl, 1969). The advent of the gas chromatograph in the early 1960's allowed a rapid, sensitive and simple assay of ethylene evolved by plant tissue without extraction or purification prior to analysis. At the present time ethylene is recognized as a powerful natural regulating substance in plant metabolism, acting and interacting with other recognized plant hormones in trace amounts, and its 21 effects are observed especially during critical periods in the life cycle of higher plants (Lieberman, 1979). Yang (1980) prOposed that due to its gaseous nature, ethylene exerts a physiological effect at or near a site where it is synthesized. For this reason the classical definition of a hormone (action at a distance from the source) does not apply to ethylene. However, l-aminocyclopropane-l-carboxylic acid (ACC) synthesized in one part of the plant may exert its effect through conversion to ethylene in another part of the plant (Yang, 1980). B. Ethylene Biosynthesis 1. Biochemical Pathway Methionine was first suggested as a possible precursor of ethylene in climacteric fruits such as apple (Lieberman, 33 31-1965, 1966), banana (Burg and Clagett, 1967) and avocado (Baur,et a1. 1971) and this was proven by Yang (1974). Carbon 1 of methionine is converted to CO2 when radioactive methionine is fed to plant tissue (Burg and Clagett,1967), C to formic acid (Siebert and Clagett, 1969), Carbon 3 and 2 4 to ethylene (Baur,gt‘a1. 1971; Hanson and Kende 1976) and CH3-S remains in the tissue (Adams and Young, 1977; Burg and Clagett, 1967). Adams and Yang (1977, 1979) identified S-adenosylmethionine (SAM) and 1-amino-cyclopropane—l- carboxylic acid (ACC) as intermediates in the pathway from 22 methionine to ethylene and proposed the following sequence for the pathway of ethylene biosynthesis in apple tissue: methionine + SAM + ACC + ethylene. Identification of SAM as an intermediate indicates that methionine must be ac- tivated by ATP and methionine adenosyltransferase (Adams and Yang, 1977; Konze and Kende, 1979; Lieberman, 1979). The conversion of 5-adenosy1methionine (SAM) to l-amino- cyclopropane-l-carboxylic acid (ACC) by ACC synthase (Adams and Yang, 1979; Boller,et 31., 1979; Yu,et a1., 1979), requires pyridoxal phosphate. The methylthio (CH3—S) group is split off at this point and is incorporated into 5'- methylthioadenosine (MTA), which is converted to 5'-methyl- thioriboside (MTR). MTR is converted back to methionine by combining with a 4-carbon acceptor such as homoserine. This is an important step because many tissues do not contain high enough levels of methionine to maintain a high rate of 02H“ production unless the S atom is recycled (Herner, 1981). The third step is conversion of ACC to C2Hu, for which oxygen is required (Adams and Yang, 1979; Lonze and Kende, 1979). 2. Factors Affecting Rate of Ethylene Biosynthesis a. Carbon dioxide - Several environmental factors af- fect ethylene production. Depending on the tissue, CO2 can inhibit, promote, or have no effect on ethylene produc- tion. CO concentrations between 10% (Potter and Griffiths, 2 23 1947) and 80% (Burg and Thimann, 1959) inhibited ethylene production in mature apple fruit tissue. However, the effect may have been indirect via the inhibition of ripen— ing by CO The effect of CO2 on overcoming or blocking the 2. action of ethylene was noted as early as 1927. Mack (1927) observed that the addition of CO2 to the gas phase of ethylene reduced the ability of ethylene to blanch celery. After discovery of the blocking effect of CO2 on ethylene action, CO2 was found to be a competitive inhibitor of ethylene action. Burg and Burg (1965, 1967) observed that . CO2 is a close structural analogue of allene, a compound which substitutes for ethylene in both the pea section growth assay and in fruit ripening. 2=CH2 carbon dioxide allene ethylene O=C=O CH =C=CH CH Because CO2 has the structural features needed for ethylene action, except that it lacks the terminal carbon atom and is negatively charged on both ends, it could act as a com— petitive inhibitor of ethylene action. They tested this possibility by measuring the growth of pea stem sections in the presence of differing concentrations of C02 and ethylene. Concentrations of less than 1.8% CO2 competitively inhibited the action of ethylene. The affinity of ethylene for the receptor site is one million-fold greater than that of CO2; 24 therefore, if enough ethylene is present C0 will not prevent 2 its action. Imaseki, et a1. (1968) observed that removal of CO2 by KOH reduced ethylene production by sweet potato roots and suggested that CO2 stimulated ethylene synthesis. How- ever, CO2 has no effect on ethylene production by citrus fruits (Ben-Yehoshua and Eaks, 1969; Rasmussen and Jones, 1969). b. Oxygen. The oxygen level also affects ethylene production. The inhibition of ethylene production by low oxygen levels or anaerobiosis has been reported by many workers for a variety of tissues (Baur, et al., 1971; Burg and Thimann,l959; Curtis,1969, Haber, 1926). Burg and Thimann (1959) showed that the effect of oxygen on ethylene production in apple sections is similar to its ef- fect on respiration, and suggested that ethylene production is dependent on respiration. Apple sections stopped produc- ing ethylene soon after being placed in nitrogen. When the sections were returned to air, ethylene production re- sumed immediately at a greater rate than that of the con- trols. Burg and Burg (1967) showed that attachment of ethylene to the receptor is enhanced by 02 and this mechanism can be inhibited by CO. An ethylene precursor may accumu- late in the tissue under anaerobic conditions; this pre- cursor may rapidly be converted into ethylene in the 25 presence of oxygen (Abeles, 1973). Adams and Yang (1979) showed that oxygen is required for conversion of ACC to ethylene. c. Temperature. The Optimum temperature for ethylene production by apple is 30°C (Burg and Thimann, 1959; Hansen, 1945; Burg, 1962; Yu, gt g1., 1980). As temperature in- creases above 30°C the rate of ethylene production falls, ceasing entirely at 40° (Burg, 1962). Yu, gt g;., (1980) found that increasing the temperature to 35° caused 1-amino- cyclopropane-l-carboxylic acid (ACC) to accumulate, while reducing the rate of ethylene production. They suggested that the conversion of ACC to ethylene can be inhibited by high temperature. d. Chemical inhibitors. Uncouplers of oxidative phos— phorylation, such as 2,4-dinitrophenol (DNP), carbonyl cyanide m-chlorophenylhydrazone (CCCP) and C02+, drastically reduced both ATP and 02H5 production, presumably by inhibit- ing the step from methionine to SAM (Burg, 1973; Murr and Yang, 1975; Apelbaum, gt gt., 1981). However, if this step is the only one inhibited, adding l-aminocyclopropane-l- carboxylic acid (ACC) should overcome the inhibition by DNP and CCCP (Herner 1981). Several analogues of enol ether amino acids inhibit ethylene production. Owen and Wright (1965) extracted 26 L-2- amino-4-(2-amino-3—hydroxypr0poxy)—trans-3-butanoic acid (rhizobiotoine) with the structure H H CHZ-CH2-CH2-O-C=C-C-COOH ‘OH ‘NH2 NH2 from pure cultures of certain strains of the soybean root nodule bacterium,Rhizobium japonicum. Owen, gt gt. (1971) reported that this compound inhibited ethylene production about 75% in both intact sorghum seedlings and senescent apple fruit tissue slices. Although addition of methionine failed to overcome the inhibition completely in either system, it reduced the level of inhibition from 75% to 60%. They concluded that rhizobitoxine may block (a) the bio- synthesis of methionine, (b) the conversion of methionine to ethylene or (c) both. Scannel, gt gt. (1972) isolated methoxyvinylglycine from the fermentation broth of Pseudo- monas aeruginosa Ach—7700. In 1974 Pruess, gt gt. iso- lated a third inhibitor L-2-amino—4-(2-aminoethoxy)-trans- 3-butanoic acid or aminoethoxyvinylglycine (AVG), from a fermentation broth of an unidentified species of Strept_r myggg_sp. X—ll-085. AVG differs from rhizobitOXine only by the loss of the terminal methoxyl group. The structural similarities between these amino acid analogues and methio- nine suggest that they may be competitive inhibitors for 27 the substrate attachment site of the ethylene forming enzyme system. AVG and its analogues inhibit many pyridoxal enzymes, and conversion of SAM to ACC tg_tttg is greatly inhibited by AVG (Yang, 1980). Therefore Adams and Yang (1979) proposed that the enzyme catalyzing the conversion of SAM to ACC is a pyridoxal enzyme. Boller, et a1. (1979) observed that AVG at low concentrations in- hibited the action of ACC-synthase isolated from tomato fruit tissue (Ki = 0.2 uM), but did not inhibit ethylene production from ACC. e. Auxin as a stimulator of ethylene synthesis. A number of investigators have reported that auxin stimu- 1ates ethylene production which in turn induces premature abscission of leaves and other organs. Zimmerman and Wil- coxon (1935) observed that treatment of tomato plants with auxin stimulated evolution of a gas which caused epinasty. Later Morgan and Hall (1962) presented evidence that this gas was ethylene. Abeles and Rubinstein (1964) found that auxin application stimulated ethylene production from roots, stems and leaves of several genera and that the endogenous level of auxin also appeared to regulate the production of ethylene from vegetative tissue. IAA treatment of mung bean hypocotyls increased ethylene production 500-fold and stimulated conversion of methionine to ethylene. Yang (1980) suggested that auxin stimulates ethylene production 28 by inducing the synthesis of ACC synthase. Yu and Yang (1979) found that IAA stimulated ACC synthase activity and showed that conversion of SAM to ACC is the rate controlling step in ethylene production. C. Ethylene Generators Ethylene gas is difficult to apply under field condi- tions. This limitation was overcome by the development of ethylene releasing compounds which can be applied as sprays, the most important commercial compound being 2-chloroethyl- phosphonic acid (ethephon or CEPA) (deWilde,l971). Kaba- chnik and Rassuskaya (1946) described the synthesis of ethephon and Mayard and Swan (1963) reported the genera- tion of ethylene from this compound. Warner and LeOpold (1967) were the first investigators to report its use as a plant regulator. Ethephon is stable in the acid form but breaks down at a pH of 3.5 or above (Abeles, 1973). The pH of the cytOplasm of plant cells is generally greater than 4, so the growth regulating activity of ethephon has been at- tributed primarily to its ability to release ethylene within the tissue (Morgan, 1969; Warner and Leopold, 1969). Plant tissues of different acidity might be expected to show dif- ferent capacities for ethylene evolution. Warner and Leopold (1969) showed that leaves from Brygphyllum plants grown under long photoperiods produced substantially more 29 ethylene after treatment with CEPA than leaves from plants grown under short photoperiods. This could be expected from the relatively higher pH of sap from the former (pH 4.6 and 4.0, respectively). Temperature has a significant effect on the rate of ethylene evolution from ethephon. Olien and Bukovac (1978) incubated the apical segments of one-year-old sour cherry shoots in test tubes at 20, 30 or 40°C. Endogenous ethylene production increased with tem- perature up to about 30°, but the effect was small com— pared with the effect of temperature on release of ethylene from ethephon. The optimum temperature range for ethylene evolution from ethephon is 16° to 29° (Amchem Products, Inc., 1969). D. Inhibitors of Ethylene Action The biological action of ethylene can be overcome by silver ion. Spraying pea seedlings with silver nitrate effectively blocks the ability of exogeneously applied ethylene to induce the classical "triple" response -- growth retardation, stem swelling and horizontal growth. AgNO3 blocks ethylene stimulated leaf abscission in cotton (Beyer, 1976). Spraying or momentarily dipping carnation flower heads in AgNO solution (50-100 ppm) extended the life of 3 cut flowers and counteracted the enhancing effect of ethe- phon on senescence (Halevy and Kofranek, 1977). Saltveit, .gt gt. (1978) observed that infiltration of apple cortical 3O cylinders or banana fruit slices with solutions of AgNO3 (0.03 mM or greater) significantly reduced ethylene produc— tion. On the other hand, cuttings of sweet potato treated with Ag+ (250 to 2500 ppm) produced more ethylene than did controls (Walker,gt gt., 1979). Beyer (1979) showed that Ag+ (100 ppm) was clearly the most potent antiethylene treat- ment of several tested on pea. However, 'Delicious' apple flowers treated with silver nitrate (20 and 200 ppm) pro- duced more ethylene than did control flowers (Greene, 1980). The mechanism of inhibition of ethylene action by Ag+ has been studied by several investigators. The data of Burg and Burg (1967) suggest that the activity of ethylene requires binding to a metal. Ethylene, like other un- saturated aliphatic compounds, forms complexes with metals, including copper (Coates, gt gt., 1968). Month-old zinc- deficient tomato plants respond very little to ethylene even when exposed overnight, whereas plants deficient in copper, iron, phosphorus or nitrogen show strong epinasty within a few hours (Burg and Burg, 1967). Beyer (1976) suggested that Ag+ may substitute for Cu+, thereby interfering with ethylene oxidation, and hence ethylene action. Because Ag+ and Cu+ have the same valence, are similar in size, and both form complexes with ethylene, Ag+ might interfere with the binding of ethylene to Cu+. Walker, gt gt. (1979) suggested that Ag+ blocks the 02H“ activation site and thereby interferes with autocatalytic regulation of 02H“ production. 31 IV. Evidence for the Role of Ethylene in Fruit Set A. Ethylene Production by Flowers and Fruits 1. Effect of Pollination Pollinated flowers of cotton (Lipe and Morgan, 1973) and carnation (Nichols 1971, 1977) produced more ethylene than unpollinated flowers. Over half of the ethylene produced by one-day-old cotton flowers is released by the combined stigma, style, and stamens (Lipe and Morgan, 1973). Burg and Dijkman (1967) observed an increase in ethylene evolution within 8 to 10 hours after pollination and flower fading in orchid. The response was duplicated by applying IAA (5 mM). They concluded that release of pollen auxin in the stigma, and its diffusion to the column and lip, induced ethylene formation in these tissues, leading to floral fading. Hall and Forsyth (1967) measured ethylene production by flowers of strawberry and lowbush blueberry following pollination. Twenty-four hours after pollination pollinated flowers produced greater amounts of ethylene than non-pollinated flowers of the same age in each of the two species. The amount of ethylene produced following self-pollination was not significantly different from that following cross-pollination. Over 90% of the ethylene pro- duced by blueberry flowers came from the style and the stigma. They concluded that pollination may stimulate IAA formation leading to an increase in ethylene production. In banana 32 the rate of ethylene production is greater in flowers having an abscising perianth than in those with a persistent peri- anth (Israeli and Blumenfeld, 1980). Plich (1977) measured ethylene evolution from strawberry beginning the first day of flower opening and continuing until a few days after petal fall. He found abscission of petals in strawberry to be dependent on pollination, because pollination did not induce C2Hu evolution. Nicholas (1977) observed that most of the ethylene was evolved from the style and petals in carnation; pollination of intact flowers promoted endogenous ethylene production and accelerated petal wilting. He suggested that pollination accelerates petal wilting, stimulates ethylene production in all flower tissues and induces ovary growth. Approximately 40-50% of the ethylene could be accounted for by the styles and most of the re— mainder by the petals. Since the styles contribute less than 4% of fresh weight of the flowers, they are the most active center of ethylene production. Blanpied (1972) found that ethylene content of 'Golden Delicious' apple flowers increased as flowers developed. The level of ethylene increased from pink stage to petal fall, then de- creased as fruit development began; however, ethylene content remained high in unpollinated flowers which abscised. In 'McIntosh' flower buds ethylene content increased from green tip to tight cluster stage. In sweet and tart cherry ethylene content was high at half green calyx, declined 33 rapidly to green calyx, then declined slowly until petal fall. Abscised flowers of both apple and cherry contained more ethylene than adhering flowers. Abscising and adhering fruits of 'Golden Delicious', 'Red Astrahan', 'Delicious', and 'McIntosh' were collected and ethylene extracted from them during the period of "June" drop in two seasons. The fruit pedicel contained 3- to lO-fold more ethylene per unit weight than fruit tissue, but tissues of abscising fruits did not consistently contain more ethylene than similar tissues of adhering fruits. 2. Effects of Auxins Synthetic auxins such as naphthalene acetic acid and its derivatives have been used as thinning agents in apple for many years. Davidson, gt gt. (1945) first showed that NAA could be used as a post-bloom spray. They obtained effective thinning when apple trees were sprayed two to three weeks after bloom. Several theories have been suggested to explain the action of NAA in apple fruit thinning. Luckwill (1953) proposed that NAA thins by inducing seed abortion. Later Luckwill and Lloyd-Jones (1962) showed that only 0.2% of the 14 C-NAA applied to apple leaves was recovered from the seed after 5 days and none of it was in the form of unmetabolized NAA. They concluded that seed abortion and consequent abscission of the fruitlet is not due to the 34 direct action of NAA itself but rather to the effect of a breakdown product which has no auxin-like properties. The mechanism of action of NAA is still not well under- stood. One hypothesis is that application of NAA stimu- lates C2Hu synthesis, and the ethylene produced induces abscission of immature fruits. Schneider (1975) found that spraying with NAA 4 days after petal fall caused ethylene evolution in leaves, fruits and pedicels of 'Golden Delicious', 'Staymared' and 'Red Rome' apple sampled 24 hours and 48 hours after spraying. Walsh gt gt. (1979) sprayed 'Golden Delicious' and 'Northern Spy' with 15 ppm NAA two weeks after petal fall. Twenty hours after application ethylene evolution from 'Golden Delicious' spurs treated with 15 ppm was 5 times greater than that from control spurs. Significant differences between control and NAA-treated spurs were still evident in both cultivars 2.5 days after treatment. B. Effects of Ethylene-Generating Compounds 1. Effects on Initial Set Ethylene reduces set when applied prior to or during bloom (Table 2); Apple fruit set was greatly reduced after applying 200 to 2000 ppm ethephon in the spring at late dormant, pink bud, full bloom and post-bloom stages of development (Amchem Products, Inc., 1969). Edgerton and 35 oHHH>Hmompmmoqu Apmccfinppo>ov 0mm : : ooom = = ficocom AomCOQmop poowv ooom Eooan xoa Amwmav Efimnppmz Amsmflv sexeapmmz councfiz Apoccfizu Lo>ov ooom Eooao Hana a mcmfipopmsx mSOfiOHHmQ somefipxpMum Apoccfinp Amwmav mBOHoHHoQ oopmnoam zaopmzoopmv OOH Eocam Hazm mucwhocfio> 0.0 ooom = m.mH com : wcficmewe .H .m m.sm o xcwm m.m coca : = m.mm o Eooan Hang o.o ooom :. H.0H oom = :.Hm o . xch 0.0 ooom : = . 0.: com : : swammmwwww smoucHoz m.©m o unwaaop commaoo a coupompm Lm>fiuaso mpoumsao AEQQV ucoEQOHm>oc mocmmmmom coaxmufiswm .ocoo we mwmpm .Eooao wcfipdc go on LOHLQ pmfiaodw cos: pom ufispm madam co cocamzum mo muommmm .m oanme 36 Greenhalgh (1969) applied ethephon to 3 apple cultivars at several stages of development from prebloom to harvest. Foliar sprays of ethephon at 200 and 2000 ppm during dor- mant and pink bud stages significantly reduced fruit set at 200 ppm and eliminated it in 'McIntosh' at 2000 ppm. Ap- plication at 1000 to 2000 ppm during the prebloom to early postbloom stages on 'McIntosh', 'Early McIntosh' and 'R.I. Greening' apple completely eliminated all fruit with little or no phytotoxicity (Edgerton and Greenhalgh, 1969; deWilde, 1971). Veinbrants (1979) applied ethephon at 100 ppm at or shortly after full bloom on several apple cultivars. 'Golden Delicious', 'Gravenstein' and 'Jonathan' were thinned adequately when followed by NAA (7.5 ppm). On lighter setting 'Richared Delicious' and 'Starkrimson Delicious' ethephon at 100 ppm resulted in adequate thinning when applied at or shortly after full bloom. Kustermans and Westerlaken (1978) compared hand thinning vs. ethephon treatment on the yield of 'Winston' apple. Ethephon at 2000 ppm applied at full bloom over thinned and reduced yield. 2. Effect on "June" Drop Effects of ethephon upon fruitlet abscission vary with concentration and time of application (Table 3). When applied 10 days after full bloom ethephon at 50 ppm had no effect on final set of 'McIntosh' but 250 and 500 ppm 37 mm com a: ms omm O: O oom mm ms omm mm amps: mm o mm m coca mm II ihmsmav OOOHOLOO HO O ON .Hm OO.OEOO m.mm oom OH WSOfiOHHGQ Won OOH 3H. OOOHOO =.m: O ad m.m oom OH OT 2:. i. l I33: ham Chomppoz 0.:m o :H .Hw um.£wamz o.o om: ma o.ma oma ma m.:m om ma :moOeHOz .m m.OA O ma m.mm omm O: m.mm ms 2: >.Hm o :: m.om omm mm o.mm 0 mm m.m oom OH 0.0H omm OH AOOOHV 0.0m om OH emaecceewo OOOOcHez O.mm O OH O couwewem Lm>fiuaso mooumSHo AEQQV AEooan Hana Lmumm mummy mocoaomom ooa\mpfistm .ocoo :OHOOOOHadm mo mafia .Eooan Hash pound moEHu msofipm> um pofiaddw con: :onmHomnm pmaufizpm madam co condonuo ho muommmm .m magma 38 :Houm:m>mho paw mSOHOHHoQ mempeeefim empeefisfiam AOAOHO emceecoe paste OOe OO OON m: .Om .mm mpeecnefio> pm>fiuaso moopmzao AEQQV AEooao Hana Loumm mmmov monopomom ooa\mufispm .ocoo coaumOfiaddm mo mEHB .OOOOAOOOO .m wanes 39 significantly reduced final set (Edgerton and Greenhalgh, 1969). Concentrations of 75 or 250 applied 28 and 44 days after full bloom had no significant effect. 'Early Mc- Intosh' responded to as little as 50 ppm applied 13 days after bloom, and 450 ppm removed all fruits. Concentrations of 200 to 400 ppm applied 35, 36 and 42 days after full bloom eliminated all fruits on 'Jonathan', 'Richared Delicious', and 'Gravenstein' in Australia (Veinbrants, 1979). Virtually all fruits abscised on 'Cortland' apple when ethephon at 1000 ppm was applied alone or with 1000 ppm Alar-85 26 days after full bloom (Lord,gt gt., 1973). Ethephon at 250 ppm had little effect on 'Mutsu' when ap- plied 35 or 44 days after full bloom; 500 ppm was much more effective at 35 than 44 days. Ebert (1980) applied ethe- phon at 711 ppm 10 and 12 days after full bloom to 'Golden Delicious' and 'Goldparmane' apple fruitlets, respectively. Abscission was not affected in 1977, but was stimulated in 1978. C. Effects of Inhibitors of Ethylene Biosynthesis Because ethylene induces flower abscission, Greene (1980) suggested that endogenous ethylene levels may be sufficiently high in apple flowers to limit fruit set. Application of AVG increases fruit set in some fruits, thus providing support to this hypothesis. Dennis, gt gt. (1978) applied an aqueous spray of AVG at 0, 2500, 5000 40 and 10,000 ppm to swelling buds of apple, cherry and plum. Fruit set of apple was increased 20 to 50% but differences were not significant at the 5% level. Archbold and Dennis (unpublished) reported that fruit set of several cvs. of apple was significantly increased by AVG applied at full bloom at 200 and 500 ppm. Maag (1979) increased fruit set in 'Golden Delicious' apples by spraying early in the spring with 200, 600 and 2,000 ppm of AVG, although 200 ppm was slightly less effective. Concentrations above 3,000 ppm often resulted in severe phytotoxicity. Greene (1980) applied AVG to 'Richared Delicious' apples at full bloom and one day after full bloom. A concentration of 200 ppm significantly increased fruit set, while greatly reducing ethylene production. The inhibitory effect of AVG on ethylene production had dissipated 15 days after treatment. AVG had no effect on viable seeds per fruit. AVG completely overcame GAu+7- and BA-induced ethylene production, reducing it to a level similar to that in the AVG treatment alone. Williams (1980a) applied 1000 ppm AVG to lS-year-old 'Delicious' and 'Golden Delicious' apple trees 2 weeks after full bloom and increased fruit set in both varieties. He also reported that AVG can increase cropping in winter pear trees either by preventing abscission or by increasing fruit set. He also reported that AVG stimulated lateral branching. However, AVG reduced fruit set in cherry and plum (Archbold and Dennis, unpublished; Vecino and Dennis, unpublished). 41 AVG also increases fruit set in other crops, such as bean, and inhibits ethylene production in cucumber. Natti and Loy (1978) reported that emasculation Of hand pollinat- ed muskmelon flowers reduced fruit set in comparison with Open pollination. AVG applied to the base of the calyx of perfect flowers improved fruit set following emasculation; IAA alone improved fruit set slightly, and IAA 4 AVG were more effective than IAA alone, but less effective than AVG .alone at optimum dosage. Foliar application of AVG at 20 or 60 ppm to 2-week-old bean seedlings stimulated growth and fruit set (Shanks, 1980). Application of AVG to soy- bean delayed leaf senescence, stimulated photosynthetic rate and increased yield about 14% (deSilva, personal com- munication). D. Effects of Inhibitors of Ethylene Action Silver nitrate inhibits ethylene action, possibly by competing With ethylene"s metabolic binding site (Beyer, 1976, 1979). Spraying explants Of 'Sprinter Scarlet' geranium inflorescences with silver nitrate effectively reduced petal abscission and slightly promoted ethylene synthesis (Miranda, 1981). Silver thiosulfate was more 'effective and less phytotoxic. 'Delicious' apple flowers treated with AgNO3 produced more ethylene than control flowers, but the effect was not significant (Greene,1980). The same author reported that AgNO3 had no effect on fruit 42 set and injured flowers when used at high concentrations (200 ppm). Applying AgNO3 at 50 to 200 ppm at full bloom to flowers of three apple cultivars ('Wealthy', 'Golden Delicious', and 'Jonathan') had no effect on fruit set (Archbold and Dennis unpublished data). SUMMARY Yields of of 'Delicious' apple are relatively low com- pared with those of other cultivars. Several reasons have been proposed for this including greater susceptibility Of flowers to frost injury, limited ovule longevity, slow growth of pollen tubes in the style, "basal gaps" between stamen filaments which permit bees to extract nectar without pollinating the stigma, low temperature, solar radiation during and immediately after bloom, and high level of endogenous ethylene. These factors may act individually or together to reduce yield in this cultivar. The role of ethylene in apple fruitset has not been critically evaluated. Because AVG both reduces ethylene production and increases fruit set investigators have assumed the later is a result Of the former. However, no studies are known in which cultivar effects on the rate of ethylene production have been examined. Nor have the quantitative effects of chemicals on both set and ethylene production have been carefully compared. VG may exert other effects such as delay of embryo abortion which are 43 more important than its effect on ethylene synthesis. Similarly, although apple fruits which abscise in the "June" drop produce more ethylene than those which do not, and thinning chemicals generally stimulate ethylene produc- tion, a causal relationship has not been established be- tween ethylene production and abscission. Likewise, the hypothesis that basal gaps limit set in the 'Delicious' has not been adequately tested. If this factor is important, why does 'Northern Spy', which exhibits the same characteristics, set so heavily that it is biennial? One way of testing the hypothesis would be to hand pol- linate open-pollinated flowers of both 'Delicious' and a cultivar which does not have basal gaps, such as 'McIntosh'. If the increase in set is greater in the former than the latter, basal gaps would appear to be an important factor in limiting yield. Some Of these approaches were used in studies reported in this thesis. ‘1 10. 11. Literature Cited Abeles, F. B. 1973. Ethylene in plant biology. Academic Press, New York. and B. Rubinstein. 1964. Regulation of ethylene evolution and leaf abscission by auxin. Plant Physiot. 39:963-969. Adams, D. O. and S. F. Yang. 1977. Methionine in plant tissue. Implication of 5-adenosylmethionine as an intermediate in the conversion of methionine to ethylene. 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Plant Physiol. 44:80. Smith, M. V. and O. A. Bradt. 1967. Fruit pollina- tion. Ontario Dept. Agr. and Food Pub. 172. 13 pp. Tukey, R. B. 1978. Secret to more Delicious. Amer. Fruit Grower 98:14, 46. Veinbrants, N. 1979. Further studies on the use of 2-chloroethylphosphonic acid (ethephon) as a thinning agent for apple. Australian J. of Expt. Agr. and Animal Husbandry 19:611-615. Visser, T. and J. J. Verhaegh. 1980. Pollen and pol- lination experiments II. The influence of the first pollination on the effectiveness Of the second one in apple. Euphytica 29:385-390. Walker, D. W., D. R. Paterson and D. R. Earhart. 1979. Silver nitrate ion increases endogenous ethylene in sweet potato vine cutting. HortScience 14:536-537. Walsh, C. 8., J. J. Swartz, and L. J. Edgerton. 1979. Ethylene evolution in apple following post bloom thinning spray. HortScience 14:704-706. 113. 114. 115. 116. 117. 118. 119. 120. 121. 122. 123. 124. 53 Warner, H. L. and A. C. Leopold. 1969. Plant growth regulation by stimulation of ethylene production. Plant Physiol. 44:156-158. Wellington, R. 1947. Pollination of fruit trees Cornell Ext. Bull. 42. 8 pp. Wertheim, S. J., A. Scholten and J. H. Bootsma. 1978. Chemical thinning of some alternate bearing apple cultivars. Fruitteelt 68:558-560 (Hort. Abstr. 48:7891). Westwood, M. N., A. N. Roberts and H. O. Bjornstad. 1976. Influence of in-row spacing on yield of Golden Delicious and Starking Delicious apple on M9 root- stock in hedgerows. J. Amer. Soc. Hort. Sci. 101: 309-311. Whitehouse, W. E. and E. C. Auchter. 1927. Cross- pollination studies with the Delicious apple. Proc. Amer. Soc. Hort. Sci. 23:157. Williams, R. R. 1965. Summary of Research 1964. Pomology and plant breeding. Pollination. Annu. Rept. Long Ashton Res. Sta. for 1964 pp. 17-18. . 1970. The effect of supplementary pol- lination on yield. p. 7-10. In R. R. Williams and D. Wilson (eds). Towards regulated cropping. Grower Books, London. . 1972. Malus species as pollinators for apple. Annu. Rept. Long Ashton Agr. and Hort. Res. Sta. 1971, pp. 26-27. Williams, M. W. 1979. Chemical thinning Of apples. Hort. Rev. 1:270-300. 1980. Retention of fruit firmness and increases in vegetative growth and fruit set of apple with aminoethoxyvinylglycine. HortScience 15:76-77. Wilson, D. and R. R. Williams. 1970. Fruit set past, passive and imperfect. p.1-6. In R. R. Williams and D. Wilson (eds). Towards regulated cropping. Grower Books, London. Way, R. 1973. Apple varieties in New York State. New York State Agr. Expt. Sta. Geneva, New York, In- formation Bull. 63. 125. 126. 127. 128. 129. 130. 131. 54 Yang, S. F. 1974. The biochemistry of ethylene bio- genesis and metabolism. Recent Adv. Phytochem. 7: 131-164. . 1980. Regulation of ethylene biosynthesis. HortScience 15:238-243. Yu, Y. B., D. 0. Adams and S. F. Yang. 1979. Regula- tion of auxin-induced ethylene production in mung bean hypocotyls: Role of 1-aminocyclopropane-l- carboxylic acid. Plant Physiol. 63:589-590. , and . 1979. l- Aminocyclopropane-l-carboxylate synthase, a key enzyme in ethylene biosynthesis. Arch. Biochem. Biophys. 198:280-286. , and 0 1980. In- hibition of ethylene production by 2,4-dinitrophenol and high temperature. Plant Physiol. 66:286-290. and S. F. Yang. 1979. Auxin induced ethylene production and its inhibition by amino- ethoxyvinylglycine and cobalt ion. Plant Physiol. 65:1074-1077. Zimmerman, P. W. and F. Wilcoxon. 1935. Several chemical growth substances which cause inhibition of roots and other responses in plants. Contrib. Boyce Thompson Inst. 7:209-229. SECTION I THE ROLE OF ETHYLENE IN FRUIT SET OF APPLE Abstract. Branch sprays of (2-chloroethyl)phosphonic acid (ethephon), an ethylene generating compound, and amino- ethoxyvinylglycine (AVG), an inhibitor of ethylene syn- thesis, were applied at full bloom to limbs of 'McIntosh', 'Red Prince', 'Delicious' and 'Golden Delicious' apple (Malus domestica Borkh.) to determine their effects on fruit set and ethylene evolution. AVG significantly increased fruit set in all cultivars when used at 200 ppm; at 100 ppm its effects were generally not statistically signifi- cant. AVG tended to reduce ethylene evolution from flowers excised l to 10 days after treatment, but the reduction was small and not statistically significant. Ethephon (40 and 80 ppm) did not affect fruit set significantly, yet markedly increased ethylene production. Evolution of ethyl- ene from ethephon-treated flowers was considerably lower in 'Delicious' than in the other two cultivars. AVG had no effect on length: diameter (L/D) ratio and seed number but reduced fruit weight. Ethephon had no effect on fruit weight, seed number or L/D ratio. Based upon these ob- servations the promotive effect of AVG on fruit set does not appear to be dependent upon its ability to inhibit ethylene synthesis. 56 57 'Delicious' was recognized as a light bearing cultivar as early as 1928 (Howlett, 1928). Several factors may be associated with poor fruit set in 'Delicious'. Frost sus- ceptibility is one of the causes of poor cropping, for 'Delicious' flowers are more susceptible than many other cultivars (Hartman and Howlett, 1954; Roberts, 1946; Meader and Blasberg, 1946; Westwood, gt al., 1976). Solar radia- tion and temperature also may affect set; Gardener, gt a1. (1949) reported that fruit set of 'Delicious' was positively correlated with high radiant energy and high temperatures shortly after full bloom. However, Dennis (1979) analyzed their data and found that correlation coefficients were low and not statistically significant. Flower structure is another factor which has been proposed as the cause of poor set of 'Delicious'. Both Roberts (1945) and Robinson (1980) reported that the structure of 'Delicious' flowers permits honey bees to remove nectar without pollinating the stigma. Both investigators observed "sideworking" of honey bees on 'Delicious' blossoms as a result of "basal gaps", or spaces between stamens. Ethylene is a naturally occurring plant hormone (Leo- pold and Kriedemann, 1975) which is involved in flower and fruit senescence and abscission. Application of ethylene directly or indirectly induces flower and fruit abscission (Abeles, 1973). Ethylene biosynthesis in flowers is stimulated by pollination in cotton (Lipe and Morgan, 1973), 58 blueberry (Hall and Forsyth, 1967), and carnation (Nichols, 1977). In grape the endogenous level of ethylene rises at full bloom, falls at fruit set, then declines during berry development (Singh and Weaver, 1976). Ethylene content of apple flowers increases as flowers develOp, then declines as the fruitlets enlarge (Blanpied, 1972). Application at full bloom of aminoethoxyvinylglycine (AVG), an inhibitor of ethylene synthesis, increased fruit set and reduced ethylene evolution in apple flowers (Greene, 1980). Application of AVG to 'Delicious' apple trees prior to harvest reduced ethylene evolution from opening buds the following spring, and increased fruit set (Williams, 1980). The compound also increased fruit set in bean (Shanks, 1978) and muskmelon (Natti and Loy, 1978). A synergistic effect was found between AVG and promalin, a commercial mixture of N-6-benzyladenine (BA) and gibberel- lins 4 and 7 (GAu+7), on fruit set of 'Delicious' (Greene, 1980). However Dennis (unpublished data) observed that promalin reduced set of 'Red Prince Delicious' when used alone and did not act synergistically with AVG on fruit set. P. B. Lombard (1981) applied AVG at 150, 300 and 600 ppm to 7-year-old 'Comice' pear trees at full bloom and at various times thereafter. Fruit set was increased at all concentrations, but the increase was statistically sig- nificant only at 600 ppm. Ethephon ((2-chloroethyl)phosphonic acid) is an 59 ethylene generating compound which releases ethylene within the treated tissues (Morgan, 1969; Warner and Leopold, 1969). The effect of ethephon on fruit set depends on concentra- tion and stage of flower bud development (Edgerton and Greenhalgh, 1969). When ethephon was applied to 3 apple cultivars during dormant and pink bud stages, 200 ppm sig- nificantly reduced fruit set and 2000 ppm eliminated it in 'McIntosh' (Edgerton and Greenhalgh, 1969). Application at 1000 to 2000 ppm during the prebloom to early postbloom stages on 'McIntosh', 'Early McIntosh' and 'R. I. Greening' apple completely eliminated all fruit with little or no phytotoxicity (Edgerton and Greenhalgh, 1969; deWilde, 1971). Greene (1980) hypothesized that the promotive effect of AVG on apple fruit set was due to its inhibition of ethylene synthesis. This hypothesis implies that the rate of ethylene synthesis is a limiting factor in fruit set under natural conditions. The purpose of this study was to determine if ethylene production by apple flowers does indeed limit set. Materials and Methods Experimental procedure, 1980. Aqueous sprays of AVG or ethephon were applied to branch units of lU-year-old trees of 3 apple cultivars, 'McIntosh', 'Red Prince Delicious', 60 and 'Golden Delicious', at the Horticultural Research Center, East Lansing, Michigan. Five uniform limbs per tree on four mature trees of each cultivar were selected and 75 to 100 flower clusters were counted on each branch at the pink stage. The first 50 clusters were used to evaluate fruit set and develOpment; the remainder were used to measure ethylene evolution. The following solutions containing 0.1% Tween 20 as a wetting agent were applied at full bloom (May 14, 16, and 19, for 'McIntosh', 'Delicious' and 'Golden Delicious', respectively), using a randomized block design with trees as blocks: wetting agent only; AVG at 100 or 200 ppm; ethephone at 40 or 80 ppm. Fruit set was recorded before and after "June drop": and two flower clusters were removed from each branch at intervals of 1 to 10 days after treatment to measure ethylene evolution. The clusters were held with their bases in water in sealed 135 ml containers in the dark at 21°C, and CO2 was absorbed by placing a filter paper wick moistened with 40% KOH in each container. The jars were ventilated after 4 hr and ethylene accumulation was monitored by removing a one ml gas sample through a serum cap after an additional 4 hr. Ethylene was determined using a gas chromatograph (Varian Aerograph Series 1700 or 1400), equip- ped with a flame ionization detector. A column (45 x 0.32 cm) of 60 to 80 mesh A1203 was used at 60° or 80°C. The amount of ethylene in samples was calculated by the following 61 formula: C H (ppm) x PH x vol. (1) -1 -1 2 ”st 3 CzHu (pi-g -hr ) = s ATTSt x PHS t x wts x hr st = standard, s = sample, PH = peak height, ATT = attenua- tion, wt = weight of tissue in g, vol = volume of container (2). Fifteen fruits were harvested at maturity from each treated limb and the weight, length (L) diameter (D) and seed num- ber were determined for each fruit. Experimental procedure, 1981. The experiment was repeated in 1981 using the following treatments: wetting agent only; AVG at 100 or 200 ppm; ethephon at 50 or 100 ppm; and AVG at 200 ppm plus ethephon at 100 ppm. Sprays were ap- plied at full bloom (May 7, 8, and 13 for 'McIntosh', 'Delicious', and 'Golden Delicious', respectively, to branch units of the same 3 cultivars using one branch per treat- ment on each of 3 replicate trees, and fruit set, size and ethylene evolution were determined as in 1980. Frost in April killed 23% of the 'McIntosh', 16% of the 'Delicious' and 2% of the 'Golden Delicious' flowers. Damage was con- fined mainly to king flowers. Therefore set was recorded as fruits per 100 flowers, rather than per 100 clusters. 62 Average orchard temperatures during sampling for 02H” measurement in 1980 and 1981 are shown in Table 1. Results Effects on fruit set and characteristics,l980. Initial set on control limbs was 104, 61, and 53 and final set 33, 35 and 26 fruits per 100 flower clusters for 'McIntosh', 'Red Prince Delicious', and 'Golden Delicious', respec- tively, in 1980 (Table 2). Both initial and final set in 'McIntosh' were significantly higher than in either 'Golden Delicious' or 'Delicious'. 'Delicious' bore significantly less fruits per cm limb circumference than 'McIntosh' or 'Golden Delicious' but fruit retention was not affected. AVG at 200 ppm increased both initial and final set sig- nificantly in all three cultivars, but the effects of AVG at 100 ppm were significant only in 'McIntosh' (Table 2). Similarly, only in 'McIntosh' was fruits per cm limb cir- cumference significantly increased by 200 ppm AVG. Al- though AVG at both concentrations appeared to increase percent fruit retention through the "June drop" in 'Mc— Intosh' the difference was not statistically significant nor was an effect apparent in the other 2 cultivars. At harvest time, fruits on limbs treated with 200 ppm AVG were significantly smaller than the controls in all cultivars, but the effects of 100 ppm AVG were statistically 63 Table 1. Mean of maximum and minimum temperatures (°C) during sampling for CgHu measurement from apple flowers and young fruits at the Horticultural Research Center, East Lansing, MI 1980-81. Cultivar Days after McIntosh Delicious Golden Delicious 616%; 1980 1981 1980 1981 1980 1981 0 11.7 12.2 9.4 7.8 13.3 2.8 1 8.9 3.9 11.7 12.8 13.9 8.3 2 9.4 7.8 12.8 13.9 15.6 10.6 3 11.7 12.2 13.3 3.9 16.7 7.8 4 12.8 13.9 13.9 2.8 18.9 6.7 5 13.3 3.9 15.6 8.3 '21.1 13.3 6 13.9 2.8 16.7 10.6 20.0 7.8 7 15.6 8.3 18.9 7.8 1n.u 8.3 8 16.7 10.6 21.1 6.7 12.2 10.0 9 18.9 7.8 20.0 13.3 17.8 12.8 10 21.1 6.7 14.4 7.8 21.1 17.2 64 a: H.« n owa w bv w H.m 9 mm 9 HF AOOHV o>¢ ©.n a H0. no «ma w 0% w b.N 9 cm 3 mm moose .mfiofiowaon cocaoo. (U a! \0 no -n- -u- -u- a no : o.m : me : ms :86: .88> a S. a 8. a $- 8 Mm a NJ. n mm 9 MI 83 888$ 8 o.s 8 Ho. m 68H a me a s.H n H- p on Ao-V cosamgpm a o.« a mo. n oHH a co m H.m a mo 8 mod Aoomv o>< a m.o a mm. a HnH a Hm a q.m n8 «q as om AooHv o>¢ a «.8 8 Ho. 8 and 8 mm a s.- 9 mm a He 268:0 .mso-ofiama ooeapm 86m. --- --- ---- s o- a H.m a me he on- 886: .aw> n m.s a om. m 08H a.mm p.mqm p.mm o.mmm Aomv nongmg-m n8 «.8 m cm. a Hmfl a me pa m.m 9 08 on mm- Ao-v nonmmg-m no m.s a em. 6 QHH 8 mm a 6.8 a mHH a com Aoomv u>¢ a m.m a be. p sma m cm aw m.m a co nu Ho- Aooav u>< n m.o a as. a sea 8 mm 9 n.H n mm so «OH 98650 4mmo-aH22. guano o-pmp Amy con-ampmn mm-®m\o mmuom\o na-m-\o A5661 \momom axq .ps pagan punch a .ason-o eaHH spam Hanna spam Haw-HaH puma-amb- ao\mefidpm .owma H2 .mawmgwq .m .oammw mo pmm>pmn aw mowpmfipmwomnwno wanna cam moamhomesohfio QEHH So pom wanna .nowpnmpon awnpm .pom awash no aonmogpo can o>< mo mpoommm .N canoe 65 .Ho>oH an .emzn hp meow can mhw>fipazo .mqazaoo Gan-H3 :o-pwnwmom cams» .myopmzao Moro-m oo- poa mpfisnmm -u- -u- -u- a 08 on -.m 6 an 6 ms Aomv Herman-m --- --- --- a so on -.m 6 me o om Aoqv sogamgsm nun uuu nun a no 8 o.¢ m,om m mva Aoomv o>< --- --- --- a Hm pm H.m 8 mm 9 can Aoofiv o>< --- --- --- a me o o.m . 6 an 8 ms 266:0 mcwozflpaoavwoha ..-- -1 --- a R a on a em a so :8: .85 a o:- w 8. a oi a Mm a Nam p m n .mwl 83 8:813 a -.m 8 mm. as mo- 8 mm a n.~ a cm a m- Ao-v conamgpm a m.- 06. o mmfi a me a H.- 6 mm a mad Aoomv o>< coda-paoou.msoH0HHoQ moo-ow. pasta o-pma Amy nausea-mg mm-®m\o mm-om\o AH-MH\8 Aaaav \memmm m\q .-3 paste sauna a .asoafiu naHH N866 Hanna 8666 Han-HaH acme-amps Eo\mpwsmm .emsaa-qoo .m mana- 66 significant only in 'McIntosh' (Table 2). AVG had no sig- nificant effect on either L/D ratio or seed number except in 'McIntosh'; in this cultivar 100 ppm AVG significantly increased seed number per fruit, although 200 ppm did not (Table 2). Ethephon had no significant effect on any of the parameters measured. When data were pooled for all cultivars the effects of AVG on both initial and final set were highly significant at both concentrations, and the effect of 200 ppm was sig- nificantly greater than the effect of 100 ppm (Table 2). Both AVG treatments also increased the number of fruits per cm limb circumference. Effects of ethephon were again non-significant. Effects on ethylene evolution, 1980. AVG generally reduced ethylene evolution from flowers excised within 5 days of application (Table 3), but the differences were statis- tically significant in only one case ('Golden Delicious', 5 days after treatment). Little effect was evident at any time in 'McIntosh'. The burst of ethylene evolution from control 'McIntosh' flowers at 5 days was unexpected; it was not associated with high orchard temperature prior to ex- cision (Table 1). Values for replicate samples were 48.5, 15.9, 8.9, 14.1 n2 per g per hr. Ignoring the highest value, the mean for the remaining 3 samples is 13.0, a more reasonable number in View of other treatment data. Ethephon at 80 ppm significantly increased ethylene 67 Table 3. Effects of AVG and ethephon on ethylene evolu- tion from apple flowers and young fruits 1 to 10 days after treatment, E. Lansing, MI, 1980. Ethylene production (n2/g f.w./hr) Treatment (ppm) 1 day 2 days 3 days 5 days 10 days McIntosh Check .8 oz 3.6 bc 8.6 b 21.9 a 1.4 a AVG, (100) 5.5 c 1.7 c 6.2 b 6.7 a 0.7 a AVG, (200) 4.8 c 4.0 bc .3 b 8.0 a 1.3 a Ethephon (40) 18.4 b 8.8 ab 12.4 b 19.0 a 1.5 a Ethephon (80) 26.2 a 13.7 a 22.2 a 18.1 a 2.5 a Red Prince Delicious Check 4 abc 4.0 ab 8.0 a 6.2 a 3.9 a AVG, (100) .2 c 2.1 be 5.9 a 5.8 a 4.2 a AVG, (200) 1.2 c 1.5 C 3.5 a 6.0 a 2.0 a Ethephon (40) 13.0 a 3.4 abc 9.1 a 7.4 a 9.5 a Ethephon (80) 11.2 ab 4.4 a 10.3 a 7.8 a 4.4 a Golden Delicious Check 5.2 b 6.0 be 6.9 c 4.8 b 1.7 a AVG (100) 2.6 b 3.1 C 4.1 c 0.9 c 1.2 a AVG (200) 3.6 b 5.3 be 3.5 C 0.8 C 0.7 a Ethephon (40) 17.4 a 12.6 ab 13.1 b 10.8 a 2.5 a Ethephon (80) 18.1 a 24.8 a 20.6 a 10.0 a 3.2 a zMean separation within columns and cultivars by DMRT, 5% level. 68 evolution in 'Golden Delicious' and 'McIntosh' flowers col- lected within 3 days of treatment (Table 3); 40 ppm was usually less effective. Effects of the higher concentra- tion were still significant in 'Golden Delicious' at 5 days, but not at 10 days. Although both concentrations of ethe- phon approximately doubled ethylene evolution from 'De- licious' flowers collected 1 day after treatment, dif- ferences were not statistically significant, and values fell to control levels within 2 days. No consistent cultivar differences in ethylene evolution were evident in control or AVG-treated flowers, but 'Delicious' flowers treated with 80 ppm ethgphon evolved significantly less ethylene than did similarly treated flowers of the other cultivars, resulting in a significant cultivar and treat- ment interaction (Table 4). Effects on fruit set and characteristics, 1981. Initial set was 20, 26, and 34 and final set 13, 16, and 9 percent for control flowers of 'McIntosh', 'Delicious' and 'Golden Delicious', respectively (Table 5). Although fruit set was consistently higher following treatment with 200 ppm AVG, differences were significant only in 'Delicious' and 'Golden Delicious' and then only for initial set. Only in 'Golden Delicious' did AVG increase fruit load significantly. However, when data were pooled for the 3 cultivars, the effects of 200 ppm AVG were significant at the 1% level 69 COPHUMLmvcw chEHmmLH x Lm>wu :5 .n_ on sensac_=m_m ._w>o_ um .pmzo an Au .o .4 .mV memos acmEHmmgu x gm>-u-so new .A- .5 .cv memos acme-mmgu .A>v memos gm>_u_=o cmmzpwa m:o_umgmamm N ~92 s o.- = 9m : 9m c 06 :8: > m.m m -.-N a e.¢_ we -.¢ u m.m uo -.o mac-o_—mo coupou > -.m o o.w o m.m u P.N u P.m no N.o mac-o-Foo woe-1a com > m.m a -.o~ a ~.m_ no -.e no m.¢ Nee o.m smoucmoz om oe com oo- comz conqusum w>< Pogucou Lm>-u_:u Asaav acmsummg- .owm_ Hz .mcpmcmn .m .ucmEumoLp mcwzo_-o% mane wmgsu m:_g:c mngopm m-aam an AL:\.3.$ m\-cv co-uoauogq mew-xgum cams co cosmos-m new w>< mo muommwu .e m—nm- 7O --- --- --- a 88 8 8.8 a 88 88 88 88821.88> 8 8.8 8 88. 88 83 8 .nl8 8 Nam. 8 m. 8 .olm. 883 8888858. + Aoomv 8>< 8 8.8 8 8.. 8 888 8 88 8 H.m 8 8H p cm Ace-v 88888888 8 8.8 8 mo. 8 088 8 88 8 8.m 8 8- p Hm A888 88888888 8 8.8 8 mo. 8 8H- 8 O8 8 8.8 8 8m 8 88 Aoomv 8>< 8 8.8 8 8o. 88 8mH 8 88 8 8.8 8 am 88 mm Aooav 8>< 8 H.8 8 mo. n8 88- 8 88 8 m.m 8 8H 8 8m 88888 .888888H8o 888888 888. -u- 1-- ..... a 08 : 8.m e 88 a 8m :88: .88> 8 8.8 8 H8. n.mmm 8.mm_ 8_mqm 8.mm 8.mm Ace-v 88888888 + Aoomv 8>8 8 8.8 8 om. 8 H88 8 88 8 8.H 8 OH 8 cm Aooav 88888888 8 8.8 8 as. 8 88H 8 mm 8 8.8 8 m- 8 8m A888 88888888 8 8.8 8 as. p s-H 8 m8 8 s.m 8 am 8 mm Aoomv 8>< 8 m.s 8 cm. 8 «NH 8 88 8 m.m 8 ma 8 mm Ace-v 8>< 8 8.8 8 as. 8 M88 8 as 8 m.m 8 ma 8 om 88888 28858:. 88:88 88888 A88 888888888 8H-8H\8 s--8-\8 «\8-8m\8 A8888 \88888 o\8 .8: 88888 88888 8 .288888 8888 888 H8888 N888 H88888H 888888889 88\88888 moaohomadonfio .Hm©H H2 .mzfimawq .m .oHQQm mo pmm>hmn pa moapmwhovowAQSQ w-dam cam 98H- Eo hon mafiSMM .aofipcopmn pwshm .988 989nm no conamspo 8:8 c>< mo mwoommm .m manna .Hm>mH Rm .emzm an 8888 8:8 .mpw>888:o .mnesaoo 88:88; nowpwnmmmm awmzx .mhmsoam 008 888 8889888 71 .u. --- ---- 8 88 88 8.8 8 88 8 8m 88888 88888888 + 88888 888 --- -a- ---- 8 88 8 8.m 8 m8 8 88 88888 88888888 --- -u- u--- 8 88 88 8.8 8 m8 8 pm 8888 88888888 .5: In .iuu 8 S 8 8.8 8 8m 8 8.8 803 83. In- nu- u--- 8 88 88 8.8 88 88 8 88 88888 8>< -u- --- ---- 8 mm 88 8.8 8 m8 8 88 88888 8880: 888888889 8 8.8 8 no. ..... 8 8m 8 8.8 8 m8 8 88 8888 .888 8 8.8 8 88. 8 888 8 mm 88 .848.” 8 Mm 88 N8 8888 88888888 + 88888 8>8 88 8.8 8 88. 8 888 8 mm 8 8.8 8 8 8 am 88888 88888888 88 8.8 8 m8. 8 m88 8 mm 88 8.8 8 8 8 88 8888 88888888 8 8.8 8 88. 8 888 8 cm 8 8.8 8 88 8 88 88888 8>8 88 8.8 8 no. 8 888 8 mm 8 8.8 8 88 8 88 88888 888 8 8.8 8 88. 8 888 8 pm 8 8.8 8 8 8 8m 88888 .mzowowaoo nowaou. 88888 88888 888 888888888 88-88\8 88-88\8 m\8.8m\8 88888 \88888 8\8 .88 88888 88888 8 .888888 8888 888 88888 8888 8888888 888888888 88\88888 .888888888 .8 88888 72 for both initial and final set, as well as for fruits per cm. limb circumference. The lower concentration did not have a significant effect. Effects on fruit retention through the "June drop" were again not significant. AVG reduced fruit weight in all cases whether used alone or with ethephon; the reduction was statistically significant at both concentrations in 'McIntosh', neither concentration in 'Golden Delicious' and at 200 ppm only in 'Delicious'. When AVG was applied with ethephon, dif- ferences in fruit weight were significant in 'McIntosh' and 'Delicious', but not in 'Golden Delicious'. All AVG treat- ments significantly reduced seed number in 'Golden De- licious' (Table 5), but L/D ratios were not affected in any cultivar. Ethephon had no significant effect on fruit set, re- tention or characteristics when used alone at either 50 or 100 ppm, except for a reduction in seed number in 'Golden Delicious' at the higher concentration. At 100 ppm ethe- phon prevented a significant fruit setting response to 200 ppm AVG when the two chemicals were applied together. Initial set Of 'Golden Deliciousi was significantly greater than that of 'McIntosh', but not of 'Delicious', but cultivar differences in final set were non-significant. The varietal mean for fruit per cm. limb circumference in 'Golden Delicious' was significantly greater than those of the other cultivars, mainly because of greater flower 73 density and less frost damage; however, fruit retention was significantly lower (Table 5). Effects on ethylene evolution, 1981. Ethylene evolution was generally lower in AVG-treated flowers of all three cultivars, but the effects were statistically significant in only one cultivar at one sampling date ('Golden Delic- ious', 5 days after treatment) (Table 6). Ethephon sig- nificantly increased ethylene evolution in all three culti- vars, although response varied with time of sampling. Ethylene evolution was 2- to lO-fold as high in ethephon- treated flowers as in controls during the 3 days following treatment; after 10 days the difference was significant only in 'Golden Delicious' flowers treated with 100 ppm. AVG treatment had no significant effect on response to ethephon although ethylene levels were generally lower when AVG was used. Ethephon-treated 'Delicious' flowers again evolved significantly less ethylene than did similarly treated 'McIntosh' flowers, and 'McIntosh' significantly less than 'Golden Delicious' following treatment with 100 ppm ethylene, with or without AVG (Table 7). Values for control and AVG-treated flowers were not significantly af- fected by cultivar, confirming the results obtained in 1980. This again resulted in significant interaction between cultivar and treatment. 7“ Table 6. Effects of AVG and ethephon on ethylene production of apple flowers and young fruits as a function Lansing, MI, 1981. of days after treatment, E. Ethylene production (nz/g f.w./hr) Treatment (ppm) 1 Day 2 Days 3 Days 5 Days 10 Days McIntosh Check 6.5 aZ 2.6 9 1.“ be “.2 bcd “.2 ab AVG (100) 1.5 a 1.680 2.3 be 2.7 cd 2.2 bc AVG (200) 6.5 a 1.3 c 0.6 c 1.5 d 2.8 bc Ethephon (50) 19.2 a 13.6 b 2.8 ab 5.5 abc 3.6 b Ethephon (100) 17.6 a 25.8 a “.“ a 7.9 a 5.6 AVG (200) + . Ethephon (100) 19.0 a 20.1 ab 2.8 ab 5.6 ab “.0 b Red Prince Delicious Check 0.9 b 1.6 bc 3.“ ab 3.“ bc 5.2 a AVG (100) 0.5 b 0.5 c 1.7 b 2.2 c 6.8 a AVG (200) O.“ b 0.6 c 1.1 b 2.“ c “.3 a Ethephon (50) 5.0 a 3.3 ab 5.9 a 5.3 a 6.1 a Ethephon (100) 3.9 a “.0 a 5.1 a “.9 ab 5.9 a AVG (200) + Ethephon (100) “.9 a 3.6 a “.8 a “.2 ab 5.3 a Golden Delicious Check 3.2 c “.2 be 13.8 be 9.7 b 2.“ be AVG (100) 1.9 c 1.6 c 3.“ 5.6 be 2.0 bc AVG (200) 1.6 c 1.“ c 3.1 2.1 c 1.8 c Ethephon (50) 12.“ b 11.5 b 26.0 ab 9.5 ab “.“ ab Ethephon (100) 18.8 a 2“.9 a 39.8 a ll;5 ab 5.2 a AVG (200) + 8 Ethephon (100) 1“.“ ab 21.9 a 28.3 a 15.2 a 6.5 a zMean separation within columns and cultivars by DMRT, 5% level. 75 :ovuumgmpcw pcmsammgp x Lm>wupzu .a_ as acmo_a_:mwm ._m>mp um .hmzo x3 Am .u .u .2 .mV cams ucwspmmga x mgo>+upao new .AP .5 .cv magma acmsummga .Ax .3 .>v mamma gm>wupau cmmzuwn m:o_umgnamm~ 5_ m.m_ > «.mp a m.- x m.~ m m.¢ z m.m o m.np cop + com com: cosmmsuu + w>< P m.m_ E N.Fp m m.w~ on m.n_ m N.¢ m o.¢ u m.m_ .uu m... cop om cozqmsam : m., c 5.. .mm .3. w No.o m m.o m -.~ m m._ com co, m>< : m.¢ com: me e.~ m=o_u__mo cme_ow m _.N mzowuppoo mu:_ga vwm Na m.m smou:_uz Fogpcou Lm>pupao Assn“ acmEummgh ._wm_ a: .ocamcss .m .ucmsummgu mcpzoppom mxmu mags» mcmgzc mgozoFm opaga an AL;\.z.w m\_=v :o_uuzuoga mcmpxgum cams co cosamcum new w>< mo muomymm .m mpnmp 76 Discussion The data obtained in this study confirm previous re; sults (Dennis, 1978; Archbold and Dennis, unpublished; Greene, 1980, Williams, 1980) that AVG increases fruit set and inhibits ethylene bioSynthesis in apple flowers. However, its fruit setting effect does not appear to be dependent upon inhibition of ethylene synthesis as pro- posed by Greene (1980) and Williams (1980). They sug- gested that apple fruit set is limited by the rate of ethylene synthesis in the flower. If this were true, re- ducing the rate of ethylene production should promote fruit set and increasing the rate should reduce set, i.e., fruit set should be negatively correlated with ethylene produc- tion. My data relating fruit set to ethylene evolution are summarized in Figure 1. High levels of ethylene evolution were not induced in 'Delicious' by treating with ethephon. However, in 'McIntosh' and 'Golden Delicious' high levels of ethylene did not reduce set. Furthermore, although AVG concentration had little effect upon ethylene evolution, set almost invariably increased as the concentration of AVG increased from 100 to 200 ppm. Correlation coefficients for initial fruit set vs. ethylene evolution were not statis- tically significant for any cultivar in 1980 or 1981 (Table 8). However, the r values for ’Delicious' were high and 77 Figure 1. Relationship between mean ethylene evolution from flowers the first three days after treatment and initial fruit set of apple. INITIAL FRUITS PER 100 CLUSTERS INITIAL FRUITS PER 100 FLOWERS 78 FRUIT SET 1980 ZSIT 200- wo- mu- 5'” Lamp '9'“. " -onu . L l . . A l x . . * 0.0“ 'o ' h ' '12 ' '1: ' '24 so FRUIT SET 1981 50!" ' ‘G- 3&- 1. Iq‘ ’ 1 l ,4; 2‘1' é ”” ‘R a \B 1% ‘9' M:- -o&. . «homfl : “2 : “a i 24 ETHYLENE PER G PER HR Figure l 79 Table 8. Relationship between initial fruit set of apple and mean ethylene evolution from flowers during 3 days following chemical treatment. Correlation coefficientz Cultivar 1980 1981 McIntosh -0.38 -0.33 Delicious -0.77 -0.76 G. Delicious -0.57 -0.58 ZNo values significant at 5% level. Five (1980) or 6 (1981) pairs of observations per cultivar. 80 might have been significant with more observations. Thus there appears to be a negative correlation between fruit set and ethylene evolution in 'Delicious' but not in 'McIntosh' or 'Golden Delicious'. Therefore the hypothesis that ethylene level controls fruit set may still be true for 'Delicious'. Untreated 'Delicious' flowers could produce more ethylene than do 'McIntosh' and 'Golden Delicious' flowers and therefore set comparatively poorly. However, the data (Tables “ and 7) indicate no consistent cultivar differ- ences in ethylene production in untreated flowers in 1980, and a lower (difference non-significant at 5%) production in 'Delicious' in 1981. Therefore this hypothesis is not valid. There remains the possibility that 'Delicious' flowers ’ are more sensitive to ethylene levels. If this were true, one should expect lower fruit set in 'Delicious' than in other cultivars at the same endogenous level of ethylene. Although the ratio of fruit set to ethylene production was higher for 'McIntosh' than for 'Delicious' in 1980, it was similar in 1981 (Figure 1). Likewise, the ratio was higher for 'Golden Delicious' in 1981, but similar in 1980. There- fore, this hypothesis too does not appear to be valid. Low concentrations (“0 to 100 ppm) of ethephon markedly increased ethylene evolution without affecting set ap- preciably. Higher concentrations (200 to 1000 ppm) of 81 ethephon probably would have reduced fruit set (see Edger- ton and Greenhalgh, 1969). I conclude that the rates of ethylene production measured in this study have little or no effect on fruit set of ap- ple. AVG had no significant effect on set when applied with ethephon, nor did it reduce the effect of ethephon on ethylene evolution from the flowers. The combined ef- fects of AVG and ethephon on fruit set are difficult to explain. If the effect of AVG is not mediated by ethylene, and if ethephon alone does not decrease set, application of AVG with or without ethephon should increase set. However, application of ethephon at 100 ppm eliminated the effect of 200 ppm AVG (Table 5), at least in 'McIntosh' and 'Delic- ious'. Perhaps a high level of ethylene within the tissue blocks the promotive action of AVG without directly reduc- ing set. This possibility should be explored in other systems. If the petals were the major source of ethylene, the ethylene production measured in this study would not have been that which is crucial in regulating set. However, petal removal had no consistent effect upon rate of ethylene evolution (Table 9). Therefore, most of the ethylene measured was probably evolved from other flower tissues. Nichols (1977) observed that approximately “0 to 50% of the ethylene evolved by carnation flowers could be ac- counted for by the styles and most of the remainder by 82 Table 9. Effect of petal removal on ethylene production by apple flowers, 1981. Ethylene production (nA per g per hr)z 1 day AFBy 2 days AFB Cultivar Petals: Intact Removed Intact Removed McIntosh 6.62x 0.91 2.57 3.57 Delicious 0.88 0.82 1.6l 1.02 Golden Delicious 3.20 3.uo “.19 “.87 zTreatment differences not significant, 5% level. yAfter full bloom. XMeans for l limb on each of 3 trees in RCB. 83 the petals. Because the styles contributed less than “% of the fresh weight of the flowers, they were considered to be the most active centers of ethylene production. Ethylene production by the style, ovary tissue or ovules might be better correlated with fruit set than production by the entire flower. Cultivar differences were evident in response to ethe- phon treatment, treated flowers of 'McIntosh' and 'Golden Delicious' producing much more ethylene than those of 'Red Prince Delicious'. Tissues with different acidities might be expected to show different capacities for ethylene evolu- tion (Warner and Leopold, 1969). Tissues of 'Delicious' flowers may have a lower pH than those of 'McIntosh' and 'Golden Delicious', resulting in less breakdown of ethephon and therefore slower release of 02H“. Breakdown of ethephon is highly dependent on tempera; ture (Olien and Bukovac, 1978; Amchem Products, Inc., 1969). In this study variation in ethylene production by flowers treated with ethephon did not appear to be an ef- fect of temperature differences (Table 6). 'Differences in ethylene evolution do not appear to (be responsible for differences in initial fruit set. and the effects of AVG in increasing set apparently are in- dependent of its effects on ethylene synthesis. The mechan- ism of action of AVG in stimulating fruit set remains to be determined. The chemical could increase ovule longevity and this possibility is explored in a subsequent study. 10. 11. Literature Review Abeles, F. B. 1973. Ethylene in plant biology. Aca- demic Press, N. Y. Archbold, D. and F. G. Dennis, Jr. 1979. Effects of ethylene inhibitors on apple fruit set. Unpublished data. Baker, E., M. Lieberman, and J. D. Anderson, 1978. Inhibition of ethylene production in fruit slices by a rhizobitoine analog and free radical scavengers. Plant Physiol. 61:886-888. Blanpied, G. D. 1972. A study of ethylene in apple, red raspberry and cherry. Plant Physiol. “9:627-630. Boller, I., R. C. Herner, and H. Kende. 1979. Assay for and enzymatic formation of an ethylene precursor, l-aminocyclo-propane-l-carboxylic acid. Planta (Ber— lin) l“5:293-303. Dennis, F. G., Jr., C. E. Crews, and D. W. Buchanan. 1978. Effect of aminoethoxyvinylglycine on bloom delay and fruit set in tree fruits. HortScience 12:386 (Abstract). Edgerton, L. J. and W. J. Greenhalgh. 1969. Regulation of growth, flowering and fruit abscission with 2— chloroethanephosphonic acid. J. Amer. Soc. Hort. Sci. 9“:11-13. Greene, D. W. 1980. Effect of silver nitrate, amino- ethoxyvinylglycine and gibberellins A“+7 plus 6- benzylamino purine on fruit set and development of Delicious apple. J. Amer. Soc. Hort. Sci. 105: 717-720. Hall, I. V., and F. R. Forsyth. 1967. Production of ethylene by flowers following pollination and treat- ment with water and auxin. Can. J. Bot. “5:1163-1166. Leopold, A. C. and P. E. Kriedemann. 1975. Plant growth and development. McGraw—Hill, Inc. 5“5 pp. Lipe, J. A. and P. W. Morgan. 1973. Location of ethylene production in cotton flowers and dehiscing fruits. Planta (Berlin) 115 93-96. 8“ 12. 13. l“. 15. 16. 17. 18. 19. 20. 21. 22. 85 Lombard, P. B. 1981. WRCC-l7 Oregon report unpublished data). Morgan, P. W. 1969. Stimulation of ethylene evolu- tion and abscission in cotton by 2-chloroethanephos- phonic acid. Plant Physiol. ““z337-3“l. Natti, T. A. and J. B. Loy. 1978. Role of wound ethylene in fruit set of hand-pollinated muskmelons. J. Amer. Soc. Hort. Sci. 103:83“-836. Nicholas, R. 1977. Sites of ethylene production in pollinated and unpollinated senescing carnation (Dianthus caryophyllus) inflorescences. Planta (Berlin) 135:155-159. Olien, W. and M. J. Bukovac. 1978. The effect of tem- perature on rate of ethylene evolution from ethephon and from ethephon treated leaves of sour cherry. J. Amer. Soc. Hort. Sci. 103:199-202. Shanks, J. B. 1980. Promotion of seedling growth with AVG. Growth Regulator Bull. 8:5-6. Singh, I. S. and R. J. Weaver. 1976. Effect of girdl- ing and gibberellic acid on endogenous level of ethylene in Black-Corinth grapes. Haryana J. Hort. Sci. 5:150-153. Warner, H. L. and A. C. Leopold, 1969. Plant growth regulation by stimulation of ethylene production. Plant Physiol. ““:156-158. Williams, M. W. 1979. Chemical thinning of apple. Hort. Rev. 1:270-300. . 1980. Retention of fruit firmness and increase in vegetative growth and fruit set of apple with aminoethoxyvinylglycine. HortScience 15:76-77. Yang, S. F. 1980. Regulation of ethylene biosynthesis. HortScience 15:238-2“3. SECTION II RELATIONSHIP BETWEEN ENDOGENOUS ETHYLENE EVOLUTION AND APPLE FRUIT ABSCISSION DURING "JUNE" DROP 86 Abstract. Sprays of aminoethoxyvinylglycine (AVG), an inhibitor of ethylene synthesis, silver thiosulfate (STS), an inhibitor of ethylene action, and (2-chloroethyl) phos- phonic acid (ethephon), an ethylene-generating compound, were applied to branch units of 'McIntosh', 'Red Prince Delicious', and 'Golden Delicious' apple (Malus domestica Borkh.) trees 18 or 21 days after full bloom. The bark on additional limbs was scored with a knife. AVG (200 ppm) had no significant effect on ethylene evolution in any of the cultivars, but significantly reduced fruit retention in 'McIntosh' in 1980. Scoring and STS treatment affected neither ethylene production nor abscission. Ethephon (200 ppm) significantly increased fruit abscission in all 3 cultivars, but 100 ppm was effective only in 'McIntosh' and 'Golden Delicious'. Ethylene evolution was measured in two populations of fruits differing in diameter. Small fruits generally produced more ethylene per unit weight and their abscis- sion potential was higher than large fruits sampled at the same time. However, ethylene production of untreated fruits appeared to be a function of fruit size rather than of abscission potential EE£.§S- The data suggest that ethylene production is not the primary factor con- trolling abscission of apple fruits during the "June" drop. 87 88 Two abscission periods are usually recognized in apple trees, one termed "first drOp" immediately following bloom and the other "June" drop approximately 2 to 3 weeks after bloom (Gourley and Howlett,l957; Teskey and Shoemaker, 1972; Childers, 1969). Several factors have been sug- gested as being responsible for the second drop, including low seed number and competition between fruits for food materials (Gourley and Howlett, 1957; Teskey and Shoemaker, 1972). Heinicke (1917) reported that apples which fell from the tree contained fewer developing seeds than those which remained on the tree. However, the mechanism of fruit abscission is still not well understood. Since the discovery of ethylene as a natural plant hormone and its action in fruit abscission, several in- vestigators have suggested that it plays a role in "June drop" (Dennis, 1970). Blanpied (1972) found that during the June drop the fruit pedicel of 'McIntosh' and 'Delicious' apple contained 3- to lO-fold more ethylene per unit weight than did fruit flesh tissues and that tissues of abscising fruits pedicel and flesh tissues did not consistently contain more ethylene than similar tissue of adhering fruits. Certain chemicals are effective in either stimulating or inhibiting the "June" drop in apple. Synthetic auxins such as naphthaleneacetic acid (NAA) and its derivatives have been used as thinning agents in apple for many years. The mechanism of action of NAA is still not well understood, 89 but one hypothesis is that application of NAA stimulates ethylene synthesis and that the ethylene produced induces abscission of immature fruits (Dennis, 1970; Schneider, 1975; Walsh, 33 al., 1979). Several investigators have reported that NAA does indeed stimulate ethylene evolution in apple fruits. Schneider (1975) found that spraying with NAA (25 ppm) “ days after petal fall stimulated both fruit abscission and ethylene evolution in leaves, fruits and pedicels of 'Golden Delicious', 'Staymared' and 'Red Rome' 'apple sampled 2“ hours and “8 hours after spraying. He suggested that NAA-induced ethylene evolution caused fruit abscission. Walsh,gt a1. (1979) sprayed branches of 'Golden Delicious' and 'Northern Spy' apple with NAA (15 ppm) two weeks after petal fall. One day after applica- tion ethylene evolution from 'Golden Delicious' spurs was 5 times greater than that from controls, and the chemical significantly thinned both cultivars. Williams (1980b) applied NAA at 5 and 10 ppm to 'Delicious' apple two weeks after full bloom. NAA at both concentrations significantly increased ethylene evolution 2“ hrs. after application and reduced fruit set. Ethephon ((2-chloroethy1)phosphonic acid), which re- leases ethylene within the tissues, also can be used as a thinning agent. Its effectiveness on apple fruits depends on the time of application and the cultivar. Walsh,gt a1. (1979) successfully thinned 'Golden Delicious' and 'Northern Spy' by applying 200 ppm 2 weeks after full bloom. When applied 10 days after full bloom 50 ppm ethephon did not thin 'McIntosh' fruits, but 250 and 500 ppm were effective; 75 and 250 ppm applied 28 and ““ days after full bloom_ had no significant effect (Edgerton and Greenhalgh, 1966). However, concentrations of 200 to “00 ppm applied 35 and “2 days after full bloom eliminated all fruits on 'Jonathan', 'Richared Delicious' and 'Gravenstein' trees in Australia (Veinbrants, 1979). Walsh,et 31. (1979) sprayed 'Golden Delicious' and 'Northern Spy' apple fruits with 200 ppm ethephon two weeks after petal fall. Ethylene evolution from spurs of 'Golden Delicious' was 7 times higher than in control spurs and the chemical significantly thinned both cultivars. Williams (1980a) applied 1000 ppm of aminoethoxyvinyl- glycine (AVG), an inhibitor of ethylene synthesis, to 'Delicious' and 'Golden Delicious' apple 2 weeks after full bloom. The treatment complete by inhibited "June" drop and Williams suggested that the effect on drop might be a result of reduced ethylene synthesis. AVG (“00 ppm) counteracted the effects of NAA (10 ppm) on both ethylene evolution and fruit abscission (Williams 1980b). Ratescxfethylene evolu- tion were “.“, 6.7, 1.8, and 1.5 ul per g per hour for control, NAA alone, AVG alone and AVG + NAA, respectively, and fruit set values (fruits per 100 clusters) were 89, “9, 116, and 80, respectively. Note that fruit set in the last 91 two treatments differed markedly, yet ethylene evolution was identical. AVG (200 ppm) applied to 'Golden Delicious' fruitlets prior to treatment with NAA (58 ppm) did not reduce the effect of the lattercn1ethylene synthesis (Ebert 1980). Silver nitrate and silver thiosulfate inhibit ethylene action rather than its biosynthesis (Burg and Burg, 1967; Reid,et al., 1980; Miranda, 1981). Spraying pea seedlings with silver nitrate effectively blocks the ability of exo- genous ethylene to induce the classical "triple" response --- growth retardation, stem swelling and horizontal growth. Treatment with AgNO3 blocks leaf abscission in cotton (Beyer, 1976) and delays senescence as well as counteracting the effects of ethephon in excised carnation flowers (Halevy and Kofranek, 1977). In banana fruit slices, AgNO3 significantly reduced ethylene production (Saltveit,;t‘al., 1978). However, sweet potato (Walker, et al., 1979) and apple (Greene, 1980) tissues treated with Ag+ produced more ethylene than the control. Several reports indicate that ringing or scoring limbs before or two weeks after full bloom can also reduce the severity of the "June drop" in apple (Murneek, 1937; Griggs and Schrader, l9“1; Batjer, 1962; Dennis and Edgerton, 1965) possibly by preventing translocation of reserve carbohydrates from the treated branch. Some evidence in- dicates that ringing alters endogenous ethylene levels 92 (Singh and Weaver, 1976). Girdling of grape canes promoted ethylene volution from the fruit at all stages of berry development (Singh and Weaver, 1976). Weaver, et a1. (1972) showed that ethephon moved from leaf to the shoot tip and suggested that the compound was translocated from source to sink. Hale and Weaver (1962) had previously shown that growing berries attract assimilates and ethylene from the foliage. The purpose of this study was to evaluate the role of endogenous ethylene production in controlling the "June drop" of 'Delicious', 'Golden Delicious' and 'McIntosh' apple fruits. Both stimulators and inhibitors of ethylene evolution were used and their effects upon abscission vs. ethephon evolution were compared. Materials and Methods In 1980 mature 'Red Prince Delicious', 'Golden De- licious', and.'McIntosh' apple trees were used and 6 uni- form limbs were selected on each of 3 trees per cultivar. Eighteen days after full bloom (FB) one of the following treatments was applied to one branch on each tree: double scoring through the bark with a knife with scores 2 cm apart; AVG at 50 or 200 ppm; ethephon at 100 or 200 ppm. One hundred fruits on each limb were counted for evaluation of fruit retention. Fifty control fruits of 'Delicious', 93 'McIntosh' and 'Golden Delicious' and 50 'Golden Delicious' fruits treated with 200 ppm AVG and ethephon were identified with numbered tags on each tree, and fruit number and di- ameter were recorded at intervals. Five small and five large fruits in the population present on a given date were harvested from each tagged limb l, 3, 6, 9 and 12 days after application. These small and large fruits were held in separate sealed 135 ml containers in the dark at 21°C and CO2 was absorbed by placing a filter paper wick moisten— ed with “0% KOH in each container.' The Jars were venti- lated after “ hours and ethylene accumulation was monitored by removing a one ml gas sample through a serum cap after an additional “ hour period. Ethylene was determined using a gas chromatograph (Varian Aerograph Series 1700 or 1“00), equipped with a flame ionization detector. A column (“5 x 0.32 cm) of 60 to 80 mesh A1203 was used at 60° or 80°C. The amount of ethylene evolved per g tissue per hour was calculated by the following formula: C H (ppm) x PH x vol(£) _l. _1 2 ”St S Hu (pi g .hr ) s ATTst x PHs C 2 t x wts x hr standard; 5 = sample; PH = peak height; ATT = st attenuation; wt weight in g, vol = volume of con- tainer (A). 9“ In 1981 treatments were applied in a similar experi- ment using 3 trees each of 'McIntosh' and 'Red Prince De- licious'. The following treatments were applied three weeks after full bloom; AVG, 200 ppm; ethephon, 100 or 200 ppm; AVG, 200 ppm, plus ethephon, 200 ppm; silver thio- sulfate (STS), 100 ppm. The STS solution was prepared by adding 1 M AgNO3 to an equal volume of “ M Na28203'5H20 with rapid stirring. Diameters of fruits receiving no treatment, 200 ppm AVG or 200 ppm ethephon were measured to the nearest mm at 3 day intervals for 18 days beginning on the day of treatment. Ethylene production by large and small fruits was measured as in 1980. Orchard tempera- tures during the sampling period were recorded for both years (Table 1). Results 1980. Double scoring did not affect fruit retention significantly in any of the 3 cultivars (Table 2), although fruit number was reduced in 'McIntosh', and increased in 'Delicious'. Because interaction between treatment and fruit size was non-significant, only main effects on ethyl- ene evolution are presented (Tables 3 and “). Although ethylene production was generally lower in fruits from scored limbs than in control fruits of 'McIntosh' and 'Golden Delicious', the reduction was not significant 95 Table 1. Mean of maximum and minimum temperatures (°C) during time of sample collection for ethylene determination in apple fruits at the Horticultural Research Center, East Lansing, MI. Red Prince Golden Days after McIntosh Delicious Delicious treatment 1980 1981 1980 1981 1980 1 15.0 16.1 15.0 12.8 10.6 2 17.2 19.“ 10.0 12.8 7.8 3 17.2 12.8 10.6 15.6 12.2 18.9 12.8 7.8 18.9 17.2 5 15.0 15.6 12.2 20.0 20.0 6 10.0 18.9 17.2 16.7 19.“ 7 10.6 20.0 20.0 21.7 11.7 8 7.8 16.7 19.“ 16.7 11.7 9 12.2 21.7 11.7 21.1 15.6 10 17.2 16.7 11.7 19.“ 16.7 11 20.0 21.1 15.6 17.8 15.0 12 19.“ 19.“ 16.7 16.7 15.0 96 .Ho>oa mm .Bmzm an mcofium>pomno new mpm>fiuaso Gazpaz coaummmdom :mozz .Amsoaoaaoa coodoov om oo H: coo Amzofiofiaoo oocaoa oomv as oo om .AomoocHozv an op om moaoee ao noose: Hofioaoe coozo o ~.m: o ~.m: o o.m: o m.om o o.mm o 0.0m om\~ Assv nooosoao passe o m.om o H.mm o 3.3: o o.mm a m.mm oo m.mm meo lav oofioooooo pasta N.ozoeoeeoa cooeoo. o o.Hm o o.Hm o m.m: o m.m: o 0.0m o m.m: mmxa Aesv oooosofio passe o o.mm o H.ma o m.m> oo ~.mo om o.wm oo m.om Hm\© ARV cofiooooon pasta N.oooeoaeon oocfina oom. o o.mm o m.mm o m.mm o o.mm o o.mm o e.mm mm\a Asev pooosoao passe o m.m o m.HH oo m.om o m.mH no H.m: Kno 0.5m om\m ARV cofiocoooo passe N . 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AVG at both concentrations significantly reduced fruit retention in 'McIntosh' (Table 2). Although AVG increased retention in both 'Delicious' and 'Golden Delicious', the differences were significant only in 'Delicious' and then only at 200 ppm. Effects on fruit size were small and in- consistent. AVG at 200 ppm appeared to reduce ethylene evolution in all 3 cultivars (Tables 3 and “), but the reduction was significant only in 'Golden Delicious' 1 day after treatment, even when means for several days were compared. At 50 ppm, AVG generally reduced ethylene evolu- tion, but differences were non-significant. Ethephon at both 100 and 200 ppm significantly in- creased fruit drop in 'McIntosh' but not in 'Delicious' or 'Golden Delicious'. Although drop was heavier on limbs of the last two cultivars which were treated with 200 ppm ethephon, the effects were non-significant (Table 2). In 'Delicious? the low concentration actually reduced drop significantly. Effects upon fruit size were small and non- significant. At the higher concentration, ethephon in- creased ethylene evolution from treated fruits regardless of cultivar or time of sampling except in 'McIntosh' fruits sampled 6 days after treatment (Table 3 and “). The ef- fects of 100 ppm were similar, but with fewer significant differences. Small 'McIntosh' and 'Delicious' fruits evolved sig- nificantly more ethylene per g fresh weight than large 100 ones with few exceptions, but fruit size had no significant effect on ethylene production in 'Golden Delicious'. 1981; AVG tended to increase fruit retention in both cultivars, but the differences were not statistically sig- nificant (Table 5). Seed number, fruit size, and L/D ratio were not significantly affected in either cultivar, although 'McIntosh' fruits treated with both AVG and ethephon ap- peared to be smaller. Data on ethylene evolution from large and small fruits were again pooled, as interaction between treatment and fruit size was non-significant. AVG alone generally reduced ethylene production, but the ef- fect was not significant, even when data for several sampling dates were pooled (Table 6). Ethephon alone did not affect fruit retention sig- nificantly in either cultivar (Table 5), nor did it alter seed number, fruit weight, or L/D ratio. Ethephon at 200 ppm consistently increased ethylene evolution; effects of 100 ppm were intermediate and were significant on 3 ('Mc- Intosh') or “ ('Delicious') of the 5 sampling dates (Table 6). AVG plus ethephon reduced fruit retention, but the effects were significant only in 'McIntosh' (Table 5). Treated fruits of both cultivars were smaller than con- trols, but effects on size and L/D ratio were non-sig- nificant. Seed number was significantly reduced in 101 Table 5. Effects of AVG, ethephon and silver thiosulfate (STS) on fruit abscission and fruit characteris- tics at harvest of 'McIntosh' and 'Red Prince Delicious' apple, 1981. Treatments applied May 28 ('McIntosh'), or 30 ('Delicious'). Fruit Treatment Retention No.seeds Fruit L/D (ppm) (%)6/l6 Per fruit wt (g) ratio 'McIntosh'Z None 36 ab 6.9 a 1“7 a 0.78 a AVG (200) 50 a 7.“ a 122 a 0.80 a Ethephon (100) 29 bc 7.2 a 150 a 0.79 a Ethephon (200) 26 bc 7.“ a 135 a 0.83 a AVG (200) + Ethephon (200 18 C 5.7 b 137 a 0.81 a STS “1 ab 7.0 1“3 a 0.77 a 'Red Prince Delicious'Z None 71 a “.1 a 1“8 a 0.91 a AVG (200) 88 a “.1 a 152 a 0.89 a Ethephon (100) 70 a “.“ a 151 a 0.93 a Ethephon (200) 62 a “.8 a 155 a 0.88 a AVG (200) + Ethephon (200) 60 a . a 133 a 0.86 a STS (100) 7“ a “.3 a 1“1 a 0.92 a zMean initial fruit numbers 90 to 97 ('McIntosh') and 6“ to 81 ('Delicious'). yMean separation within columns and cultivars by DMRT, 5% level. —~r. .Ho>oH mm .Bsz an mmNHm new mcesaoo .mao>HoH:o Gonna: coaumpooom coozu 102 a :H.m a :0.m a 0m.0 05.0 s ma.m 0 onaooe owoon a ma.: a Hm.= a 00.0 0:.H a 02.0 0 onfisoo HHoEm pom com: o m0.m oo mn.m mm.0 o ma.0 n mm.m o no new no on.m no ou.0 05.0 o A0.H n me.m o o A00m0 oonnonnm + A00m0 0>< o 0A.0 o 05.5 m0.H o m0.m o He.a o o A00m0 nonoonnm n am.= on no.2 0m.0 n :H.H o Hm.: n no A00HV :onnonnm o 00.0 o 05.0 00.0 o 00.0 n 0N.H o n A00mv 0>< oo mm.H o 0m.H mm.0 o 00.0 n 0N.H o n noono soaoaaon oonfion com. 5 No.3 2 0:.m a 0m.0 a nm.a s ~0.H a a manage owoon a 50.0 a 0m.a 0 Hm.0 a m0.fi 0 No.0 0 0 ooasoe Heoam now cow: 0 mm.m o 00.H o mm.0 n 0:.H n 00.0 o o mam no 00.0 no 0m.HH n 00.0 n m0.0 no mm.m n no A00mv conoonnm + Aoomv c>< o 0o.0H o m0.me o 00.0 o mm.m o mm.= o o A0000 connonnm n no.0 n me.» n no.0 n 00.H n H0.m n on A00Hv nonoonnm o mm.H o am.H o :m.0 n 03.0 o 00.0 o o A00N0 0>o o mm.n o m0.n o 0H.0 n No.0 o HA.0 o No xoono .nooooHoz. 0+m+H m+H ma 0 0 Agony psoEumoLB uCoEpmmpB Loum< mama no .Aanchozv mm an: oofiaooo mucoEummpe .AmSOHOfiHoQ oocfipm pomv om .mea done zoc:h= on pofipo mafiSLM madam .msofiofiaom monopm pom. new .cmOchoz. 80pm Ap£\w\a:v cofiusao>o ocoamnuo so mummasmOHSp ao>HHm cam conoocpo .Ao>zxonpoocfiem mo mpoommm .o magma 103 'McIntosh', but not in 'Delicious'. Ethylene evolution was consistently reduced in comparison with ethephon alone with one exception ('Delicious' at 3 days), differences being significant in 5 of 10 cultivar-date combinations (Table 6). However, values were significantly higher than control values in all but two cases ('McIntosh', 9 days, 'Delicious', 6 days after treatment). Silver thiosulfate did not significantly affect fruit retention, fruit size, seed number or L/D ratio in either cultivar (Table 5), and ethylene production was affected (increased) significantly in only one case ('McIntosh', 6 days) (Table 6). Small fruits evolved consistently more ethylene per g fresh weight than did large ones, and differences were significant in 10 of l“ comparisons (Table 6). In the previous paper I observed that 'Delicious' apple flowers consistently evolved less ethylene following treat- ment with ethephon than did similarly treated 'McIntosh' or 'Golden Delicious' flowers. However, cultivar dif- ferences in the response of developing fruitlets was in- consistent. Treated 'Delicious' fruitlets evolved more ethylene in 1980, but less in 1981, than did 'McIntosh' fruitlets (Tables 3, “, and 6). Abscission data obtained from untreated, tagged fruits which were measured periodically in 1980 indicated that abscission potential decreased as fruit size increased 10“ (Figure l) as previously demonstrated by Zucconi (1975, 1978) for peach and orange fruits. In 'McIntosh', for example, fruits 9 mm or less in diameter as of June 3 had all abscised by June 15, whereas no fruits 15 mm or greater in diameter had fallen. As fruit diameter increased from 10 to 1“ mm, fruit drop decreased. Data for 'Delicious' and 'Golden Delicious' fruits were parallel, but abscission potential was greater than for 'McIntosh' fruits of similar size. To determine the relationship between ethylene evolu- tion and abscission potential, the fruits used for ethylene determination were stored in plastic bags at “°C for two weeks. The diameter and weight of each fruit was then measured and a curve constructed relating the two (Figure 2). This curve was used to convert the mean weights of both large and small fruits to mean diameters for each sampling data (Table 7). Data for abscission potential obtained from tagged fruits were then plotted against diameter of the fruits remaining at each sampling date (Figure 3) and the abscission potentials of large and small fruits estimated from these graphs using their mean diam- eters as indices (Table 7). The data indicate that small fruits generally had both a higher abscission potential (Table 7) and a higher rate of ethylene production (Table 7, Figure “A and C) than did large fruits. These differences in ethylene evolution could be responsible for differences in abscission potential. On 105 Figure 1. Effect of cultivar on relationship between fruit diameter on June 3 (McIntosh), 8 (Delicious) or 9 (Golden Delicious) and abscission during "June” drop 1980. Surviving fruits counted June 15 (McIntosh), 20 (Delicious) or 21 (Golden Delicious). 106 09 a opswfim aEEv honorams :35“. a s n. 0 o _.. .a 2.22.096 25.0.30. coEood . 5350.26 n. o .ION .0? do 60 :00, uogssgosqv 95 107 Figure 2. Relationship between diameter and weight of 'McIntosh' fruits in 1980 and 1981. 108 m opswfim A3 2225 :3... m V «a: «pm 00o.. 50. ...-..--. 0 dl - u up“ an“ .-F €20.22 .oN (mm) Jemmegp mug 109 Table 7. Average fruit weight, diameter, abscission po- tential and ethylene production of untreated control apple fruits prior to and during "June" drOp, 1981. Mean Mean 2 C H Sampling Weight Diameter Abscissiony 2 “ Date (g) (mm) Potential (%) (nZ/g/hr) McIntosh Small fruits 5/29 1.0 11.0 2“ 1.22 5/31 0.9 10.5 68 3.03 6/3 1.9 1“.8 “5 0.73 6/6 2.7 16.“ 62 0.8“ 6/9 “.8 21 6 0.2“ Large fruits 5/29 1.9 l“.8 0 1.21 5/31 1.6 13.6 25 1.03 6/3 2.3 15.8 28 0.69 6/6 3.9 19.0 15 0.39 6/9 7.0 2“.2 0 0.13 Red Prince Delicious Small fruits 5/31 0.8 9.0 56 0.91 6/2 1.1 10.8 “3 2.02 6/5 2.0 13.8 32 1.5“ 6/8 3.1 16.“ 6 1.00 6/11 “.3 18.6 0 0.51 Large fruits 5/31 1.“ 10.8 “ l.“3 6/2 1.7 13.0 18 1.17 6/5 3.2 16.2 0 0.86 6/8 “.6 19.0 0 0.30 6/11 6.3 21.“ 0 0.20 zEstimated from mean weight using data in Figure 2. yEstimated from mean diameter using data in Figure 3. 110 Figure 3. Relationship between fruit diameter at various sampling times and fruit abscission as of June 12 (McIntosh) or 1“ (Delicious) 1981. Per cent absclsslon 111 1001» . . _e g _ f - Delicious . \ 60“» 60+- a - b c d 1130 of sampling .1961 4°i' Ms mu. ‘ a. 5129 ' 5131 b. 6131 612 204- c. 613 616 d. 616 '6/6 0. 619 6111 0 __ill Ar 7' r ‘r ‘r uL I? 7r 100» McIntosh 80d- 60.. 40+- 204- o'-_',’7 .f 7' r 7 r fl - r O 4 6 6 10 12 14 16 16 20 22 Dlamotor on day sampled Figure 3 Figure “. 112 Ethylene production by large vs small 'Mc- Intosh' (A + B) and 'Red Prince Delicious' (C + D) apple fruits as a function of day of sampling (A + C) or fruit weight (B + D). Day 0 was May 27 ('McIntosh') or 30 ('Delicious'), 1981. Ethylene evolved nI pet a per hr 113 u 2.51: 2.09 fl! McIntosh °-—° Smell fruits m Large fruits Q“. v v z . 6 9 Day of sampling It 6 Fruit weight“) Figure 4 11“ the other hand, the rate of ethylene evolution declined as fruit size increased; therefore differences in size, per se, could be responsible for differences in ethylene evolu- tion. When the data were replotted as ethylene evolution _s. fruit diameter (Figure “B and D), the curves for large and small fruits were nearly coincident, suggesting that differences in rates of ethylene production are indeed due to differences in size rather than abscission potential. Discussion Williams (1980) suggested that the "June" drop can be reduced or eliminated by inhibiting endogenous ethylene formation; applying AVG at 1000 ppm to apple fruitlets two weeks after full bloom completely eliminated this drop. Although AVG at 200 ppm appeared to increase fruit reten- tion in “ of 5 comparisons in my experiments (Table 8), the increase was significant in only one of the “. In a first comparison (1980-McIntosh), AVG actually increased drop significantly. In retrospect either higher concentra- tions of AVG or more replications should have been used. AVG reduced ethylene evolution from treated fruits in “ of 5 comparisons (Table 8), all differences being non-signifi- cant; furthermore, reductions in ethylene evolution were not necessarily associated with increases in fruit reten- tion. 115 .poommo pcoumfimcoo o: n o momoopocfi u + momoopooo I IN .emzn an Ho>oH am no Hooncoo goon noonoanno mnonoofioaomnmw o *++ *++ n .0Ho>o ooonznnm o I I + cowucouom H00H «nooonofieon ooonnn oom. + *++ *++ o .0Ho>o oooaznnm o +II I + coapcopom H00H ..noonoHoz. *++ I I .3H0>o ocoachm I + o coaucouom 000a ..osoHoHHon nooaoo. x++ I o .SHo>o ocoaznpm I: *+ + noeoconom 000a ..ooofioneon oonfina oom. a++ I I .SHo>o ocoahcum *II *I I coaucopom 000a ..nooooHoz. AEQQ ooav Anomo Eon oomv AEQQ oomv AEQQ oomv weapoom "waspm new oonoonnm+0>< connonnm 0>< no onooeem uCoEpoopB ocoahnpo .Hmlomma soapSHo>o poo coaucouop uHSLM wagon co mucoEpooLp no nmuoommo go apneezm .m magma 116 Ethephon consistently reduced fruit retention, but the effect was significant only in one case (1980-McIntosh) (Table 8). More pronounced differences were expected in view of the effectiveness of ethephon at 200 or 250 ppm in previous studies with apple (Walsh, et al., 1979; Edgerton and Greenhalgh, 1966; Veinbrants, 1979). For example, Walsh, et al. (1979) increased fruit drop in 'Golden De- licious' and 'Northern Spy‘ by applying 200 ppm two weeks after petal fall. Ethylene evolution from treated tissues was consistently high following application of ethephon in my experiments, thus breakdown of the chemical did not appear to be limiting activity. The combined effects of AVG and ethephon paralleled those of ethephon alone, although ethylene evolution was consistently reduced by AVG treatment (Tables 6 and 8). AVG actually increased the thinning effect of ethephon in 'McIntosh' in 1981. Williams (1980 and unpublished data) observed that “00 ppm AVG overcamelfluapromotive effects of NAA (10 ppm) on both fruit abscission and ethylene evolu- tion, but did not reduce the effects of a very high concen- tration (1000 ppm) of ethephon. These data were inter- preted to mean that AVG inhibited the thinning effect of NAA by interfering with NAA-stimulated ethylene synthesis, but was unable to prevent ethephon-induced abscission because biosynthesis of ethylene was not involved. Neither scoring nor spraying with silver thiosulfate had appreciable or consistent effects on either fruit 117 retention or ethylene evolution. Limbs may have been scored too late; previous workers treated limbs within two weeks of full bloom (Murneek, 1937; Griggs and Schrader, 19“1; Batjer, 1962; Dennis and Edgerton, 1965). Greene (1980) treated apple flowers with AgNO3; ethylene produc- tion was stimulated slightly and fruit set was unaffected. However, silver thiosulfate reduces both ethylene evolution and petal abscission in geranium (Miranda, 1981). Both abscission potential and ethylene production were generally higher in small than in large fruits on a given sampling date (Tables 3, “, 6). However, ethylene produc- tion was more closely associated with fruit size, 933 se, than with abscission potential (Figure “). Ethylene evolu- tion decreased as fruit weight increased, confirming the observations of Blanpied (1972), who noted that ethylene content of 'Golden Delicious' fruitlets decreased as growth commenced. Although these data are not conclusive, they do not support the hypothesis that ethylene production is the controlling factor in apple fruitlet abscission during "June" drop. The effect of ethephon is undoubtedly mediated by the ethylene released within the tissues, but ethylene produCtion in untreated fruits remains far below that ob- served in ethephon-treated ones within 2“ hours of treat- ment. The effect of competition between large and small 118 fruits for nutrients may play a more important role in "June" drop than does ethylene. Abscising fruits generally contain more aborted seeds than adhering ones (Heinicke, 1917; Blanpied, 1972). Hormones probably play a major role in this competition. Young fruits are dependent on their seeds as centers of hormone production for attract- ing nutrients. Large fruits with more developed seeds probably produce more hormone(s) and can therefore attract more metabolites to them than small ones, and the shedding of fruits occurs at a time when the hormone content of the seeds is low (Luckwill, l9“8). 10. 11. Literature Cited Batjer, L. P. 1962. Effect of pruning, nitrogen and scoring on growth and bearing characteristics of young éDelicious' apple trees. Proc. Amer. Soc. Hort. Sci. 2:5-10. Beyer, E. M., Jr. 1976. A potent inhibitor of ethylene action in plants. Plant Physiol. 58:268-271. . 1979. Effect of silver ion, carbon di- oxide and oxygen on ethylene action and metabolism. Plant Physiol. 63:169-173. Blanpied, G. D. 1972. A study of ethylene in apple, red raspberry and cherry. Plant Physiol. “9:627-630. Burg, S. P. and E. A. Burg, 1967. Molecular require- ments for the biological activity of ethylene. Plant Physiol. “2:1““-152. Childers, N. F. 1969. Modern fruit science. Somer- set Press, Inc., New Brunswick, New Jersey. 912 pp. Dennis, F. 0., Jr. 1970. Effects of gibberellins and naphthalene acetic acid on fruit development in seed- legs apple clones. J. Amer. Soc. Hort. Sci. 95:125— 12 . and L. J. Edgerton. 1965. The effects of gibberellins and ringing upon apple fruit develop- ggnt and flower bud formation. Proc. Amer. Soc. Hort.Sci. :l“-2“. Ebert, A. 1980. Hormonale Aspekte der Fruchtbehangsreg- ulierung beim Apfel. Ph.D. Thesis. University of Hohenheim. Edgerton, L. J. and W. J. Greenhalgh. 1969. Regula- tion of growth, flowering and fruit abscission with 2-chloroethanephosphonic acid. J. Amer. Soc. Hort. Sci. 9“:1l-13. Gourley, T. H. and F. S. Howlett. 1957. Modern fruit production. MacMillan Company. New York, 579 pp. 119 12. 13. l“. 15. 16. 17. l8. 19. 20. 21. 22. 120 Greene, D. W. 1980. Effect of silver nitrate, amino- ethoxyvinylglycine and gibberellins Au+ plus 6- benzylaminopurine on fruit set and deve1opment of 'Delicious' apple. J. Amer. Soc. Hort. Sci. 105: 717-720. Griggs, W. H. and A. L. Schrader. l9“1. Effect of branch ringing before and after blossoming on fruit set of Delicious apple. Proc. Amer. Soc. Hort. Sci. 38:89-90. Hale, C. R. and R. J. Weaver. 1962. The effect of development stage on direction of translocation of photosynthate in Vitis vinifera L. Hilgardia 33:89- 131. Halevy, A. H. and A. M. Kofranek. 1977. Silver nitrate treatment of carnation flowers for reduction of ethylene damage and extending longevity. J. Amer. Soc. Hort. Sci. 102:76-77. Heinicke, A. J. 1917. Factors influencing the ab- scission of flowers and partially developed fruits of the apple (Pyrus malus L.) Ph.D. Thesis. Cornell University, 11“ pp. Luckwill, L. C. 19“8. The hormone content of the seed in relation to endosperm development and fruit dr0p in the apple. J. Hort. Sci. 2“:32-““. Miranda, R. M. 1981. Studies of petal abscission in hybrid geranium. Ph.D. Thesis, Michigan State Uni- versity, East Lansing, MI. Murneek, A. E. 1937. Branch ringing and fruit set of Minkler and Arkansas (Black twig) varieties of apple. Proc. Amer. Soc. Hort. Sci. 35:2“-26. Reid, M. S., D. S. Farnham, and J. L. Paul. 1980. Control of cut flower senescence. Proc. 27th Annual. Congr. Amer. Soc. Hort. Sci. (Tropical Region) (in press). Saltveit, M. E., K. J. Bradford and D. R. Dilley. 1978. Silver ion inhibition of ethylene synthesis and action in ripening fruits. J. Amer. Soc. Hort. Sci. 103: “72-“75. Schneider, G. W. 1975. Ethylene evolution and apple fruit thinning. J. Amer. Soc. Hort. Sci. 100:356-358. 23. 2“. 25. 26. 27. 28. 29. 30. 31. 32. 33. 3“. 121 Singh, 1. S. and R. J. Weaver. 1976. Effect of girdl— ing and gibberellic acid on endogenous level of ethylene in Black Corinth grapes. Haryana, J. Hort. Sci. 5:150-153. Teskey, B. J. E., and J. S. Shoemaker. 1972. Tree fruit production. AVI Publishing Company, Inc. 336 pp. Veinbrants, N. 1979. Further studies_on the use of 2- chloroethylphosphonic acid (ethephon) thinning agent for apple. Australian J. of Expt. Agr. and Animal Husbandry 19:611-615. Walker, D. W., D. R. Paterson and D. R. Earhart. 1979. Silver nitrate ion increases endogenous ethylene in sweet potato vine cutting. HortScience 1“:536-539. Walsh, 0. 8., H. J. Swartz and L. J. Edgerton. 1979. Ethylene evolution in apple following post-bloom thin- ing sprays. HortScience l“:70“-706. Weaver, R. J., H. A. Abdel-Gawad, and G. C. Martin. 1972. Translocation and persistence of (2-chloroethyl)- phosphonic acid in Thompson Seedless grape. Physiol. Plant. 26:13-16. Westwood, M. N. 1978. Temperate-zone Pomolo y. W. H. Freeman and Company, San Francisco. “2 pp. Williams, M. W. 1979. Chemical thinning of apple. Hort. Rev. 1:270-300. ' Williams, M. W. 1980a. Retention of fruit firmness and increase in vegetative growth and fruit set of apple with aminoethoxyvinylglycine. HortScience 15:76-77. Williams, M. W. 1980b. Control of ethylene production from naphthalenacetic acid and with aminoethoxyvinyl- glycine and the effect on fruit abscission of apples. HortScience 15:3“5 (Abstract, and unpublished data). Zucconi, F. 1975. Reassessment of the relationship between hormonal and development changes during ab- scission with particular reference to peach (Prunus perSica L.) fruit. Ph.D. Thesis, Michigan State Uni- versity, East Lansing, MI. Zucconi, F., S. P. Monselise and R. Goren. 1978. Growth abscission relationship in developing orange fruit. Sci. Hort. 9:137-l“6. SECTION III EFFECT OF AMINOETHOXYVINYLGLYCINE ON THE EFFECTIVE POLLINATION PERIOD OF APPLE 122 Abstract. Aqueous sprays of aminoethoxyvinylglycine (AVG), an inhibitor of ethylene synthesis,were applied at full bloom to flowers on bagged limbs of 'McIntosh' and 'De- licious' apple (Malus domestica Borkhi.)to determine its effect on the effective pollination period. AVG-treated and control flowers were hand pollinated at 1 to 3 day intervals beginning at anthesis. AVG at 200 ppm increased both initial and final fruit set in 'Delicious' but not in 'McIntosh'. In both cultivars, fruit set decreased as the time of pollination was delayed, and response of AVG— sprayed flowers paralleled that of control flowers. These results suggest that AVG has little or no effect on the effective pollination period. The period during which pollination of the flower re- sults in fertilization is termed the effective pollination period (EPP) (Williams 1965a). It is a function of both ovule longevity and the rate of pollen tube growth. Williams (1965b) reported that EPP, which varied with cul- tivar and season, was the principal factor controlling fruit set. The EPP of various apple cultivars ranged from 2 to 10 days after anthesis (Williams 1965a). Ovule longevity is an important factor affecting fertilization. Hough (19“7) found that in ovules of 'Delicious' the most frequent abnormality was either a 123 12“ tardy initiation of the megaspore mother cell or slow rate of development of megaspore and embryo sac. Normal embryo sacs broke down soon after the flower opened even though it had been pollinated with compatible pollen. Hartman and Howlett (195“) found that when pollination of 'Delicious' flowers was delayed until “8 hours after anthesis, fer- tilization, which seldom occurred in less than 72 hours, was greatly reduced. They attributed this largely to early ovule degeneration. Rootstock may have an effect on embryo sac degeneration. Fruit set of 'Richared Delicious' apple flowers pollinated at full bloom and petal fall was greater in trees propagated on seedling than on M9 rootstocks (Marro, 1976). Temperature during anthesis influences both pollen tube growth and ovule longevity. Lapins and Arndt (197“) considered the optimum temperatures for pollen tube growth in 'Delicious' flowers to be between 7.2 and 12.8°C; pollen grains failed to germinate below “.“°. However, low tempera- tures prolong ovule longevity, increasing the chance of fertilization. Stott (1972) found that ovule longevity ranged from 9 to 12 days after pollination in many apple cultivars and 5 to 6 days were required for compatible pollen tubes to reach the embryo-sac at low temperatures (9.“°). However, tubes required twice as long (12 days) to reach the ovule following self-pollination. Because pollen tubes penetrate more slowly following self-pollina- tion than following cross-pollination, cultivars with a 125 short EPP are less likely to be self-fruitful than those with a long EPP. EPP was only 3 to “ days for those cul- tivars with limited ovule longevity (Stott,l972). Lapins and Arndt (197“) reported that pollen tubes could reach the base of the style in 3 days at 12.8° vs. 5 days at 7.8°. In strawberry and lowbush blueberry, pollinated flowers produce more ethylene than non-pollinated flowers (Hall and Forsyth, 1967). In orchids pollination enhances ethylene production and flower fading (Burg and Dijkman, 1967). In carnations pollination of intact flowers pro- motes endogenous ethylene production and accelerates petal wilting within 2—3 days from pollination (Nichols,l977). The role of ethylene in regulating EPP and ovule longevity has not been investigated. However, inhibitors of ethylene synthesis such as aminoethoxyvinylglycine (AVG) both in- crease fruit set and inhibit ethylene evolution in apple (Dennis et al., 1978; Greene, 1980; Williams, 1980). The purpose of this study was to determine the role of AVG on EPP in 'Delicious' and 'McIntosh' apple flowers. Materials and Methods Experimental Procedure Eight uniform branches per tree were selected on three 'Starking Delicious' and three 'McIntosh' apple trees about “0 years old in a commercialorchard at Leslie, MI 126 in 1981. These branches were enclosed in cheesecloth bags before full bloom (pink stage) to prevent pollination by insects. At full bloom the "king" flower and all frost— damaged flowers were removed and about 100 viable flowers were counted on each branch. Four branches on each tree were sprayed with AVG (200 ppm) at full bloom (May 6 for 'McIntosh'; May 8 for 'Delicious') and four branches were left untreated. The following pollination treatments were randomly assigned to four AVG-treated and four control limbs on each tree, using a randomized block design with trees as blocks: hand pollinated 0, 2, 5 or 6 days (Mc- Intosh) or 0, 3, “ and 5 days ('Delicious') after AVG ap- plication, using pollen previously collected from 'Empire' apple flowers. All limbs remained bagged, except at the time of pollination, until 10 days after each treatment. The flowers were counted at the time of pollination and initial and final set were recorded. Ten fruits were harvested at maturity from each treated and non-treated limb and the length, diameter and seed number were de- termined for each fruit. Results In 'McIntosh', both initial and final set declined as pollination was delayed (Table 1; Figure l), the effect becoming significant after 5 (initial) or 6 days (final 127 Table 1. Effect of AVG on effective pollination period and fruit characteristics at harvest of 'McIntosh', Leslie, MI 1981. Flowers on bagged limbs treated with AVG (200 ppm) at full bloom (May 6). Time of hand pollination (day after full bloom) AVG (ppm) 0 2 5 6 Mean Initial setZ (z), 5/27 0 71 62 “0 23 “9 1y 200 _i l§ fl 23: 57 1 Mean 75 ay 70 a “2 b . 26 b Final Setz (%), 6/16 0 35 20 17 9 20 l 200 gl_ 3; g; lg 23 1 Mean 28 a 27 a 19 ab 12 a Seeds_per fruit, 9/1“ 0 “.7 5.2 “.1 “.0 “.5 1 200 112 5;; 3;§ 3;6 “.0 1 Mean “.3 a 5.0 a 3.9 a 3.8 a Fruit weight ($13 9/1“ 0 126.3 116.0 098.0 101.7 110.5 200 90.3 95.7 98.7 86.3 92.6 m Mean 108.3 a 105.8 a 98.3 a 9“.0 a L/D ratio, 9/1“ 0 .80 .7 .79 .7 .79 l 200 .80 L82 1&1 L80 481_m Mean .80 a .80 a .80 a .79 a zFruits per 100 flowers. yMean separation within rows, observations, and sets by DMRT, 5% level. Figure l. 128 Effect of AVG on effective pollination period (E.P.P.) of 'McIntosh' and 'Starking Delicious' apple 1981. AVG (200 ppm) applied May 6 ('McIntosh') or 8 ('Starking Delicious') to bagged limbs. Flowers hand-pollinated with 'Empire' pollen at indicated times. Broken lines indicate initial set of AVG-treated (a) and control (b) flowers, solid lines final set of AVG-treated (c) and control (d) flowers. H ogsmfim AEOOJm .33“. mm..:...< m>