”fl WW \ Wm “M who \\\ \ m M \\ MSU LIBRARIES 1—”- your record. be charged if book is stamped below. ____:::#_.__T_._ g27> p .. - 'h‘L a -.¢$§ ' SE9 533:2!" ‘- '2 ,. _ ‘7}?R'ag ' ‘ C R“ z/ BEIURNING MATERIALS: P1ace in book drop to remove this checkout from imam] returned after the date L. ' , Wm " 9 can? INFLUENCE OF SILVER THIOSULPHATE AND FUNGICIDES ON PLANT MORTALITY CAUSED BY PYTHIUM ULTIMUM IN THE SEED PROPAGATED GERANIUM (PELARGONIUM x HORTORUM) By Mary Kay Hausbeck A THESIS Submitted to Michigan State University in partial fulfillment of the requirements for the degree of MASTER OF SCIENCE Department of Horticulture 1985 To Jane, my sister and childhood companion. ii ACKNOWLEDGEMENTS Special appreciation and thanks are extended to my major professor, Dr. Royal Heins as well as the members of my graduate guidance committee including Dr. Christine Stephens and Dr. Art Cameron. Their direction and critical evaluation were an integral part of this research. I also extend special gratitude to Dr. Jack Kelly for his unwavering support and interest throughout my studies at Michigan State. The continuous faith of my parents deserves special recognition. Also, the emotional contribution my husband, Greg, has made for my professional development is lovingly 'acknowledged. iii TABLE OF CONTENTS Page LIST OF TABLES .............. ................ ...... vi LIST OF FIGURES ..................................... xi LITERATURE REVIEW . ................ .................. 1 List of References . ..... ......................... 18 SECTION 1: Verification of induced Pythium ultimum mortality in 'Ringo Scarlet' geraniums treated with silver thiosulphate ................................. 28 Abstract .. ............. ....... ........... . ....... 29 Introduction ............ ....... .................. 30 Material and Methods .... ........................ . 50 Results ........... . ..... ......................... 33 Discussion ....................................... 48 Literature Cited .... ............ ......... ........ 55 SECTION II: Efficacy of selected fungicides in controlling crown and root rot in 'Ringo Scarlet' geraniums caused by Pythium ultimum in the presence or absence of silver thiosulphate ................... 58 Abstract ......... ............ . ...... .. ..... ...... 59 Introduction ..................................... 60 Material and Methods ...... . ........ ...... ........ 62 Results .......................................... 66 Discussion ............. . ........................ . 80 Literature Cited ... ....... . ................ . ..... 86 iv TABLE OF CONTENTS (Continued) Page SECTION III: Variation in sensitivity to Pythium ultimum of selected seed propagated hybrid geranium oultfvars .................................. 89 Abstract ......................................... 90 Introduction .... ............... ..... ....... ...... 91 Material and Methods ............................. 92 Results ......... ..................... . ........... 97 Discussion ....................................... 106 Literature Cited ........ .......... .............. . 112 APPENDIX A APPENDIX B LIST OF TABLES Table Page SECTION I Percent mortality of 'Ringo Scarlet' geraniums 50 days STS application and at experiment termination when transplanted into 4 levels of Pythium ultimum-infested medium at 55 days an treated with .25 mM silver thiosulphate (STS) at 4 timings ........................................... 59 Percent mortality of 'Ringo Scarlet' geraniums 50 days STS application and at experiment termination when transplanted into 4 levels of Pythium ultimum—infested medium at 49 days and treated with .25 mM silver thiosulphate (STS) at A timings ..................................... ..... . 45 SECTION II Experiment 1 Percent mortality of 'Ringo Scarlet' geraniums 110 days after seeding and 80 days following transplant into Pythium-infested medium. Plants were sprayed with .25 mM silver thiosulphate (STS) on day 80 and treated with Pythium controlling fungicides at selected timings .................................. 67 Average size of 'Ringo Scarlet' geraniums 51 days after seeding and 21 days following transplant into non-infested or Pythium-infested medium when treated with Pythium controIIing fungicides at selected timings .................................. 69 Average size of 'Ringo Scarlet' geraniums 98 days after seeding and 68 days following tranplant into non-infested or Pythium-infested medium when treated with Pythium controIIIng fungicides at selected timings ........ .......... . ............... 70 vi LIST OF TABLES (Continued) Table 4. Average days to flower of 'Ringo Scarlet' geraniums when transplanted into Pythium-infested medium on day 50 when treated wItE Pythium controlling fungicides at selected timings ..................... 5. Average days to flower of 'Ringo Scarlet' geraniums when treated with Pythium controlling fungicides at selected timings ................................. Experiment 2 6. Percent mortality of 'Ringo Scarlet' geraniums 155 days after seeding and 114 days following transplant into Pythium-infested medium. Plants were sprayed with .25 mM silver thiosulphate (STS) on day 89 and treated with Pythium controlling fungicides at selected timings ................... ...... .......... 7. Average size of 'Ringo Scarlet' geraniums 155 days after seeding and 114 days following transplant into non-infested or Pythiumuinfested medium when treated with Pythium controIIing fungicides at selected timings ............................................ Experiment 5 8. Percent mortality of 'Ringo Scarlet' geraniums 112 days after seeding and 75 days following transplant into Pythium-infested medium. Plants were sprayed with .25 mM silver thiosulphate (STS) on day 89 and treated with Pythium controlling fungicides at selected timings ................................... 9. Average size of 'Ringo Scarlet' geraniums 84 days after seeding and 47 days following transplant into non-infested or Pythium-infested medium when treated with Pytfiium controlling fungicides at selected timIngs ................................ 10. Average size of 'Ringo Scarlet' geraniums 105 days after seeding and 68 days following transplant into non-infested or Pythium-infested medium when treated with Pythium controIIing fungicides at selected timings ....................... ....... ..... 11. Average days to flower of 'Ringo Scarlet' geraniums when treated with'Pythium controlling fungicides at selected timings ...... ...... . ..... . ................ Page 72 75 74 76 77 79 81 LIST OF TABLES (Continued) Table Page SECTION III 1. Selected seed propagated hybrid geranium cultivars, seed sources, flower colors and foliage characteristics grouped into 4 experiments. Each cultivar was screened for sensitivity to crown and root rot disease when grown in P. ultimum- infested medium and treated witH or witEout silver thiosulphate ................... ..... ............... 95 Experiment 1 2. Percent mortality of selected geranium cultivars grown in non-infested (-P) or Pythium- infested (+P) medium and treated with 0.25 mM silver thiosulphate (STS) .................. ........ 98 5. Average size and days to flower of selected geranium cultivars grown in non-infested (—P) or Pythium-infested (+P) medium . ..... ....... .......... 99 Experiment 2 4. Percent mortality of selected geranium cultivars grown in non-infested (-P) or P thium- infested (+P) medium and treated with 0.25 mM silver thiosulphate (STS) .......................... 100 5. Average size and days to flower of selected geranium cultivars grown in non-infested (-P) or pythium-infested (+P) medium OOOOOOOOOOOOOOOOOOOOOOO 101 Experiment 5 6. Percent mortality of selected geranium cultivars grown in non-infested (—P) or Pythium- infested (+P) medium and treated with 0.25 mM silver thiosulphate (STS) ......................... 105 7. Average size and days to flower of selected geranium cultivars grown in non-infested (-P) or Pythium-infested (+P) medium ....................... 104 viii LIST OF TABLES (Continued) Table Page 10. A1. A2. A3. A4. A50 A6. A7. Experiment 4 Percent mortality of selected geranium cultivars grown in non-infested (-P) or Pythium- infested (+P) medium and treated with 0.25 mN silver thiosulphate (STS) .......................... 105 Average size of selected geranium cultivars grown in non-infested (-P) or Pythium-infested (+P) medium .... ...... ................................... 107 Average size and days to flower of selected geranium cultivars grown in non-infested (-P) or Pythium-infested (+P) medium .............. ......... 108 APPENDIX A Average size of 'Ringo Scarlet' geraniums 51 days after seeding when treated with Pythium controlling fungicides at selected timings. Average size of 'Ringo Scarlet' geraniums 51 days after seeding and 21 days following transplant into Pythium- infested medium when treated with Pythium controIIing fungicides at selected timings. Average size of 'Ringo Scarlet' geraniums 98 days after seeding when treated with Pythium controlling fungicides at selected timings. Average size of 'Ringo Scarlet' geraniums 98 days after seeding and 68 days following transplant into Pythium- infested medium when treated with Pythium contro ing fungicides at selected timings. Average size of 'Ringo Scarlet' geraniums 155 days after seeding when treated with Pythium controlling fungicides at selected timIngs. Average size of 'Ringo Scarlet' geraniums 155 days after seeding and 114 days following transplant into Pythium- infested medium when treated with Pythium controIIing fungicides at selected timings. Average size of 'Ringo Scarlet' geraniums 84 days after seeding when treated with Pythium controlling fungicides at selected timings. ix LIST OF TABLES (Continued) Table A8. A9. A10. B1. B2. B40 B5. Page Average size of 'Ringo Scarlet' geraniums 84 days after seeding and 47 days following transplant into Pythium- infested medium when treated with Pythium controIIing fungicides at selected timings. Average size of 'Ringo Scarlet‘ geraniums 105 days after seeding when treated with Pythium controlling fungicides at selected timings. Average size of 'Ringo Scarlet' geraniums 105 days after seeding and 68 days following transplant into Pythium- infested medium when treated with Pythium controIIIng fungicides at selected timings. APPENDIX B Average size of selected geranium cultivars grown in non-infested (-P) or Pythium-infested (+P) medium. Average size of selected geranium cultivars grown in non-infested (-P) or Pythium-infested (+P) medium. Average size of selected geranium cultivars grown in non-infested (—P) or Pythium-infested (+P) medium. Average size of selected geranium cultivars grown in non-infested (-P) or Pythium-infested (+P) medium. Average size of selected geranium cultivars grown in non-infested (-P) or Pythium-infested (+P) medium. LIST OF FIGURES Figure Page SECTION I Symptoms of Pythium ultimum infection on geranium showing chlorotic, reddish and wilted lower leaves ................................. ....... 55 . Size comparision between geraniums grown in non-infested (LEFT) and Pythium ultimum- infested medium(RIGHT) ..................... ......... 56 First symptoms of crown rot phase due to Pythium ultimum showing black, watersoaked stem Iesions at the base of the plant just above the soil line ................................. 57 Percent mortality over time of 'Ringo Scarlet' geraniums transplanted into control, low, medium or high levels (0.0, 0.75, 1.50 or 5.0 g inoculum liter4medium) of Pythium ultimum-infested medium 55 days after seeding. PIants were treated with 0.25 silver thiosulphate (STS) at O, 7 or 14 days following infestation or not at all. The arrow G—+) indicates the first STS application. A second application (----) was made thirty days after the initial STS treatment ............................... 41 Average plant height, width and volume of 'Ringo Scarlet' geraniums over time transplanted into control, low, medium or high levels (0.0, 0.75, 1.50 or 5.0 g inoculum liter”1 medium) of Pythium ultimum- infested medium at 55 days. Letters indiEate significant differences in treatment means based on Tukey's test ..... ............ ... ....... .. ........... 45 xi LIST OF FIGURES (Continued) Figure Page 6. Percent mortality over time of 'Ringo Scarlet' geraniums transplanted into control, low, medium or high 1evels(0.0, 0.75. 1.50 or 3.0 g inoculcum uter-1 medium) of Pythium ultimum-infested medium 49 days after seeding. Plants were treated with 0.25 silver thiosulphate (STS) at 0, 7 or 14 days following infestation or not at all. The arrow G—+) indicates the first STS application. A second application (----) was made thirty days after the initial STS treatment ....... ......................... ....... 47 7. Average plant height, width and volume of 'Ringo Scarlet' geraniums over time transplanted into control, low, medium or high levels (0.0, 0.75, 1.50 or 5.0 g inoculum liter'1 medium) of Pythium ultimum-infested medium at 49 days. Letters Indicate significant differences in treatment means based on Tukey's test ... ............ 50 xii ABSTRACT INFLUENCE OF SILVER THIOSULPHATE AND FUNGICIDES ON PLANT MORTALITY CAUSED BY PYTHIUM ULTIMUM IN THE SEED PROPAGATED GERANIUM (PELARGONIUM X HORTORUM) By Mary Kay Hausbeck Crown and root rot symptoms caused by Pythium ultimum were described for the first time on the seed propagated geranium. 'Ringo Scarlet' geraniums transplanted into low. medium or high levels (0.75, 1.50, or 5.0 g inoculum liter--1 medium) of P. ultimum-infested medium and sprayed with 0.25 mM silver thiosulphate 0, 7 or 14 days following transplant showed greater mortality than geraniums not treated with silver thiosulphate. Geraniums not treated with silver thiosulphate but grown in P. ultimum—infested medium often appeared healthy except for reduced plant size. Fenaminosulf, ethazol or metalaxyl applied as soil drenches at selected times reduced crown and root rot disease caused by P. ultimum in geraniums not treated with silver thiosulphate. Only metalaxyl consistently controlled the increased incidence of crown and root rot in geraniums sprayed with silver thiosulphate. Genetic resistance to P. ultimum was not identified in the 57 cultivars screened. LITERATURE REVIEW The seed propagated hybrid pelargonium (Pelargonium x hortorum) belongs to the Geraniaceae family. Family members are characterized by the long-beaked fruit or 'Cranesbill' from which the Greek word geranos is derived (32,39,68,82). Geraniaciae was first published in Materia Medica, a book of herbal remedies written by Dioscorides about 50 A.D. (32,68). The simple, irregular (zygomorphic) flower shape and necter tube are responsible for the separation of Pelargonium from the other 10 genera of Geraniaceae. The Dutch of the Cape Colony of South Africa discovered the pelargonium in the opening years of the seventeenth century. Its natural habitat is the arid semi-desert of southern Africa. With few exceptions, all members of the genus Pelargonium are of African origin (32,39,68,82). By the 1650's several species were under greenhouse cultivation in Europe (39). Approximately 300 spp. of pelargoniums exist today varying in habit, leaf form and flower color (82). Hybridization has changed the flower form from irregular to rounded, single, semi-double, double, and the quilled types currently available. Pelargonium x hortorum originally arose from a cross between P; inquinans and P; zonale with subsequent 2 characteristics incorporated from other species (39,82). This garden or bedding plant is best known as "geranium", a name more correctly describing its hardy, European native cousin of the Geranium genus. However, further use of the term geranium in this thesis will be in reference to the hybrid geranium Pelargonium x hortorum. Until the 1960's geraniums were almost exclusively propagated by cuttings. Erratic germination, non-uniformity and a flowering time of 12-15 months made seed propagation undesirable (39). Extensive breeding and development in the 1960's made growing geraniums from seed commercially feasible. The open-pollinated 'Nittany Lion Red' developed by Dr. Richard Craig at Pennsylvania State University was the first true breeding hybrid geranium produced by seed (96). Subsequent improvements followed with the introduction of the 'New Era' and 'Carefree' F Hybrids from the Harris Seed Co. and Pan-American Seed Co. (37). Through the joint effort of Goldsmith Seeds, Inc. and Sluis and Groot, a group of cultivars called the Sprinters were released in 1973 and were the first seed propagated geranium to successfully flower in the pack. Subsequently, cultivars referred to as the Ringos were introduced by Sluis and Groot in 1978 which flowered earlier and were more compact for pack production (Personal communication, Dr. L. Ewart; Michigan State University). In 1983, approximately 10 seed companies were developing improved hybrid seed cultivars. More than 100 3 cultivars have been introduced, bringing annual production in 1983 to 90-100 million seeds (37). The development and availability of early flowering cultivars makes the production cost of the seed propagated hybrid geranium competitive with the traditional cutting propagated geranium. Petal shatter has been a problem in transporting and marketing the seed propagated hybrid geranium (10). The degree of shattering varies among cultivars (11). Double or semi-double flowers lacking nectaries do not shatter easily (120). However, the "low shatter" cultivar may not have the most desirable or aesthetic characteristics. Petal abscission in the seed-propagated hybrid geranium is increased by exogenous ethylene (11,120). Premature petal abscission in the geranium can be reduced by preventing ethylene accumulation and transporting under cool temperatures (1-5o C) (11). However, cool storage in combination with frequent irrigation was found to increase ethylene sensitivity and decrease flower life after storage removal in studies on cut carnations (71). In addition to promoting petal abscission in seed propagated geraniums, ethylene also plays in important role in the keeping quality of several floricultural crops (“6). Crocker and Knight (38) first suggested exogenous ethylene as the deleterious component in illuminating gas causing cut carnation "sleepiness". Naturally emanating ethylene from cut flowers measured under contolled conditions was believed 4 to have been produced by fungus which became contaminants during investigations. Nichols (88), however, showed an ethylene surge accompanies wilting in carnation independent of fungal infection. This confirmed Smith's (106) proposal that the ethylene surge after fungal attack was due to the breakdown of the host tissue and was not generated by the fungus. Ethylene production has been correlated with flower longevity in cut carnations (“0). During a low steady-state ethylene production phase, carnation flowers remained turgid. This ethylene level was followed by a high log linear phase which resulted in flower wilting and senescence. Kende and Hanson (58) concluded from studies with morning glory tissue that this ethylene-induced senescence is an acceleration of natural senescence. The silver ion is effective in blocking ethylene actions such as senescence and petal abscission (16,17). Silver ion appears to inhibit ethylene effects at a very early stage in the events leading to abscission (94). Precisely how silver inhibits ethylene action is not known. Yang (123) recently presented many of the current hypotheses. Beyer (18,19) proposed that a small fraction of ethylene is metabolized and incorporated into tissue or converted to C02. Burg and Burg (2“) first suggested that that ethylene binds to a metal-containing site. Sisler and + Goren (102) support this theory. Copper (Cu ) is the 5 suggested metal because of its affinity to ethylene (18,26). Sisler (109) showed that ethylene binding is prevented by reagents containing copper enzymes. Ethylene can form mono and diligand complexes with silver (57) however, the bond is not irreversible (16). It is the conclusion of many investigators that ethylene biosynthesis is not inhibited by silver (6,16,18). Silver appears to counter the effect of ethylene by blocking ethylene action at its receptor sites (6,16,18). Studies in which silver lowered ethylene binding by a plant extract support this theory (102). Sisler and Goren (103) Propose that this ethylene/receptor complex triggers a series of metabolic events and then diffuses out or is degraded. The binding of ethylene to its receptor increased the pool size of the ethylene precursor aminocyclopropane—1- carboxylic (ACC) as well as ethylene production in studies with cut carnation flowers (25). Exogenous ACC caused wilting and increased ethylene levels. Veen and Kwakkenbos (117) showed increased ethylene levels but no flower wilting in cut carnations when a STS treatment was followed by ACC. The blocking of ethylene to the receptor site may inhibit the autocatalytic ethylene increase and the accompanying ACC content increase. Similar studies substantiate this conclusion. Veen (114) blocked the ethylene surge preceding the wilting of cut carnation by the anionic silver thiosulphate (STS) solution. In preclimacteric fruit STS inhibited ethylene-induced ethylene production (55). 6 The receptacle tissue may be an important factor in the silver inhibition of ethylene action as silver accumulates in receptacle tissue (20,35,116,119). Veen (114) showed that this tissue is also capable of producing ethylene in large quantities. Although the primary response of silver is considered to be ethylene action inhibition, other physiological plant processes are also affected. Abscisic acid (ABA) influences senescence by affecting ethylene production (98). With ethylene treatment, ABA levels rose during senescence but Nowak and Veen (90) blocked this increase with STS. STS also prevented endogenous cytokinin levels from rising in cut flower pistils (113). It is postulated that high cytokinin levels in reproductive organs create a sink enhancing flower senescence. STS pretreatment prevents gynaceium enlargement due to carbohydrate accumulation and allows continued transport and utilization of carbohydrates in the petals (41,118). Silver is also recognized for its germicidal effect. In postharvest physiology studies, Kofranek and Paul (61) immersed the basal end of cut carnations in 1200 ppm AgNO for varying time periods. The vascular and pith tissues 3 absorbed silver which moved upward with time. The surface and cut end tissue were impregnated with silver which functioned to inhibit bacterial plugging. Nichols (89) concluded that the silver nitrate prevented bacterial stem blockage, thereby allowing absorption of sucrose-containing 7 preservative. Kofranek (60) pulsed cut chrysanthemums with basal treatments of silver nitrate plus 5% sucrose and increased flower longevity under a range of shipping temperatures. Silver alone had a much smaller effect than the combination of silver and sucrose in cut carnation longevity. Dilley and Carpenter (A0) attributed the delayed senescence benefits of sucrose to the delay but not inhibition of the autocatalytic production of ethylene. They postulated that sucrose could stimulate respiration creating higher CO levels which could delay autostimulation of ethylene production by endogenous ethylene. Halevy and Kofranek (“5) compared basal and foliar silver treatments on cut carnations to determine if longevity is due to bacteria action or by countering ethylene. They found the benefits of a stem treatment were due to bactericidal properties whereas senescence delay of directly treated flowers was due to anti-ethylene properties of silver. Because of its ability to block ethylene action, and thereby delay senescence, silver has become an important tool in commercial floriculture. The anionic STS complex is most commonly used because this negatively charged complex is not as likely to be involved in absorption and exchange processes and moves freely within the plant (35). The silver ion formulated as silver nitrate (AgNO ) has low 3 mobility in the plant (61,62). STS reduces phytotoxicity 8 from silver oxidation which often occurs with silver nitrate application (44,45), yet the inhibitory effect of silver on ethylene action is preserved. STS is effective in preventing premature senescence and petal abscission in a variety of floricultural crops (7,29,42,43). STS inhibited senescence even in detached petals (86) STS does not, however, produce uniform results in all horticultural crops (115). Cut carnations differed in response to STS with 'standard"flowers more responsive than spray carnations (110). STS has also been shown to remain effective over an extended time period. Swart (109) immersed lily bulbs in STS prior to planting. The flowers eventually produced from these bulbs were of higher quality and not as sensitive to exogenous ethylene than those not treated with STS at pre- plant. A variety of potted plants treated with STS 2-3 weeks prior to harvest with STS were protected from premature flower abscission under simulated harvest, transit and retail conditions (28). STS is effective in controlling petal shatter in the seed propagated hybrid geranium (27,79). In geranium production, a STS foliar application is commonly applied just prior to transit. A molar ratio of 1:8 (silver nitrate (AgNO ) to sodium thiosulphate (Na S 0 .SH 0)) has been guggested by Veen and Van de ée$n3(11g). Reid et a1. (95) recommended a 1:4 ratio. STS mixing procedures have been simplified by Heins et a1. (49). 9 However, during STS formulation trials a higher incidence of crown and root rot symptoms on STS treated geraniums was observed (49). Pythium ultimum was identified as the causal agent. Commercial growers reported similar findings (Personal communication, Dr. Royal Heins Michigan State University, 1983). Pythium is a large genus with members found worldwide including saprophytes in water or soil and parasites on algae, other fungi, or higher plants (75). The damping-off and crown rot soil pathogens belong to the Oomycete class. They are characterized by coenocytic, elongated mycelium, zoospore production in zoosporangia and resting spores called oospores produced by the union of two morphologically different gametes. Pythium spp. cause disease problems in greenhouse grown crops (92,100). Pythium spp. are ubiquitious pathogens. Stephens et al. (108) showed that Pythium spp. may be recovered from dust and soil mix particle samples collected from walkways, floors and greenhouse beds and flats. Pythium spp. also infect a wide variety of ornamental vegetable and fruit crops in production, storage or transit (30,36,50,52,53,59,91). Diseases caused by Pythium spp. include seedling damping off, rotting of unrooted cuttings, and root rot of established plants. Seedling damping off caused by Pythium Spp. can develop quickly killing large numbers of seedlings in local areas (66,67,97). Prior to damping off, Campbell and Sleeth (30) 10 noted purplish or reddish cotyledon color in Pythium infected guayule cotyledons. Pythium disease symptoms for rooted and unrooted cutting propagated geraniums are well documented and are generally referred to as "blackleg" (87). On cutting propagated geraniums, brown watersoaked lesions originate at the cut base or wounds on young plants. The rooted area enlarges and turns black progressing up 3-4" from the base (22,23,87). On the seed propagated geranium, Pythium disease symptoms have not been thoroughly described although the disease is commonly referred to as blackleg (92). Plant stunting is a typical symptom of root rot due to Pythium infection (30,59,74). Stunted, yellow Pythium infected pineapple showed reduced fruit yields (59). Pythium disease on woody ornamentals resulted in greatly reduced feeder roots and necrotic lesions on existing roots. Root rot symptoms include leaf chlorosis, partial defoliation, reduced growth and vigor and frequent iron deficiency problems (50,54). The amount of stunting depends on on the degree of Pythium injury to the root system (30). Pythium ultimum enters a plant through the root system (51). On young peach roots, 3. ultimum penetrated within 5-8 hours of contact, primarily at epidermal cell junctions (76). Young sweet potato rootlets showed decay within 36 hours after Pythium infection occurred (91). Established infections continued to develop even under drying conditions. Although considered to be primarily a root 11 pathogen, Pythium spp. can infect stems and foliage of some plants provided environmental conditions are favorable for the pathogen. Braun (24) showed increased disease of stems due to P. complectans on succulent geranium cuttings that were crowded, well-fertilized and well-watered. Environmental factors also play a role in root disease development and infection potential (1,36,83). Powell et al. (93) noticed increased severity of peach decline following excessive rainfall. Pythium ultimum was one of several Pythium Species isolated from affected peach root systems. Mircetich (80) correlated high soil moisture with increased P. ultimum saprophytic activity. Loblolly pine decline was associated with trees on poorly drained, fine- textured soils (69). Saprophytic colonization by Phytophthora and Pythium spp. appeared to weaken the trees allowing brooding and subsequent destruction by bark beetles. Increasing the moisture holding capacity (MHC) to approximately 30 to 40% did not greatly increase disease severity, although at 70% MHC P. ultimum became a serious problem. However, Hanan et al. (48) determined deficient aeration was not a prerequisite for Pythium infection based on symptom severity on snapdragons in well-aerated medium. To control disease they reduced irrigation but found this method ineffective and to be further detrimental to plant growth. 12 Sleeth (105) attributed Winter Haven citrus decline to recurring periods of high and low soil moisture and large P. ultimum populations. Biesbrock and Hendrix (21) correlated P. vexans severity with water excess allowing zoospore production. Since P. irregulare forms germ tubes from sporangia it was unaffected by soil water. Specific temperature ranges are an important requirement for many Pythium spp. Pythium irregulare, P. spinosum Sawada, P. ultimum and related species are most damaging at lower temperatures while P. myriotylum, P. aphanidermatum, P. arrhenomanes, P. polytylum Drechs., P. carolinianum Matthews and P. volutum Vanterpool & Truscolt are damaging at higher temperatures (51). Although P. ultimum is reported to produce more damping-off at lower temperatures (18oC-21OC) with little disease at 30°C (12,67), Halpin el al. showed P. ultimum to be more pathogenic on red clover seedlings at 24-2800 than 16-200C. (47). Fertilization also influences disease development. Kraft and Erwin (64) showed favorable nitrogen sources necessary for P. aphanidermatum infection of mung bean seedlings at low inoculum densities. Moore et al. (84) showed calcium deficiency to have the most pronounced influence on susceptilbilty of Highland bentgrass to P. ultimum. Nitrate nitrogen and potassium reduced damping off in moist soil, whereas ammonium nitrogen and phosphate did not (122). Similar results were not obtained in drier soil. 13 It was proposed that fertilizer influences microbial antagonistic activity in moist soil and corrects plant mineral deficiencies. However, Agnihotri and Vaartaja (3) concluded from their investigations that added nutrients may disrupt a delicate biological balance in the soil which under natural conditions prevents germination of P. ultimum sporangium. A suggested mechanism for increased disease incidence following herbicide application is the inhibition of microflora competing with potential pathogens (9). Plant exudations into the surrounding environment may play a role in stimulating or preventing root rot fungal growth. Seed or root exudates have been shown to stimulate oospore germination of P. afertile (3). P. aphanidermatum (31,64), P. mamillatum (14), and P. irregulare (4). High numbers of P. ultimum sporangia germinated when parts of different plants were incorporated into the soil (5). Zoospores of P. aphanidermatum supplemented with mung bean exudates showed greater virulence than zoospores not supplemented with exudates (65). Royle and Hickman (99) showed specific substances present in root exudates served as a nutrient source for P. aphanidermatum. They identified sugar or amino acids as the most common promoting factors. A drench of 1000 ppm glucose, fructose, maltose or sucrose induced high germination of P. ultimum Sporangia in the soil (5). Mixtures of sugars and amino acids also stimulated germination. Specific herbicides have been shown to affect plant exudation and stimulate pathogen growth (9). 14 Herbicide application has been correlated with increased disease incidence. Compounds produced by host plants have also been identified which are toxic to fungi and bacteria. An extensive review has been written on this topic (63). Phytoalexins have been shown to be promoted by ethylene production stimulated by host tissue breakdown due to pathogen infection. However, compound identification and host pathogen relationships are not conclusive. Physiological plant changes may be involved in resistance to Pythium disease. McCarter and Littrel (73) showed oats, wheat and cucumber were more susceptible in the early stages of germination than in late growth stages. Root rot severity in cotton seedlings decreased as the plants grew older (97). Peach tree seedlings escaping pre- emergence and early postemergence mortality in infested soil grew to a size comparable to those in non-infested soil (81). Differences in P. ultimum blackleg resistance in cutting geraniums were attributed to the time required for the cut surface to heal (33). A decrease in disease caused by P. ultimum in cutting geraniums was correlated to the suberization of wound periderm (34). Susceptibility of two safflower cultivars (Carthamus tinctorius) differed, according to the period in which elongation of the hypocotyl first internode ocurred (112). This resistance was not correlated to age alone but rather 15 the physiological processes which occur at the time of internode development. Genetic resistance to P. ultimum has been demonstrated for a variety of crops (51) including bean cultivars (Phaseolus vulgaris) (2). Schroth and Cook (101) found resistance to Pythium disease in three bean varieties to be correlated with the amount of seed exudation; the greatest exudation occurred in the more susceptible variety. Alicbusan et al. (8) showed that P. ultimum grew more abundantly in the rhizophere of susceptible plants than in that of resistant plants. Mathre and Otta (70) were unable to find genetic resistance to P. solani or P. ultimum among species and varieties of cotton. McCarter and Littrell (72) screened crops for genetic resistance to Pythium spp. Tbmato, bean and rye were more susceptible to disease caused by Pythium spp. than cotton and corn. Resistance to P. ultimum has not been thoroughly investigated for seed propagated geranium cultivars. Stephens and Powell (Personal communication, Dr. Christine T. Stephens; Michigan State University) screened many bedding plant crops including two hybrid geranium cultivars for resistance to P. ultimum. Although neither cultivar showed disease resistance, there was a difference in damping—off susceptibilty between them. Powell (92) observed apparent cultivars differ to P. ultimum in naturally occurring disease instances. 16 In the absence of identified Pythium-resistant geranium cultivars, primary control of the pathogen is achieved through sanitation and/or fungicides. Powell (92) and Stephens (107) recommended the fungicides ethazol (Truban), fenaminosulf (Lesan): metalaxyl (Subdue) and ethazol plus dimethyl 4,4-0-0 phenylenbis (Banrot) for Pythium control on seedling geraniums. Moorman (85) recommended heat or fumigant soil treatments as the preferred control method due to the adverse effects of some fungicides on various plant species. In many reported cases, fenaminosulf controlled disease due to Pythium spp. resulting in reduced mortality and increased plant growth (56,97,111). However, Miller and Sauve (78) found fenaminosulf to be less effective than other fungicides for control of Pythium stem rot of cutting geranium. This substantiates earlier work by Wheeler et al. (121). Miller and DeNeve (77) found ethazol superior to fenaminosulf in controlling Pythium crown and root rot on bedding plants. Baker and Harman (13) showed ethazol drenches more effective than soil steaming in controlling Pythium spp. on chrysanthemums. Metalaxyl produced varying results. They also determined that a combination soil steaming plus a Pythium controlling fungicide was more effective in increasing chrysanthemum growth and flower production than either treatment used alone. This effect 17 was attributed to disease eradication through steaming and subsequent fungicide protection from recontamination. SUMMARY The seed propagated hybrid geranium (Pelargonium x hortorum) has become a popular bedding plant in recent years. Breeding programs have produced bright colors, consistent germination and compact growth habits (57). New, early flowering cultivars make the production cost of the seed propagated geranium competitive with the traditional cutting propagated geranium. Petal shatter is a problem in transporting and marketing seed propagated hybrid geraniums (10). STS prevents petal abscission in seed propagated geraniums and is commercially important in seed propagated geranium production (27.43.79). Michigan State University researchers observed P. ultimum crown and root rot symptoms on geraniums treated with STS during STS formulation trials (49). Pythium spp. are often pathogens of greenhouse crops (92,100). Objectives of this study were threefold: 1) To verify the interaction of silver thiosulphate and P. ultimum crown and root rot; 2) To test fungicides and application times for control of mortality due to P. ultimum of geraniums grown in P. ultimum-infested medium and treated with or without STS: 3) To screen geranium cultivars for P. ultimum and P. ultimum/STS resistance. LIST OF REFERENCES 1. Adegebola, M.O.K. and D.J. Hagedorn. 1969. Symptomatology and epidemiology of Pythium bean blight. Phytopathology 59:1113-1118. 2. Adegbola, M.O.K. and D.J. Hagedorn. 1970. Host resistance and pathogen virulence in Pythium blight of bean. Phytopathology 60:1477-1479. 3. Agnihotri, V.P. and 0. Vaartaja. 1968. 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Plant Disease Reporter 51:864-866. 71. Mayak, S. and A. M. Kofranek. 1976. Altering the sensitivity of carnation flowers (Dianthus caryophyllus Z. ) to ethylene. J. Amer. Soc. Hort. Sci. 161: - . 72. McCarter, S.M. and R.H. Littrell. 1970. Comparative pathogenicity of Pythium aphanidermatum and Pythium myriotylum to twere plant species and intraspecific variation in virulence. Phytopathology 60:264-268. 73. McCarter, S.M. and R.H. Littrell. 1968. Pathogenicity of Pythium myriotylum to several grass and vegetable crops. Plant DiSease Reporter 52:179-183. 74. McCarter, S.M. and R.W. Roncadori. 1971. Influence of low temperature during cotton seed germination on growth and disease susceptibility. Phytopathology 61:1426-1429. 75. Middleton, J.T. 1943. The taxonomy, host range and geographic distribution of the genus Pythium. Mem. Torrey Bot. Club 20:1-171. 76. Miller, C.R., W.M. Dowler, D.H. Peterson and R.P. Ashworth. 1966. 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Pennsylvania Flower Growers. 410 pp. 83. Moore, L.D. and H.B. Couch. 1961. Pythium ultimum and Helminthosporium vagans as foliar pathogens of Gramineae. Plant Disease Reporter 45:616-619. 84. Moore, L.D., H.B. Couch and J.R. Bloom. 1963. Influence of environment of diseases of turfgrasses III. Effects of nutrition, pH, soil temperature, air temperature and soil moisture on Pythium blight of highland bentgrass. Phytopathology 53:53-57. 85. Moorman, G.W. 1983. Soilborne diseases. Pennsylvania Flower Growers Bulletin 7:6. 86. Mor Y., M.S. Reid and A.M. Kofranek. 1980. Role of the ovary in carnation senescence. Scientia Hortic. 13:377-383. 87. Nichols, L.P. and P.E. Nelson. 1982. Pythium blackleg. Pages 208-210 in: Geraniums III. J.W. Mastalerz and E.J. Holcomb, eds. Pennsylvania Flower Growers. 410 pp. 88. Nichols, R. 1966. Ethylene production during senescence of flowers. J. Hort. Sci. 41:279-290. 89. Nichols, R. 1973. 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Phytopathology 52:1285-1287. 123. Yang, S.P. 1985. Biosynthesis and action of ethylene. HortScience 20:41-45. Section I: Verification of induced Pythium ultimum mortality in 'Ringo Scarlet' geraniums treated with silver thiosulphate 29 Verification of Induced Pythium ultimum Mortality in 'Ringo Scarlet' Geraniums Treated with Silver Thiosulphate Mary K. Hausbeck, Christine T. Stephens* and Royal D. Heins Graduate Research Assistant and Associate Professors, respectively, Departments of Horticulture and Botany and Plant Pathology*, Michigan State University, East Lansing, MI 48824-1312 ABSTRACT Hausbeck, M.K., C.T. Stephens and R.D. Heins. 1985. Verification of induced Pythium ultimum mortality in 'Ringo Scarlet' geraniums when treated with silver thiosulphate. Plant Disease 0:0. Pythium ultimum crown and root rot symptoms on the seed propagated geranium are described. Root rot caused by P. ultimum often resulted in plant stunting, otherwise plants appeared healthy unless compared to an uninfected plant. Severe root rot symptoms frequently resulted in plant mortality. Stem blackening or crown rot commonly occurred on plants which initially showed only plant stunting or a reduced root system. Geraniums transplanted into a P. ultimum-infested medium and sprayed with silver thiosulphate (STS) had a higher mortality incidence than geraniums transplanted into P. ultimum-infested medium and not treated with STS. Increasing inoculum level resulted in increased mortality when seedlings were transplanted into the P. ultimum-infested medium 35 days after seeding but not when seedlings were transplanted 49 days after seeding. 30 Additional key words: Pelargonium hortorum, Pythium ultimum, silver thiosulphate. INTRODUCTION Premature flower petal shatter has been a severe problem in marketing the seed propagated geranium (4). Foliar application of silver thiosulphate (STS) solution prevents petal abscission in seed propagated hybrid geraniums (6,11,23). Heins, et al. (13) observed crown and root rot symptoms on geraniums treated with STS during STS formulation trials. Commercial growers reported similar findings (Personal communication, Dr. Royal Heins; Michigan State University). The symptoms of this root rot were dark, water-soaked stem lesions and death. Isolations from plants with these symptoms confirmed the identity of the pathogen as Pythium ultimum Trow. The objective of this study was to determine if a relationship existed between plant mortality incidence due to P. ultimum crown and root rot and application of STS on geraniums grown in P. ultimum-infested medium. Information is also presented on the relationship between plant mortality and inoculum levels of P. ultimum. MATERIALS AND METHODS 'Ringo Scarlet' geranium seeds (Sluis & Groot B.V., Enkhuizen, Holland) were individually sown in round 2.0 cm in diameter cells, covered with fine vermiculite and placed 0 under intermittant mist at 24 C in a glass greenhouse. 31 After germination (7 days), the seedlings were removed from the mist and grown at 22°C day and 20°C night temperatures under natural light in the greenhouse. They were fertilized at each watering with 200 mg liter- each of nitrogen and potassium. Preliminary investigations using a P. ultimum isolate cultured from rotted geranium roots and a highly virulent P. ultimum isolate (#248) used in previous studies (28) showed both isolates to cause crown and root rot symptoms (Results not shown). The #248 P. ultimum isolate was used throughout the remainder of this study. Pythium ultimum inoculum was prepared using the potato media procedure first developed for culture of Rhizoctonia solani Kuhn by K0 and Hora (17). Fifty grams of finely chopped potato was added to 500 m1 of a commercial soilless mix (Sunshine Media Mix, Blend 1, Fisons-Western Corp., Vancouver, B.C., Canada) containing 2 parts vermiculite, 2 parts peat moss and 1 part perlite. After mixing, the potato media-mixture was autoclaved twice for one hour, 24 hours apart. Cultures of P. ultimum were maintained on 20 ml of water agar (Difco Laboratories, Detroit, MI 48232) and grown in 10 cm petri plates for 2 days at 240C. Six mycelial discs 12 mm in diameter were selected from the perimeter of the growing culture and used to infest 1.5 liters of sterilized potato soil medium. The inoculum was grown in 2.0 liter closed flasks for 2 weeks and shaken daily. The 32 inoculum mixture was air dried for 1-2 days and sieved through a #10 (2mm) screen. The inoculum was then mixed with the soilless medium at three infestation levels; 0.75 3 liter”1 (low), 1.5 g liter.1 (medium) and 3.0 g liter.1 (high). The low inoculum level was chosen as a level sufficient to induce P. ultimum disease symptoms (Personal communication, Dr. H. Hoitink; Ohio State University). After thoroughly mixing, the infested soil was placed into single cells (8 x 8 cm) of 18 pack flats (25 x 53 cm). Eight single cells (replicates) were used per treatment. Seedlings were transplanted 35 and 49 days after seeding into the P. ultimum-infested medium or into a non- infested medium. The P. ultimum-infested and non-infested treatments were randomized in a growth chamber. Temperatures were maintained at 210C day and 18°C night. Irradiance was controlled at 135.1(111018-1m-2 for 12 hours per day using VHO cool white fluorescent lamps. The medium pH varied between 5.5 and 6.5 during the experiment. Plants were fertilized at each watering with 200 mg liter“1 each of nitrogen and potassium. Foliar applications of 750 ppm chlormequat chlorine (American Cyanamide Co., Wayne, N.J. 07470) (CCC) were applied to control height as growth necessitated (8). A freshly prepared 0.25 mM STS solution (silver to thiosulphate ratio of 1:4 ) (13,25) was sprayed on the foliage the day of transplant into P. ultimum-infested 33 medium (0 days), 7 or 14 days following transplant. A second STS application was applied 30 days after each original STS treatment. The control received no STS. Plants were observed for symptoms of Pythium crown and root rot. The number of days following transplant into P. ultimum-infested medium at which death occurred was recorded. Percent mortality was calculated. Plant height and width were recorded at 36,41, and 56 days or 27,36,42 and 49 days following tranplant into P. ultimum infested- medium on seedlings transplanted at 35 and 49 days respectively. Plant volume (size) was calculated from height and width measurements by assuming the plant to be a cylinder. Reisolation of P. ultimum was conducted on geraniums that died during the study. At the termination of the experiment, surviving geraniums in P. ultimum-infested medium and non-infested controls were randomly sampled. One half inch sections of roots, stem and petioles were surface sterilized in 10% sodium hypochlorite for approximately 20 seconds and plated on water agar. The plates were observed for a minimum of seven days for Pythium hyphal growth. The agar plates with hyphal growth were allowed to dry at room temperature for three weeks to promote encysted sporangia development. The fungus was identified as P. ultimum. RESULTS Disease Symptoms Geraniums grown in P. ultimum-infested medium showed two different types of disease symptoms. The root rot phase 34 was characterized by a number of foliar and root rot symptoms. Lower leaves became chlorotic and also showed a reddish cast associated with anthocyanin due to an apparent breakdown of chlorophyll. Affected leaves eventually wilted (Figure 1). Plants with severe foliar symptoms also showed considerable growth reduction. Root systems of affected plants were often minimal with extensive necrosis and rotting of feeder roots. Geraniums either remained in this condition with no further apparent disease progression or rotted at the soil line resulting in death. Geraniums which showed many root rot symptoms generally did not progress into the stem blackening associated with the crown rot phase. Other geraniums showed stunted growth but otherwise appeared healthy (Figure 2). Root systems of these plants were reduced in size but did not show extensive necrosis or rotting. These plants often developed crown rot symptoms. The first symptoms of the crown rot phase were black, watersoaked stem lesions at the base of the plant just above the soil line (Figure 3). Plants remained green and turgid until stem blackening progressed up the stem to the base of the petioles. Total stem blackening usually occurred during a 24 to 48 hour period depending on the size of the plant. Once the disease symptoms had progressed substantially, the plant toppled over and the blackened tissue hardened. White mycelium at the base of the plant was evident approximately five days following plant death if the plant was kept moist. 35 Figure 1. Symptoms of Pythium ultimum infection on geranium sfiowing cEIorotic, reddish and wilted lower leaves. 36 Figure 2. Size comparision between geraniums grown in non-infested (left) and Pythium ultimum- infested medium (right). 37 Figure 5. First symptoms of crown rot phase due to Pythium ultimum showing black, watersoaked stem Ie51ons at the base of the plant just above the soil line. 38 Response of 35 day old plants When grown in P. ultimum-infested medium but not treated with STS, final plant mortality varied from 0 to 38% (Table 1). Mortality was greater after STS application regardless of P. ultimum infestation level, although total plant mortality increased as P. ultimum inoculum level increased. As the time between transplanting (infestation) and STS application increased (0 to 14 days), plant mortality increased during the first thirty days after transplanting into the P. ultimum-infested medium (Table 1, Figure 4). However, by the termination of the experiment, the number of geraniums that died was similiar within a P. ultimum infestation level. When surviving plants were retreated with STS four weeks following the initial spray, there was additional mortality. The greatest plant loss occurred when STS was reapplied to plants initially treated with STS at transplant (Figure 4). This was due at least in part to the fact that more plants in these treatments survived for 30 days compared to plants in other treatments. Plants growing in the P. ultimum-infested medium were smaller than plants grown in the non-infested medium. At days 36 and 41, plant size in the low, medium and high P. ultimum-infested medium did not differ significantly (Figure 5). By 52 days, plants growing in the high level of P. ultimum-infested medium were significantly smaller than 39 .vouanao mm: mam scans um aawvoa woumowcwlezsquaa . .anvma vmummmcfiusaswuas . M.uo coumouafinco: cuca wawuaoamosouu nouns hon: M.uo woumomaunco: coca ucanmcouu wound name on uaoauuoaxu mo couuoauauosx .coaunuaanam mam Howuwaw uuuuu ammo» .aSwvoa mmoaawom mo nouwu non asdaooaw anawuaa «M.maouuu cos as mm mm on on mm o mane SS cos Hm mm mg an ca 0 o «use A cos mm mm _m as aw mm o amsue c an a c o .S o o o Houuaoo Hanan on Hanan on Hangs on Hanan on .11 x a hoe-a acqunoqaas< mam H\w c.« “\w om.~ H\w m~.o NH\w o.o mao>oa cowumumouca asaauas aawnumm .mmcaaau S no Amemv muons-smofizu uo>awm :3 mm. su-s Bouncy» vac mace mm as esfivoa voummwcwnssefiuaa esfisuzm mo mHm>m~ S cucw nouamanmcmuu coca cofiumcfiauou unmaauoaxo um vcm cowumowaaam mew umuum mxmv 0m mes-:mumm .uoaumom owe-m. mo Sufi-ounce unmouom .H manmh Figure 4. 40 Percent mortality over time of 'Ringo Scarlet' geraniums transplanted into control, low, medium or high levels (0.0, 0.75, 1.5 or 5.0 g inoculum liter‘1 medium) of Pythium ultimum- infested medium 55 days after seeding. Plants were treated with 0.25 mM silver thiosulphate (STS) at 0, 7 or 14 days following infestation or not at all. The arrow G—o) indicates the first STS application. A second application (----) was made thirty days after the initial STS treatment. 41 zo—pchwwuz~ ta~zh>‘ Kuhn: urco zo—bcpmu z— tauth>m Kuhn: m>¢o O. .r . 8 ' '--.g _ ‘ O DDDDDD . Inn-Innmvn m L '——' a R A A 8 § ' 11118180“ 1M33833 13.... 52.... 4 1‘8}. . atti. .55.... 55.6. 41118180" JNBOUBJ t: 024. +222 .5. .t .... .0. £3 3 a; Ki 1) £78 zouhc—wum2_ ::~:»»L muhuc w>¢o zo——¢~muuz~ t3~zh>L xuhuc m>¢o on on, 3 on o AIIWUIUOH 1N3383d All‘UlUOW 1NJDUBJ Figure 5. 42 Average plant height, width and volume of 'Ringo' Scarlet' geraniums over time transplanted into control, low, medium or high levels (0.0, 0.75, 1.5 or 5.0 g inoculum liter”1 medium) of Pythium ultimum- infested medium at 55 days. Letters indicate significant differences in treatment means based on Tukey's test. PLANT VOLUME (04’) PLANT HEIGHT (CM) 43 7 W ‘I’ V V I 1 W Y T Y Y ' T I f 35 4o 15 ‘ =50 55 DAYS AFTER PYTHIu'M INFESTATION 12- ’2‘ 8"- a E ‘ . §1O-1 WWI-WIH E hVGIS 5 oNoP o. vLPyt 9MP)" chPyt a- ...,Tfi..,....,-..:— 35 4o 45 so 55 DAYS AFTER PYTHIUM INFESTATION 4 300 r I’ r V r I’ fl I I I I Yfijfi V T T V 35 4O 45 50 55 DAYS AFTER PYTHIUM INF ESTATION 44 those growing in the low level of P. ultimum-infested medium. Response of 49 day old plants Plant mortality due to P. ultimum was low in the non-STS treated plants regardless of P. ultimum infestation level (Table 2): STS application increased percent mortality due to P. ultimum at all levels of P. ultimum infestation. Plant mortality incidence was 63% or greater in treatments combining both P. ultimum and STS (Table 2). No trends were observed as inoculum level increased. However, 30 days after transplant into P. ultimum-infested medium, plant mortality was at least 63% greater when STS was applied 14 days after infestation compared with STS application 0 or 7 days after infestation (Table 2). STS timing influenced rate of plant loss. During the 30 days following initial spraying, STS application at 0 or 7 days after transplanting into the P. ultimum-infested medium resulted in plant mortality similar to that of the control treatments (Figure 6). Application of STS 14 days after transplanting into the P. ultimum-infested medium, however, resulted in up to 90% plant death during a similar 30 day period. A second STS application 30 days after the first STS application to plants originally sprayed O or 7 days after transplant into P. ultimum-infested medium resulted in up to 45 .voaanao mos mam nuan3.uo aswvos woumwmcwuesaausa .M.uo voumoucwnao: cuafi waaunmanucouu nouns hon: .aswvoa voumowcuaEsEHuns aM.uo woumowcglno: can“ acmanmamuu nouwm mama oh acoauuomxo mo aoauuauauosx .cowuoowaamm mam Houuwca umuum whoa“ .asavoa mooHHuon mo umuHH won asaauoaw asawuas mM.maunuu «a om SS ca mm mm mm Nu Show e“ me o me o mm o Na o show u as o cos NH AA NA NA NA ammue o c 0 mm a ma 0 - o Houucoo Hanan on Hanan on Hmcfim on Hmcwm on x a omega ao-uuuwang< mam H\w o.m .~\w on.A H\m mh.o ~H\m o.o mHo>oH cowuoummucfl anaguas sawszM .mwcaswu e um Ampmv oumnaasmowsu uo>HHm :5 mm. zuas vmumouu can mmmv me um finance woummmcwuesswuas aswsuzm mo mHo>oH S can“ voucmaamcmuu cons :ofiumcaehou ucosfiuonxo um can :owumoaaaam mam umumm mzmv om mesficwuow .umHumom owcwm. mo Suwfimuuos ucouuom .N canoe Figure 6. 46 Percent mortality over time of 'Ringo Scarlet' geraniums transplanted into control, low, medium or high levels (0.0, 0.75, 1.5 or 5.0 g inoculum liter'1 medium) of Pythium ultimum- infested medium 49 days after seeding. Plants were treated with 0.25 mM silver thiosulphate (STS) at 0, 7 or 14 days following infestation or not at all. The arrow @—.9 indicates the first STS application. A second application (----) was made thirty days after the initial STS treatment. 47' zo:¢hwumz_ cauzzm «UTE wrco om on 8 4111618014 11433836 zoahchwuk: «5:32. xuhmm m>¢o om p>u-..v 2.2.... 2:9 Asanxzxr +23 0 a5 5 A1 118180“ “43383:! zcchnuuz_ ta~zp>m cure mrco h?‘ 2.7 h»; 1...: pt a... :c 021 .23: a5 A AIIWUIUDHIINHJNJQ .txi ..a..ie .€4¢ it!1 8.0 Oz if? A1 I 18180)! 1N3383d 48 100% plant mortality by the termination of the experiment (Table 2, Figure 6). Twenty-seven days after tranplanting, plants growing in the non-infested and low level of P. ultimum-infested medium were significantly larger than those plants growing in medium and high levels of P. ultimum-infested medium (Figure 7). At day 36, the plants grown in non-infested medium were significantly larger than those plants grown in P. ultimum-infested medium. By day 42 and 49, however, plants grown in low levels of P. ultimum-infested medium were not statistically smaller than the non-infested control. DISCUSSION Pythium ultimum disease symptoms on the seed propagated geranium were documented by this study. Geraniums in the P. ultimum-infested medium may not exhibit all of the typical root rot symptoms commonly associated with infection of Pythium spp. This study showed that symptoms present may be limited to stunted plants and a reduced root system. Pythium ultimum was isolated from plants with these symptoms. Abscence of chlorotic and wilted lower leaves may be misleading in diagnosing the cause of reduced plant size because several growing conditions may also result in low plant vigor including improper soil pH, high soluble salts and low nutrition levels. It is conceivable that an entire crop could be infested with Pythium spp. without the grower Figure 7. 49 Average plant height, width and volume of 'Ringo' Scarlet' geraniums over time transplanted into control, low, medium or high levels (0.0, 0.75. 1.5 or 5.0 g inoculum liter”1 medium) of Pythium ultimum- infested medium at 49 days. Letters indicate significant differences in treatment means based on Tukey's test. HEIGHT (CM) 7‘ 5O 5.. 5.. 1 ‘ W—f V Y r r V Y r 1 T ' ' '—[ ' ff V I fl j T 25 35 4O 45 50 DAYS AFTER PYTHIUM INFESTATION 12 4 [Pythium ‘1 levels 4 9 No Pyt A 10‘ o LPyt 2 U 1 4' MPyt V 9* v H t 33 , P' . 9 a a- a 7.. 6 Y j U—f T Y ' fi' 1' V r if 1 I Y fir V I j V 1 V 25 30 .35 4O 45 50 DAYS AFTER PYTHIUM INFESTATION 1 100 . 7 {Pythium 900. levels 0 No Pyt n’“ q I. P t 5 700- o y 8 + MPyt Lu '1 t a ' "” , _’ 500‘ O > - 3004 100 rrvTfi'vvrITfifIVfVVIVVVT .. 25 30 35 40 45 50 DAYS AFTER PYTHIUM INFESTATION 51 being aware of the resulting decrease in plant size if healthy plants were not available for comparision. Stunting is a typical symptom of root rot due to Pythium infection (7,16,22). Disease symptoms due to Pythium spp. include leaf chlorosis, partial defoliation, reduced growth and vigor on woody ornamentals. Such specimans exhibited a greatly reduced feeder root system and necrotic lesions on existing roots (14,15). Campbell and Sleeth (7) correlated the amount of plant stunting to the degree of Pythium injury to the root system. This study documented that there was significantly higher plant mortality among STS treated geraniums grown in P. ultimum-infested medium than plants grown in P. ultimum- infested medium without STS. This correlation between STS application and increased disease incidence has not been previously reported. Other chemicals appear to increase plant mortality incidence due to Pythium spp.. The herbicide glyphosate applied to bean seedlings at levels ineffective in causing mortality in sterilized soil increased disease incidence due to Pythium spp. in non-sterilized soil (12). Altman and Campbell (3) addressed the interaction of increased disease incidence following herbicide application. They summarized three major herbicide effects possibly leading to increased plant disease: 1) reduction of host structural defense; 2) stimulation of host exudation stimulating pathogen growth; 52 3) inhibition of microflora competing with potential pathogens. It is conceivable that STS may affect the plant resuling in increased P. ultimum disease incidence in a manner similiar to those proposed for herbicides. Another possibility is that STS may reduce the geranium's natural defense system. Phytoalexins produced by the plant have been shown to be toxic to fungi and bacteria. Phytoalexins have also been shown to be promoted by ethylene production stimulated by host tissue breakdown due to pathogen infection (18). Since the silver can block the action of ethylene (5), phytoalexin production or some other defense system may be reduced by STS application. Another possible mechanism involves exudation of STS through the root system resulting in exudates favorable for pathogen growth. This exudation could inhibit microflora which normally suppress pathogen growth. Studies show that STS is readily tranlocated within the plant (10), however, STS movement in the root system has not been documented. Favorable seed exudates have been shown to increase pathogen growth (2,9,19,20,27). In contrast, it has also been suggested that altering the delicate soil balance may result in increased Pythium disease incidence (1). No specific evidence is available to support one hypothesis over another. Results of this study suggest that foliar sprays of STS increased the decline in plants already infected with 53 P. ultimum. The longer the time for possible infection before STS application the quicker the plant death due to P. ultimum (Figure 4,6). Geraniums transplanted into the P. ultimum-infested medium 35 days after seeding required a minimum 7 day infection period to result in increased death during the first 30 days after STS application. However, geraniums transplanted into P. ultimum-infested medium 49 days after seeding required a minimum 14 day infection period to result in increased disease incidence within 30 days after STS application. This may be due to increased resistance of older plants to infection. Volume measurements of geraniums transplanted into P. ultimum- infested medium 35 and 49 days after seeding also support this. Over time, P. ultimum at higher levels continued to significantly affect plant size in geraniums transplanted into the P. ultimum-infested medium at 35 days. However, geraniums tranplanted into the P. ultimum-infested medium at 49 days showed no significant size difference between the non-infested and P. ultimum-infested treatments. This may indicate that geraniums transplanted at 49 days are more resistant or tolerant to invasion by P. ultimum than those geraniums transplanted at 35 days. Several studies substantiate this hypothesis. For instance, McCarter and Littrel (21) showed oats, wheat and cucumber were more susceptible in the early stages of germination than in late growth stages. Root rot severity in cotton seedlings decreased as the plants grew older (26). 54 Peach tree seedlings escaping pre-emergence and early postemergence mortality in infested soil grew to a size comparable to those in the non-infested soil (24). However, delaying transplant of geraniums from the plug tray to suitable growing containers is not recommended as a method of controlling P. ultimum or the interaction of P. ultimum with STS application. A second STS spray to plants transplanted at a later date resulted in a high mortality. LITERATURE CITED 1. Agnihotri, V.P. and 0. Vaartaja. 1968. Seed exudates from Pinus resinosa and their effects on growth and zoospore germination of Pythium afertile. Can. J. Bot. 46:1135-1141. 2. Agnihotri, V.P. and O. Vaartaja. 1970. Effect of seed exudates of Pinus resinosa on the germination of sporangia and the population of Pythium irregulare in the soil. Plant Soil 32:246-249. 3. Altman, J. and C.L. Campbell. 1977. Effect of herbicides on plant diseases. Ann. Rev. Phytopathol. 15:361-385. 4. Armitage, A.M. 1978. Seed geranium: timing, growth regulators and environmental problems. Pro. XI Intern. Bedding Plant Conf. pp. 149-151. 5. Beyer, E., Jr. 1976. Silver ion: a potent antiethylene agent in cucumber and tomato. HortScience 11:195-196. 6. Cameron, A.C. and M.S. Reid. 1981. The use of silver thiosulphate anionic complex as a foliar spray. II. Prevention of shattering in potted geraniums. HortScience 16:405. 7. Campbell, W.A. and B. Sleeth. 1945. A root rot of guayule caused by Pythium ultimum. Phytopathology 35:636-639. 8. Carlson, W.H. 1976. How growth retardants affect seed geranium varieties. Michigan State University Research Report 302:2-7. 9. Chang-Ho, Y. 1970. The effect of pea root exudate on the germination of Pythium aphanidermatum zoospore cysts. Can. J. Bot. 48:1501-1514. 10. Cook, E.L. and J. VanStaden. 1984. The translocation of pulsed radioactive silver thiosulphate within cut carnation flowers 2. Pflanzenphysiol. Bd. 113:177-181. 11. Farthing, J. and F. Chappell. 1982. Controlling petal shatter in Pelargoniums. Grower Feb. 18:32. 55 56 12. Gurmukh, J.S. and J.E. Rahe. 1984. Effect of soilborne plant-pathogenic fungi on the herbicidal action of glyphosate on bean seedlings. Phytopathology 74:950-955. 13. Heins, R.D., H.N., Fonda and A. Cameron. 1984. Mixing and storage of silver thiosulphate. BPI News 15:1-2. ' 14. Hendrix, F.F., Jr. and W.A. Campbell. 1966. Root rot organisms isolated from ornamental plants in Georgia. Plant Disease Reporter 50:393-395. 15. Hendrix, F.F, Jr., W.M. Powell and J.H. Owen. 1966. Relation of root necrosis caused by Pythium species to peach tree decline. Phytopathology 56:1229-1232 16. Klemmer, H.W. and R.Y. Nakano. 1964. Distribution and pathogenicity of Phytophthora and Pythium in pineapple soils of Hawaii. PlantfiDisease Reporter 48:848-852. 17. K0, W. and F.K. Hora. 1971. A selective medium for the quantitative determination of Rhizoctonia solani in soil. Phytopathology 61:707-710. 18. Kosuge, T. 1969. The role of phenolics in host response to infection. Ann. Rev. Phytopathol. 7:195-222. 19. Kraft, J.M. and D.C. Erwin. 1968. Effects of inoculum substrate and density on the virulence of Pythium aphanidermatum to mung bean seedling. Phytopathology 20. Kraft, J.M. and D.C. Erwin. 1967. Stimulation of Pythium aphanidermatum by exudates from mung bean seeds. Pfiytopathology 57:866-868. 21. McCarter, S.M. and R.H. Littrell. 1968. Pathogenicity of Pythium myriotylum to several grass and vegetable crops. Plant Disease Reporter 52:179-183. 22. McCarter, S.M. and R.W. Roncadori. 1971. Influence of low temperature during cotton seed germination on growth and disease susceptibility. Phytopathology 61:1426-1429. 23. Miranda, R.M. 1981. Studies on petal abscission in hybrid geranium. Ph.D. Thesis, Mich. State Univ., East Lansing. 24. Mircetich, S.M. and H.W. Fogle. 1969. Role of Pythium in damping-off of peach. Phytopathology 59:356-360. 57 25. Reid, M.S., J.L. Paul, M.B. Faroomand, A.M Kofranek and G.L. Staby. 1980. Pulse treatments with silver thiosulphate complex extend the vase life of cut carnations. J. Amer. Soc. Hort. Sci. 105:25-27. 26. Roncadori, R.W. and S.M McCarter. 1972. Effect of soil treatment, soil temperature and plant age on Pythium root rot of cotton. Phytopathology 62:373-376. 27. Royle, D.J. and C.J. Hickman. 1964. Analysis of factors governing in vitro accumulation of zoospoores of Pythium aphanidermatum on roots. II. Substances causing response Can. J. Microbiol. 10:201-219. 28. Stephens, C.T. and C.C. Powell. 1982. Pythium species causing damping-off of seedling bedding plants in Ohio greenhouses. Plant Disease 66:731-733. Section II: Efficacy of selected fungicides in controlling crown and root rot in 'Ringo Scarlet' geraniums caused by Pythium ultimum in the presence or absence of silver thiosulphate 59 Efficacy of Selected Fungicides in Controlling Crown and Root Rot in 'Ringo Scarlet' Geraniums Caused by Pythium ultimum in the Presence or Absence of Silver Thiosulphate Mary K. Hausbeck, Christine T. Stephens’ and Royal D. Heins Graduate Research Assistant and Associate Professors, respectively, Departmenta of Horticulture and Botany and Plant Pathology‘, Michigan State University, East Lansing, Michigan 48824-1312 ABSTRACT Hausbeck, M.K., C.T. Stephens and R.D. Heins. 1985. Efficacy of selected fungicides in controlling crown and root rot in 'Ringo Scarlet' geraniums caused by Pythium ultimum in the presence or absence of silver thiosulphate. Plant Disease 0:0. Fenaminosulf, ethazol or metalaxyl applied as soil drenches at selected times were effective in reducing crown and root rot disease caused by Pythium ultimum in geraniums when plants were not treated with silver thiosulphate (STS). Only metalaxyl consistently controlled the increased levels of disease caused by P. ultimum when plants were sprayed with silver thiosulphate. At selected application times, all three fungicides effectively decreased plant stunting symptoms and the delay of flowering associated with root rot caused by P. ultimum. However, metalaxyl and ethazol were generally, more effective than fenaminosulf. Fungicides also significantly affected plant size and flowering time of geraniums grown in the non-infested medium. 60 Additional key words: Pythium ultimum, Silver thiosulphate, fenaminosulf, metalaxyl, ethazol. INTRODUCTION Unlike the geraniums grown from cuttings, seed propagated geraniums experience premature flower petal abscission (1). This flower shattering generally occurs during transportation and has been a serious problem in marketing. Foliar application of silver thiosulphate (STS) solution prevents petal abscission in seed propagated hybrid geraniums (5,8,18). A correlation between STS application and increased disease incidence due to Pythium ultimum of plants grown in P. ultimum-infested medium has been documented (9). Plants grown in P. ultimum-infested medium and not treated with STS showed low mortality but exhibited a marked reduction in growth. Crown and root rot due to Pythium spp. is a common disease of geraniums and other greenhouse plants (11). Infection by Pythium on the seed propagated hybrid geranium may cause plant stunting, chlorotic and wilting of lower leaves, stem blackening, reduced root growth and root rot. One or more of these symptoms may be present. Plants with severe root rot generally will not develop black watersoaked stem lesions, which initiate the stem blackening and plant death phase of the disease. Plants with severe root rot symptoms may rot just below the soil line which ultimately 61 leads to death. However, apparently healthy plants with reduced root systems and stunted growth may develop a blackened stem and die with little warning to the grower after application of STS. The blackening progresses rapidly up the stem and into the base of the petioles. Control of Pythium crown and root rot is currently achieved through sanitation and/or preventative fungicides. Moorman (19) recommended heat or fumigant soil treatments as the preferred control method due to the suspected adverse effects of some fungicides on various plant species. Although Pythium spp. can be eliminated by heat or fumigant soil treatments, reintroduction of the pathogen is common in the bedding plant greenhouse (24). The fungicides ethazol (Truban), fenaminosulf (Lesan), metalaxyl (Subdue) and ethazol plus dimethyl 4,4-0-0 phenylenbis (Banrot) are recommended for prevention of crown and root rot caused by Pythium spp. (20,22). The objective of this study was to determine the effectiveness of selected Pythium-controlling fungicides in preventing plant mortality due to crown and root rot disease caused by P. ultimum in geraniums treated with or without STS. Due to the nature of this experiment, it was possible to determine the fungicide effect on geranium plant growth on plants grown in a non-infested medium and not treated with STS. 62 MATERIALS AND METHODS 'Ringo Scarlet' geranium seeds (Sluis A Groot B.V., Enkhuizen, Holland) were individually sown in round 2.0 cm in diameter cells, covered approximately 0.5 cm with 0 fine vermiculite and placed under intermittent mist at 24 C in a glass greenhouse. The seedlings were removed from the o mist after germination (7 days) and grown at 21 C night and o 24 C day temperatures under natural light in a glass greenhouse until tranplanting at 35 days from seeding. Inoculum of Pythium ultimum Trow was prepared using the potato media procedure developed for Rhizoctonia solani Kuhn culture (14). Fifty grams of finely chopped potato was added to 500 ml of growing medium (Sunshine Media Mix, Blend 1, Fisons-Western Corp., Vancouver, B.C., Canada) containing 2 parts vermiculite, 2 parts peat moss and 1 part perlite. After mixing, the potato media-mixture was autoclaved twice, 24 hours apart, for one hour. Pythium ultimum cultures were maintained on 20 ml of water agar (Difco Laboratories, Detroit, MI 48232) and grown in 10 cm petri plates for two days at 24°C. A highly virulent isolate of P. ultimum shown to cause crown and root rot in preliminary studies was used (25). Six mycelial discs 12 mm in diameter taken from the growing perimeter were used to infest 1.5 liters of sterilized potato soil media. After two weeks growth in 2.0 liter closed flasks, 63 the inoculum was air dried for 1-2 days and sieved through a #10 (2 mm) screen. Three grams of inoculum were mixed with a liter of the soilless medium. Previous studies indicate that high levels of P. ultimum crown and root rot occur in geraniums transplanted into this ratio of inoculum and medium (9). After thoroughly mixing, the infested soil was placed into single cells (8 x 8 x 6 cm) of 18 pack flats (25 x 53 cm) for the first experiment and in plastic pots (10 cm) for experiments 2 and 3. Eight single cells or pots were used to replicate each treatment. Non-infested and P. ultimum-infested treatment containers were randomized on greenhouse benches constructed of 14.0 cm wide wooden planks spaced 4.0 cm apart. To minimize P. ultimum contamination of non-infested plants, non-infested and P. ultimum-infested treatments were placed on alternating wooden planks. Treatments were maintained at 200C night and 22°C day temperatures. Medium pH varied between 5.5-6.5 during the expermiment. Plants were fertilized at each watering with a constant liquid feed containing 200 mg liter.1 each of N and K. Foliar applications of 750 ppm chlormequat chlorine (American Cyamide Co., Wayne, N.J. 07470) (CCC) were applied for height control as growth necessitated (2,7). A series of three fungicide screenings were conducted over the greenhouse growing season: Experiment 1- 64 September through November; Experiment 2- January through April; Experiment 3- April through July. Fungicide drenches were applied at the following times for each screening: Experiment 1 1) seeding 2) seeding and transplanting 3) seeding, transplanting and 1 week after transplanting 4) transplanting 5) 1 week after transplanting (1 week before STS application) 6) 2 weeks after transplanting (day of STS application) Experiment 2 1) seeding 2) seeding and transplanting 3) seeding, transplanting and 1 week after transplanting 4) transplanting 5) 8 weeks after transplanting (1 week before STS application) 6) 9 weeks after transplanting (day of STS application) Experiment 3 1) seeding 2) seeding and transplanting 3) seeding, transplanting and 1 week after transplanting 4) transplanting 5) 1 week after transplanting 6) seeding, transplanting and 7 weeks after transplanting 65 7) 7 weeks after transplanting (1 week before STS application) 8) 8 weeks after transplanting (day of STS application) The following fungicides were used at the label rate: fenaminosulf, 0.43 g ai/liter (Lesan 35% WP, Mobay Chemical Corp., Kansas City, MO 64120); ethazol, 0.11 g ai/liter (Truban 30% WP, Mallinkrodt, Inc., St. Louis, MO 63147); metalaxyl, 0.101 g ai/liter (Subdue 2E, 25% emulsifiable concentrate, Ciba-Geigy, Agricultural Division, Greensboro, NC 27409). A 0.25 mM silver thiosulphate (STS) solution (10,21) was sprayed to runoff 50, 61 or 52 days following transplant into P. ultimum-infested medium in Experiments 1-3 respectively. A second STS application was applied 14 days after each original STS treatment. Control plants received no treatment. Plants were observed daily for symptoms of Pythium crown and root rot. The number of days following transplant into P. ultimum-infested medium at which death occurred were recorded from time of transplant (day 0) into P. ultimum- infested medium through day 80 (Experiment 1), day 114 (Experiment 2) and day 75 (Experiment 3). Percent mortality was calculated. Plant height and width were recorded at the following times: 1) Experiment 1- days 51 and 98; 2) Experiment 2- day 135; 3) Experiment 3- days 84 and 105. Plant volume (size) was calculated from the height and width measurements, assuming the plant to be a cylinder. Days from seeding to flower were also recorded for Experiment 1 (non-infested and P. ultimum-infested treatments) and Experiment 3 (non-infested treatments). Plant measurements and flowering time were recorded only for plants that had not been treated with STS because the effect of STS on plant growth was not being investigated. Measurements on plants not treated with STS insures an accurate evaluation of the _ effects of fungicide and P. ultimum on plant growth. Reisolation of P. ultimum was conducted on geraniums that died during the study. At the termination of the experiment, surviving geraniums in P. ultimum-infested medium and non-infested controls were randomly sampled to determine if P. ultimum was present. One half inch sections of roots, stem and petioles were surface sterilized in 10% sodium hypochlorite for approximately 20 seconds and plated on water agar. The plates were observed for a minimum of seven days for Pythium hyphal growth. The agar plates with hyphal growth were allowed to dry at room temperature for three weeks to promote encysted sporangia development. The fungus was idenified as P. ultimum. RESULTS Experiment 1 No plant death occurred in the non-infested medium with or without STS (Results not shown) or the P. ultimum- infested medium without STS (Table 1). However, 88% mortality was observed on plants growing in the P. ultimum- infested medium and treated with STS. 67 .wawu u=05ummgu m>_gumammc sumo so» mcwumom Loewe mxmc we go um .om .o~ mm o u - u - up N- o N- o o + - - - o o o 0 mm o - + u u o o o o cm o - - + 1 o o o 0 mm o u + + + o o o 0 cm o u u + + oo- o mm o cm 0 - - u + mhm+ mew- mhm+ mpm- mkm+ mew- mpm+ mpm- mew mew mgowmn ucmpnmcmgp mcwummm Fo~osum Pxmemumz c-zmocwamcom Pocucou mo mama gum: F mvao-mcsm 1prpmugoz ucmugma Neowumow_aa< muqumczm co mauve?- .mm:wswu umuom_mm an mmuwowmcac mcw_PoLa:oo e=_;uxa ;a_z cognac» use ow ace :0 Am-mv mum;a_=mo_sp Lm>__m ze mN. zuwz umxmcam mew: mace-a .aawuoa umummccw-aaw;uxa oucw ace-amcmgu mcwzoppow mzmu om new ocwvomm cwucm mxau opp mazwcmcmm .um-cmom omcwm. co xpw_aagoa ucmucma .w opac- 68 Fungicide drench(es) reduced mortality on STS-treated plants grown in P. ultimum-infested medium. Metalaxyl and ethazol were more effective than fenaminosulf in preventing mortality of geraniums grown in the P. ultimum-infested medium and treated with STS. A fenaminosulf drench one week before STS or an ethazol drench at seeding did not reduce mortality in STS treated geraniums grown in the P. ultimum- infested medium compared to those without fungicide. Fifty-one day old geraniums grown in P. ultimum- infested medium with no fungicide treatment were 75% smaller than plants grown in the non-infested medium and not treated with fungicide (Table 2) (See Appendix A, Tables A1, A2 for corresponding height and width measurements). After 98 days, the plants were still 53% smaller (Table 3) (See Appendix A, Tables A3, A4 for corresponding height and width measurements). Fungicide drenches at 1) seeding or 2) one week after transplanting did not reduce plant stunting due to P. ultimum. Further, fenaminosulf drenches, regardless of application timing, did not reduce plant stunting in comparision to plants grown in the P. ultimum-infested medium without fungicide. The fungicide drenches affected size of plants grown in the non-infested medium (Table 2). On day 51, geraniums grown in the non-infested medium and treated with fenaminosulf drench(es) at 1) seeding, transplanting and 1 week after transplanting or 2) at transplanting only were significantly smaller than plants not drenched with 69 .—o>op mucouvcwco.a Achy npx .ov-u_m:=u guoo cvsu.x com.cogsou cowuau.—na_uuoamos sumo so; ac—coum Lowe. «new we so um .on .o~ 4:.2 8.88... 2 s: :3.- 8.882 m 833:9: :53 ~2sz ... «3295. m .m.z .u.a mosaom Nu I Aamv an: M.No mc— on- am pm pm om «mu 9" mm . mum esp um cow «an an opp on— no 2.8.2 8.832 S 3 x33... $.28- m 538:5 3.... 3888 N 3.29.3 u .m.: .u.a ousaom hmop - Anny an: ~m~ u u u . mum mw— map + u u 1 map o—w 0mm 1 + u . 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Ethazol applied 1 week after transplanting or metalaxyl 2 weeks after transplanting also resulted in significant growth reduction in comparision to the no fungicide control. By day 98, no significant size difference existed between plants treated or not treated with fungicide (Table 3). Plants drenched with fenaminosulf one week after transplant were, however, larger than plants not drenched with fungicide. Geraniums without fungicide and grown in the f. ultimum-infested medium showed a flowering delay in comparision to plants without fungicide and grown in non- infested medium (Table “,5). Flowering delay due to f. ultimum of geraniums grown in f. ultimum-infested medium was prevented by a metalaxyl drench at transplanting or a combination of ethazol drenches at seeding and transplanting (Table H). Time to flower of geraniums grown in non-infested medium was affected by application timings of fungicides (Table 5). Fenaminosulf drench(es) at 1) seeding, transplanting and 1 week after transplanting or 2) transplanting resulted in delayed flowering. An ethazol drench at seeding also delayed flowering. Experiment 2 Twelve percent mortality occurred due to 3. ultimum in geraniums grown in the f. ultimum-infested medium and not treated with either STS or fungicide drenches (Table 6) and 75% mortality when treated with STS. Metalaxyl, regardless ._m>m. acmuac_=m_m A..V a: .muwowmcam some :_;p:3 comwgmnsoo :o_umo:—aa< .mmmsmc pmowmopowmxzn op oat :o:um=:m>m Lo: mpnmpww>m megwume “cape acmwowwezmcux 72 Lozopu on when Neo:umowpaa< onwowmcam mo macmswh .mm:_ewu umuompmm um mmuwuwmcaw mcwppogucou sawcuxw gym: emummgu cog: on see no E=:cme umummmcpusawguxm ope: amazoFchphymE::cmgmm .umpgmum omcwm. co gmzopm op mxmu mmmgm>< .Q open: .ms_u acmspmmgu m>wuomammg some so: meromom Loewe want we so Rm .om .om Fm.p o~.mm: a < x m «emm.m m~.~mm m co:umo::na< gq.m¢.m ep.mmn N mewupmcal u .m.: .u.o mogaom xw.mp u Axmv om: m.oe_ - - - - ~.wmp o.emp 9.5m: + - - - m.mep m.mm— xmpnsem o: u + u . ¢.mmp m.m- «.mmp - - + - m.o~: 5.:mp o.mep - + + + mémp NKNP odmp u u + + ~.mmp :.oep o.mmp - - - + Fo~mgpm F»memumz epamocPEmcmm Fogucou pcmpnmcmgk pcm~amcmeh pcmpnmcmgp unwemmm gmowm ax: N Lmuwm x: P mmuwuwmczu 73 .mepu acmsummgu ._o>mp mucauwepcmvm A.v me .mewupmcze some cvsawz comwgmqeoo cowumoP—aamuomammg some Low mcwummm gmuwm ammo cc so mm .om .o~ NN.P mo.mN_ O: < x m x«PF.m NN.oNN m cowumUP—aa< we.: mm.mo_ N mu:o:m==u .u .m.z .u.n mogaom mo.NF u Ava om: N.NNF - - - . 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N: N: o mN o + - n - . mN o o o mN o - + - - - mm N: N: o mN o - - - + . mm o o o cm 0 - - + + + mu o o 0 mm Np - - - + + N: N: mm o mN o - u - - + mpm+ mpm- mhm+ mhm- mhm+ mhm- mhm+ mhm- mew mpm ucopamcmgp ueapamcagh m=Puomm pouoguu pzxupouoz epsmoe:2ocuu Posucou mo mxmo ogomon x: p Lopes x: p m¢u:u:qc:u Neowuou:pan< uu:u:me=m mo mmc:evh 3:33: Sue: .mmcwspu umuuopum «a mouvupmeaw mc:——ogpcou 53:;uxn my»: venues» use mm xav co Amhmv «ungapamo:zu Lu>~am :5 mN. saw: uoxagam use: mucupg .e=:uae umumom::-e::;u a one: acopamcugu ac::oppop mxuu «p. ecu m::uamm Loam. mane mm: 23::ogmm .uo—Loum ou=:¢. mo zu:pnasos Hecate; ..0 upon» 75 of application time, decreased plant mortality due to crown and root rot of geranium grown in the E. ultimum-infested medium. Specific drenches of fenaminosulf or ethazol were also effective. Metalaxyl drenches were more effective in preventing plant mortality on plants grown in the f. ultimum-infested medium treated with STS than either fenaminosulf or ethazol drenches (Table 6). However, 38% mortality still occurred when metalaxyl was applied at seeding. Plants grown in non—infested medium for 135 days and drenched with metalxyl were similar in size to plants not treated with a fungicide drench (Table 7) (See Appendix A, Tables A5, A6 for corresponding height and width measurements). However, plants grown in the non-infested medium and treated with a fenaminosulf drench at transplant or an ethazol drench 9 weeks after transplanting were larger than plants not treated with fungicide. Experiment 3 There was 14% plant mortality due to E. ultimum in STS treated plants grown in the non-infested medium (Results not shown). Thirty-eight percent mortality occurred in STS treated plants grown in the f. ultimum-infested medium without fungicide (Table 8). Drenches of fenaminosulf or metalaxyl were more effective than ethazol in preventing death due to E. ultimum in plants not treated with STS and grown in f. ultimum-infested medium. However, a metalaxyl 715 .Fu>o_ uu::u:.::m:m Rev mm .o Aeev n—x .oo:o:m:=. :uoo :psu—z :om:.:asou :opu:u_—anpuuoamuu gas: so» a:.:omm :aamo axe: no :o co .vc .um .o~ 3:8. 8 - - - - - 3:8 : 2 a 8. N. + - - - - 328 2 as: m... 8 2 - + - - - 8. 3 o o 3 a - + - + + m: 3 o N. 328 c - - + - - 838. N: o a 8 o - - - + - m: a o o :2 o - - + + + £58. 8 o o 8. c - - - + + 8. 3 E28. NP 8. 2 - - - - + 3+ m5- 3? as. E 3? m5- m5. as 2235.; 2235.; 83 .223. 3532 £3222: .228 3 £3 283 .... p :3: .... . awn—ummcmu‘ Nco'uouv—nn< ou.u.m:=u ma ma:.s.h xu.—~u::: u:ou:o: .mm:+s'u uuuuopum a: mac—u.m::m m:.ppo:u:ou Ezvguxx guwz nouoo:« v:~ am An: :o Amhmv ouogapamo.:u :u>p_m x: mw. :u.3 vuxagnm as»: m»::—: .sapuu: voumov:.-s:r:uxx ou:' u::—:m:~:u m:.x¢p—ou who: ms u:u m:*:oam :oaw: who: mp— ms:*:::mm .uu—:oum oa:*¢. :o aa_pau::s u:ou:o: .0 upon: 78 drench 1 week before STS application to plants grown in the E. ultimum- infested medium resulted in the same mortality as the no fungicide control. Foliar STS sprays applied to geraniums grown in the 3. ultimum-infested medium resulted in up to 100% mortality due to E. ultimum (Table 8). Properly timed metalaxyl drenches controlled mortality due to 3. ultimum on plants grown in the E. ultimum-infested medium and treated with STS with the exception of plants treated with a drench at seeding. Fenaminosulf and ethazol were ineffective in preventing mortality. Geraniums grown in the non-infested medium without fungicide treatment were 75% larger on day 84 than geraniums not treated with fungicide and grown in the 3. ultimum- infested medium (Table 9) (See Appendix A, Tables A7, A8 for corresponding height and width measurements). On day 8&, any fungicide drench applied at 1) seeding or 2) one week after transplant did not reduce plant stunting. Fenaminosulf drenches, regardless of timing, did not reduce plant stunting. Drenches at 7 or 8 weeks following transplant into E. ultimum-infested medium had not been applied at this measurement. Eighty-four days after seeding, plants grown in the non-infested medium and treated with fungicide were similar in size to those plants without fungicide (Table 9). However, plants treated with a metalaxyl drench one week after transplanting were significantly smaller compared to 79 «>0 ou:au. :u m .uku_m::u :uoo :.:w.3 hom.sonsww :wvuwmww :um .05» 2953.: 233:3: 53 :8 @533 not: «an: no .8 no .3 .5 .ON 3:. a 8. 233— : rand £6333 .— 18. mm -. 588: N 558292 8; 8.228 s 5582:? :2. 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On day 105, geraniums grown in the non-infested medium without fungicides were 83% larger than plants grown in the 3. ultimum-infested medium without fungicides (Table 10) (See Appendix A, Tables A9, A10 for corresponding height and width measuremnts). Regardless of fungicide, drenches at 1) seeding, 2) 1 week after transplant or 3) 8 weeks after transplanting did not reduce plant stunting. Some fungicide applications to plants grown in the non- infested medium resulted in reduced plant size on day 105 in comparision to plants without fungicide drenches (Table 10). The fenaminosulf treatment consisting of drenches at seeding, transplanting and 7 weeks after transplanting reduced plant growth significantly compared to plants not treated with fungicide (Table 10). A delay in flowering occurred in plants treated with drenches of fenaminosulf at seeding, transplanting and 1 week after transplanting (Table 11). DISCUSSION Fungicides at specific application times were effective in reducing crown and root rot disease caused by f. ultimum without STS treatment (Table 6,8). Other studies show fenaminosulf, ethazol and metalaxyl effectively control €31 .—o>ap «2.3.39.3 7.; a... 6339.3 53 553 5352.3 5382.3: .23 «5.58.5 03383.. 53 .3» 9:33 .33. a»... ma .3 3 .3 .sm .o~ :2 no no“ 333 2. (x... aw .p 3. $33 3. 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STS application to plants grown in f. ultimum-infested medium increased mortality due to f. ultimum as observed previously (9). Metalaxyl drenches at specific application timings consistently prevented increased incidence of plant death due to E. ultimum when plants were grown in E. ultimum- infested medium and treated with STS (Table 1,6,8). In contrast, fenaminosulf was ineffective in controlling increased death due to E. ultimum in response to foliar treatment with STS. Ethazol treatments gave inconsistent responses. In two of the three trials, ethazol treatments did not reduce mortality due to f. ultimum when plants were grown in 3. ultimum-infested medium and treated with STS. This investigation concluded that metalaxyl was superior in preventing mortality due to 2. ultimum in STS- treated geraniums grown in f. ultimum-infested medium. The inability of fenaminosulf and ethazol to control I. ultimum after STS application when these same fungicides were effective against 3. ultimum without STS is a matter of concern to the grower. It is proposed that STS application favors the pathogen (9) requiring stronger control fungicides. Apparently, metalaxyl is able to control 3. ultimum under these conditions. Results from this study suggest that plant size may be significantly affected by fungicides meant to control 84 Pythium spp. when the pathogen is not present. Whether a decrease or an increase of plant size occurs depends on the fungicide used and when it is applied. Plant size data were not consistent enough among the three screenings to determine at which developmental stage a geranium is most sensitive to a particular fungicide. Fungicide phytotoxicity has been suggested in other bedding plant studies. Stephens (23) showed that phytotoxicity of fenaminosulf and ethazol was dependent on the rate of application and growing medium used. Bolton (3) found geraniums to be very sensitive to fungicides applied as drenches. For instance, transplant into fenaminosulf treated medium resulted in plant chlorosis and stunting. Bolton's work is supported by the plant measurements taken on day 51 in the first screening. Plants grown in non- infested medium with a fenaminosulf drench at transplant were 68% smaller than the no fungicide control (Table 3). Significant stunting has been shown on the geranium 'Ringo Scarlet' when grown in 3. ultimum-infested medium compared to plants grown in a non-infested medium (9). In the first and third screenings, geraniums grown in the f. ultimum infested medium showed severe plant stunting typical of that caused by Pythium spp. (6,13,15). This was not the case in the second screening. However, the high mortality (62%) of STS-treated plants grown in non-infested medium suggests that these controls were contaminated with f. ultimum and do not constitute a control for comparision. 85 Metalaxyl or ethazol treatments at specific application timings to plants grown in E. ultimum-infested medium effectively reduced time to flower in comparision to the control. Fungicide application also affected time to flower when applied to geraniums grown in non-infested medium. In the first and third screenings, drenches of fenaminosulf at seed, transplant, and 1 week after transplant consistently delayed flowering (Table 7,10). Fungicide screening for effectiveness in controlling Pythium spp. has been investigated on several crops (“,12,16,17,26,27). However, recommendations from these studies may not be effective in situations where STS is utilized. Growers planning to use STS should consider a fungicide program which will provide the added control. Fungicide testing is important in formulating grower recommendations. Frequently, percent mortality is the only parameter evaluated in these studies. This particular study has attempted to define fungicide effectiveness through a variety of plant parameters. Data from this study are helpful in emphasizing that fungicide recommendations for control of f. ultimum on seed propagated geraniums should involve several factors; 1) the increased disease incidence due to STS if 3. ultimum is present, 2) the subtle symptoms of f. ultimum infection including growth stunting and delay of flowering that may occur without fungicidal control, and 3) the possible fungicide effects of geraniums without a pathogen. LITERATURE CITED 1. Armitage, A.M 1978. Seed geranium: timing, growth regulators and environmental problems. Pro. XI Intern. Bedding Plant Conf. pp. 1u9-151. 2. Ball, G.V. 1982. Growing geraniums from seed. pages 183-192 in: Geranium III. J.W. Mastalerz and E.J Holcomb, eds. Pennsylvania Flower Growers. #10 pp. 3. Bolton, A. Toxicity of Fungicide Drenches. Bull. Agriculture Canada. A. Baker, R. and G. Harman. 1981. Increased flower production by application of fungicides. Colorado Greenhouse Growers Assoc. Inc. Research Bulletin 376:1-fl. 5. Cameron, A.C. and M.S. Reid. 1981. The use of silver thiosulphate anionic complex as a foliar spray. II. Prevention of shattering in potted geraniums HortScience 16:”05. 6. Campbell, W.A. and B. Sleeth. 19fl5. A root rot of guayule caused by Pythium ultimum. Phytopathology 35:636-639. 7. Carlson, W.H. 1976. How growth retardants affect seed geranium varieties. Michigan State University Research Report 302:2-7. 8. Farthing, J. and F. Chappell. 1982. Controlling seed petal shatter in Pelargoniums. Grower Feb. 18:32. 9. Hausbeck, M.K, C.T. Stephens and R.D. Heins. 1985. Verification of induced plant mortality in 'Ringo Scarlet' geraniums when treated with silver thiosulphate. Plant Disease. In Review. 10. Heins, R.D., H.N. Fonda and A. Cameron. 198A. Mixing and storage of silver thiosulphate. BPI News 15:1-2 11. Hendrix, F.F., Jr. and W.A. Campbell. 1973. Pythiums as plant pathogens. Annu. Rev. Phytopathology 11:77-98. 86 87 12. Jones, J.P. 1961. Comparative effects of soil fungicides treatments on soil rot and damping-off of cucumber. Plant Disease Reporter ”5:376-379. 13. Klemmer, H.W. and R.T. Nakano 196A. Distribution and pathogenicity of Phytophthora and Pythium in pineapple soils of Hawaii. Plant’Disease Reporter 38:8u8-852. 1“. K0, W. and F.K. Hora. 1971. A selective mediumfor the quantitative determination of Rhizoctonia solani in soil. Phytopathology 61:707-710. 15. McCarter, S.M. and R.W. Roncadori. 1971. Influence of low temperature during cotton seed germination on growth and disease susceptibility. Phytopathology 61:1“26-1u29. 16. Miller, H.N. and R.T. DeNeve. 1971. Disease control on bedding plants with the use of 5-ethoxy-3- (trichloromethyl)-1,2,u-thiadiazole. Plant Disease Reporter 55:587-589. 17. Miller, H.N. and R.J. Saueve. 1975. Etiology and control of Pythium stem rot of geranium. Plant Disease Reporter 59:122-126. 18. Miranda, R.M. 1981. Studies on petal abscission in hybrid geranium. PhD Thesis, Mich. State Univ., East Lansing. 19. Moorman, G.W. 1983. Soilborne diseases. Pennsylvania Flower Growers Bulletin 7:6. 20. Powell, C.P. 1982. Diseases of seedling geraniums. Pages 262-265 in: Geraniums III. J.W. Mastalerz and E.J. Holcomb, eds. Pennsylvania Flower Growers. "10 pp. 21. Reid, M.S., J.L. Paul, M.B Faroomand, A.M. Kofranek and G.L. Staby. 1980. Pulse treatments with silver thiosulphate complex extend the vase life of cut carnation. J. Amer. Soc. Hort. Sci. 105:25-27. 22. Stephens, C.T. 198A. The glasshouse ornamental disease control handbook. Michigan State University extension bulletin E-1750:29-30. 23. Stephens, C.T. The effects of fungicides in different soil mixes. Bedding Plant Foundation Report. 2“. Stephens, C.T. and C.C. Powell. 1982. Pythium species causing damping-off of seedling bedding plants in Ohio greenhouses. Plant Disease 66:731-733. 88 25. Stephens, C.T., L.J. Herr, A.F. Schmitthenner and C.C. Powell. 1983. Sources of Rhizoctonia solani and Pythium spp. in a bedding plant greenhouse. Plant Disease 67:272-275. 26. Tammen, J. and D.F. Muse. 1961. Control of Pythium root and basal stem rot of Chrysanthemum morifolium with Dexon. Plant Disease Reporter 35:863-865. 27. Wheeler, J.E., R.B. Hine and A.M. Boyle. 1970. Comparative activity Dexon and Terrazole against Phytophthora and Pythium. Phytopathology 60:561-562. Section III: Variation in sensitivity to Pythium ultimum of selected seed propagated hybrid geranium cultivars 90 Variation in Sensitivity to Pythium ultimum of Selected Seed Propagated Hybrid Geranium Cultivars Mary K. Hausbeck, Christine T. Stephens* and Royal D. Heins Graduate Research Assistant and Associate Professors, respectively, Departments of Horticulture and Botany and Plant Pathology*, Michigan State University, East Lansing, Michigan “8824-1312 ABSTRACT Hausbeck, M.K., C.T. Stephens and R.D. Heins. 1985. Variation insensitivity to Pythium ultimum of selected seed propagated hybrid geranium cultivars. Plant Disease 0:0. While genetic resistance to Pythium ultimum was not identified in 37 cultivars screened, tolerance appeared to be present. Cultivars varied in the amount of plant stunting and flowering delay caused by P. ultimum. Silver thiosulphate increased plant mortality due to P. ultimum when plants were grown in a P. ultimum-infested medium. Additional key words: Pythium ultimum, geranium (Pelargonium x hortorum), silver thiosulphate 91 INTRODUCTON The seed propagated hybrid geranium (Pelargonium x hortorum) is commanding significant attention as an important crop in the bedding bedding plant industry. In 1983, approximately ten seed companies were developing hybrid geranium seed cultivars. More than 100 cultivars have been introduced, bringing annual production in 1983 to 90-100 million seeds (7). However, flower petal shatter is a severe problem in marketing the seed propagated geranium (2). This premature abscission occurs primarily during transportation. Foliar application of silver thiosulphate (STS) solution prevents petal abscission in seed propagated hybrid geraniums (6,8,17). Foliar sprays of STS increase disease incidence due to Pythium ultimum Trow in geraniums grown in E. ultimum- infested medium (10). Geraniums not treated with STS but grown in Pythium-infested medium may not show typical black stem lesions commonly associated with P. ultimum crown and root rot, but may have stunted plant growth, reduced root systems and chlorotic, wilted lower leaves indicative of Pythium root rot (10). The objectives of this experiment were: 1) to screen geranium cultivars for possible resistance to crown and root rot disease caused by P. ultimum in plants treated with or without STS and 2) to screen geranium cultivars not treated 92 with STS for possible tolerance to plant stunting due to root rot disease caused by P. ultimum. MATERIALS AND METHODS Forty-one seed propagated geranium (Pelargonium x hortorum) cultivars were chosen, encompassing a variety of colors, leaf types and seed sources (Table 1). The cultivars were grouped into four separate experiments due to variable seed and greenhouse space availability. Seeds were individually sown in round 2.0 cm in diameter cells, covered with fine vermiculite and placed under intermittant mist at 2AOC in a glass greenhouse. After germination (7 days), the seedlings were removed from the mist and grown at 220C day and 200C night temperatures under natural light in the greenhouse. They were fertilized at each watering with 200 mg liter.1 of N and K throughout the experiment. Pythium ultimum inoculum was prepared using the potato media procedure originally developed for culture of Rhizoctonia solani Kuhn (13). Fifty grams of finely chopped potato were added to 500 ml of a commercial soilless mix (Sunshine Media Mix, Blend 1, Fisons-Western Corp., Vancouver, B.C., Canada) containing 2 parts vermiculite, 2 parts peat moss and 1 part perlite. The potato media mixture was autoclaved twice for one hour, 24 hours apart. 93 Table 1. Selected seed propagated hybrid geranium cultivars, seed sources, flower colors and foliage characteristics grouped into 4 experiments. Each cultivar was screened for sensitivity to crown and root rot when grown in Pythium ultimum-infested medium and treated with or without silver thiosulphate. Cultivar Seed Source Flower Color Foliage Experiment 1 Jackpot S&Gz Scarlet Zoned Rosita Improved S&G Rose-pink Green Smash Hit Rose Pink Bally Br. rosewpink Zoned Smash Hit Salmon Ball Deep salmon Zoned Experiment 2 Cameo Ball Deep salmon Zoned Cheri Improved Goldx Soft salmon Zoned Encounter Salmon Ball Midsalmon Zoned Marathon Ball Deep scarlet Green Red Orbit Gold Bright red Zoned Showgirl Ball Rose-pink Green Smash Hit Ball Scarlet Zoned Snowdon Ball White Green White Orbit Gold White Green Experiment 3 Capri Deep Red PM" Dark red Zoned Jackpot S&G Scarlet Zoned Mustang S&G Red Zoned Picasso S&G Violet-cerise Zoned Red Elite Gold Red Green Red Pimpernel S&G Red Green Ringo Dolly S&G Bicolor Green Ringo Salmon S&G Deep salmon Zoned Ringo Scarlet S&G Bright scarlet Zoned Rosita Improved S&G Rose—pink Zoned 94 Table 1. (Cont.) Cultivar Seed Source Flower Color Foliage Experiment 4 Appleblossom Orbit Imp Gold Soft-pink Zoned Cherry Diamond Wale Cherry—red Zoned Expermental Rose Walz Soft-pink Zoned Heidi S&G Bicolor Green Hollywood Red Ball Red Zoned Hollywood Salmon Ball Deep salmon Zoned PAC Adretta Walz Coral red Zoned Pink Orbit Gold Rose-pink Zoned Pinwheel Salmon Harrisu Medium salmon Zoned Quix Walz Orange scarlet Green Ringleader Light Pink VJt Light pink Zoned Ringleader Red VJ Red Zoned Ringleader Salmon VJ Salmon Zoned Ringo Dolly SG Bicolor Green Ringo Scarlet SG Bright scarlet Zoned Scarlet Diamond Walz Bright scarlet Zoned Sitta Walz Salmon pink Zoned Sprinter Scarlet Gold Bright scarlet Green leuis & Groot B.V., Enkhuizen, Holland yBan Seed Co., West Chicago, IL 60185 xGoldsmith Seeds, Gilroy, CA 95020 wPan-America Seed Co., West Chicago, IL 60185 vWalz, Germany uHarris Seeds, Rochester, NY 1 14624 Vaughan—Jacklin, Downers Grove, IL 60515 95 Cultures of P. ultimum (#2u8) (20) were maintained on 20 ml of water agar (Difco Laboratories, Detroit, MI 48232) and grown in 10 cm petri plates for two days at ZAOC. Six mycelial discs 12 mm in diameter from the growing perimeter were used to infest 1.5 liters of sterilized potato soil media. The inoculum was grown in 2.0 liter closed flasks for approximately two weeks and shaken daily. The inoculum was air dried for 1-2 days and sieved through a #10 (2mm) screen. Three grams of inoculum (prepared as described above) were added to each liter of non-infested medium. This rate had been shown to incite a high level of disease due to P. ultimum (10). After thoroughly mixing, the infested medium was placed into 10 cm plastic pots. There were 12 replications for each treatment. Non-infested and P. ultimum-infested treatment containers were randomized on greenhouse benches comprised of 1u.o cm wooden planks ”.0 cm apart. To minimize P. ultimum contamination of non-infested plants, non-infested and P. ultimum-infested treatments were placed on alternating wooden planks. Benches were sterilized between experiments with a 10% sodium hypochlorite solution. Treatments were maintained at 200C night and 22°C day temperatures. Medium pH varied between 5.5-6.5 during the experiment. Foliar applications of 750 ppm chlormequat chlorine (American Cyanamid Co., Wayne, N.J. 07H?) (CCC) were applied for height control as growth necessitated (5). 96 A 0.25 mM foliar silver thiosulphate (STS) treatment (11,18) is commonly applied at flower bud color. STS was applied 100, 105, 102 or 95 days after seeding for Experiments 1 to a respectively. Each plant was individually sprayed. A second STS spray was applied two weeks after the original application. The control plants received no treatment. Plants were observed daily for symptoms of P. ultimum crown and root rot. The number of days following transplant into P. ultimum-infested medium at which death occurred were recorded, from time of transplant (day 0) into 3. ultimum-infested medium through day 9“, 93, 93 or 100 after transplanting for Experiments 1 to u respectively. Percent mortality was calculated. Plant height and width were recorded 83, 90 or 100 days after seeding for Experiments 1-3 and 67 and 103 days after seeding for Experiment u. Plant volume (size) was calculated from the height and width measurements, assuming the plants to be a cylinder. Days from seeding to flower were also recorded. Reisolation of P. ultimum was conducted on geraniums that died during the study. At the termination of the experiment, surviving geraniums in the 3. ultimum-infested medium and non-infested controls were randomly sampled. One half inch sections of roots, stem and petioles were surface sterilized in 10% sodium hypochlorite for approximately 20 seconds and plated on water agar. The plates were observed for a minimum of seven days for Pythium hyphal growth. The agar plates with hyphal growth were allowed to dry at room 97 temperature for three weeks to promote encysted sporangia development. The fungus was identified as P. ultimum. RESULTS Cultivar Set 1 Mortality prior to application of STS ranged from 9-3uz (Table 2). STS application increased mortality due to P. ultimum with mortality following STS application ranging from 27 to 53% The cultivar 'Smash Hit Rose Pink' showed the greatest loss throughout the experiment. All tested cultivars showed a significant reduction in plant size when transplanted into 3. ultimum-infested medium (Table 3) (See Appendix B, Table B1 for corresponding height and width measurements). This was accompanied by a delay in flowering. Surviving 'Smash Hit Rose Pink' geraniums in P. ultimum-infested medium showed the greatest volume reduction in comparision to control plants as well as the greatest flowering delay. Cultivar Set 2 The nine cultivars investigated had lower mortality overall than the previous cultivar set (Table A). The overall low mortality was also reflected in a smaller plant volume reduction due to P. ultimum-infestation (Table 5) (See Appendix B, Table B2 for corresponding height and width measurements. The cultivar 'Cheri Improved' showed a significant size reduction in P. ultimum-infested medium as well as a significant 5% delay in flowering. 98 Table 2. Percent mortality of selected geranium cultivars grown in non-infested (-P) or P thium-infested (+P) medium and treated with 0.25 mM si ver thiosulphate (STS). PERCENT MORTALITYZ Before STS Treatment After STS Treatmenty Cultivar -STS +STS -P +P -P +P -P +P Rosita Improved 3 9 0 0 0 53 Smash Hit Salmon 9 9 0 0 0 50 Jackpot 9 28 0 0 0 27 Smash Hit Rose Pink 3 34 0 0 0 50 2120 days after seeding and 94 days following Pythium infestation. YPercent mortality based on surviving plants at STS application on day 100. 99 ._m>m_ mocmowcwcmwm Acv um Lo Away u_x .ummuuu no woman pw>mp Aav am so Aaav up an acmuemmcm_m a+ tea a- consume mmocogwmmwcx .co_uaummwc. savsuxm ocpzoppom name Pm new unpuwwm Looms mxcu mm cmxau mucoEmgamumz~ .em.m n~.mk m axe ..Am.a~ mm.emo~e~m m axe ..Nm.mm ae.-o_ _ .exa .em5.mmp me.~mmogm~m _ .ssa 2.84 2.2 m .5 5.8.8, 8.3323 n .5 a .m.: .n.o unison a .m.: .n.o esteem P. .em.eo. ~.mm mm ..Nem. __mm seed deem s_: ennEm e .m.mm P.ee em ..mNOF Noe. eeepem s_= emuEm m ..m.eo_ _.mm mm ..oewp Pug, ee>eeees es.mem A ..~.mop ~.ea mw >weemm_ mmep neexuee ampmv a+ a- a+ a- gmzopu nosey; coeuusuwg A auv oE=~o> Lm>wupsu a o» made ma=Po> u N m acnpd .s=eeee Aa+v enamece_-e=_esxm go Aduv woummucwlcoc cw exocm mgn>_u_=o ancmgom vmuompmm mo Lozopm on name use m~wm mmmgm>< .0 open» 100 Table 4. Percent mortality of selected geranium cultivars grown in non-infested (-P) or Pythium-infested (+P) medium and treated with 0.25 mM silver thiosulphate (STS). PERCENT MORTALITYZ Cultivar Before STS Treatment After STS Treatmenty -STS +STS -P +P Ifi““'1b IP““‘:$ Showgirl o o o 31 9 o Cheri Improved 0 0 0 0 0 7 Red Orbit 0 0 0 0 8 8 White Orbit 0 0 0 7 l3 8 Encounter Salmon 0 0 0 0 O 9 Snowdon 0 0 0 0 0 9 Marathon 0 0 0 0 10 l3 Cameo 0 0 0 O 9 l5 Smash Hit 0 4 0 0 9 0 2140 days after seeding and 93 days following Pythium infestation. yPercent mortality based on surviving plants at STS application on day l05. 101 Table 5. Average size and days to flower of selected geranium cultivars grown in non-infested (-P) or Pythium-infested (+P) medium. % Plant Days to Cultivar volume (cm3)z % golume flower flower -P +P re uction :5—-—-;p- de ay White Orbit 1161 1262 103.5 104.5 1 Marathon 2676 2591 3 110.7 116.3” 5 Snowdon 1778 1661 7 103.1 100.6 Showgirl 2140 1964 8 102.7 104.3 2 Red Orbit 1809 1646 9 99.3 98.6 Cheri Improved 1648 1415 14 107.9 113.2** 5 Encounter Salmon 2099 1788 15 93.9 95.7 2 Cameo 1394 1183 15 96.3 98.1 2 Smash Hit 1723 1438 17 96.3 104.4* Source D.F. M.S. F Source D.F. M.S. F Cv. 8 9277263.l8 24.60"x Cv. 8 873.32 28.67** Pyt. l 2846307.23 7.55** Pyt. 1 292.32 9.60** Cxp 8 173117.75 .46 Cxp 8 61.36 2.01* zMeasuFeman? gaken 90 days after seeding and 62 days following Pythium n es a o . yDifferences between -P and +P significant at 1% (**) or 5% (*) level based on T-test. x1% (**) or 5% (*) significance level. 102 Cultivar Set 3 In the ten cultivars screened in Experiment 3, mortality ranged from 0 to 33% before STS application (Table 6). STS greatly increased the mortality of geraniums grown in P. ultimum-infested medium. The largest difference was seen in the cultivar 'Mustang' with a mortality increase of 55%. 'Ringo Dolly' showed no mortality either before or after STS. While 'Ringo Dolly' geraniums grown in P. ultimum- infested medium showed no mortality, plants were significantly smaller in comparision to the control (Table 7) (See Appedix B, Table B3 for corresponding height and width measurements). All the cultivars grown in P. ultimum-infested soil also showed reduced plant size in comparision to control plants. Flowering was delayed in 'Picasso' and 'Capri Deep Red' grown in P. ultimum-infested medium. Cultivar Set 4 Eighteen geranium cultivars were screened in this experiment. Overall, there was more P. ultimum mortality in this group (Table 8) than in the previous three screenings. Percent mortality before STS treatment ranged from 0 to 63%. STS application to geraniums grown in the P. ultimum- infested medium resulted in increased mortality due to P. ultimum. The cultivar 'Sprinter Scarlet' showed no plant loss prior to STS but 69% mortality after application. 103 Table 5- Percent mortality of selected geranium cultivars grown in non-infested (-P) or P thium—infested (+P) medium and treated with 0.25 mM si ver thiosulphate (STS). PERCENT MORTALITYZ Cultivar Before STS Treatment After STS Treatmenty -STS +STS -P +P -P +P -P +P Ringo Dolly 0 0 0 0 0 0 Ringo Scarlet 0 0 0 0 8 0 Red Elite 0 0 0 0 0 7 Red Pimpernel 0 0 0 0 0 8 Ringo Salmon 0 0 0 0 0 17 Rosita Improved 0 O 0 0 0 42 Picasso 0 4 0 0 8 42 Mustang 0 8 0 8 O 55 Jackpot 0 10 0 0 0 11 Capri Deep Red 6 33 0 13 11 20 2120 days after seeding and 93 days following Pythium infestation. Percent mortality based on surviving plants at STS application on day 102. 104 Table 7. Average size and days to flower of selected geranium cultivars grown in non-infested (-P) or Pythium-infested (+P) medium. . Plant % volume Days to % Cultivar volume (cm3)z reduction flower flower -P +P -P +P delay Ringo Scarlet 1983 1664**Y 16 97.9 99.3 1 Picasso 1849 1420** 23 99.9 108.3* 8 Red Elite 1448 1059** 27 99.3 102.4 3 Ringo Dolly 1518 1070** 30 100.1 101.9 2 Red Pimpernel 1897 1331** 30 98.5 101.4 3 Mustang 2390 1520** 36 100.5 101.7 1 Ringo Salmon 1542 948** 39 99.3 102.8 3 Capri Deep Red 1938 1001** 48 104.8 116.8* 10 Rosita Improved 2065 1060** 49 92.6 96.9 4 Jackpot 2261 1133** 50 92.9 96.9 4 Source D.F. M.S. F Source D.F. M.S. F Cv. 9 2805388.00 13.12**x Cv. 9 303.92 12.48** Pyt. 1 46397214.11 217.08** Pyt. 1 803.84 33.01** Cxp 9 905612.23 4.24** Cxp 9 50.24 2.06* ZMeasurements taken 100 days after seeding and 73 days following Pythium infestation. YDifferences between -P and +P significant at 1% (**) or 5 (*) level based on t-test. x1% (**) or 5% (*) significance level. 105 .mm awn co copuoow_aao new on mucopa mcw>w>gam co woman zappougoe ucmogodx .co»uoummwc_ sswguxu mcwzoppom mxmu cor can mcwpuomm Loewe mama omru mm mm om o mm o um>ogos~ pence sommopampaa< mm a mm o om m~ one: poucmswcoaxm mm o mm c on m ouumgu< oh~4_n_=o .Amhmv myocapamomzu Lm>pwm :5 mN.o cur: umummgu vcm Ezwume Am+v umummmcw Eamguxa Lo Aouv coummwcw-=oc cw czogo mco>wapao ancmcmm umuoopom we >u_Poucos acmugoa .m epoch 106 'Pink Orbit' showed 29% and 100% mortality before and after STS respectively. Mortality before STS was not a reliable indicator of subsequent response to STS. For example, 'Quix' showed only 12% mortality before STS but 89% after STS while 'Appleblossom Orbit Improved' showed 63% loss before STS treatment and 83% after application. . Pythium ultimum infestation resulted in a significant size reduction in all cultivars at«day 67 (Table 9) (See Appendix B, Table B4 for corresponding height and width meausurements). Plants growing in the P. ultimum-infested medium were still smaller at day 103 but not statistically in the three cultivars, 'Hollywood Salmon','Quix', and 'Sitta' (Table 10) (See Appendix B, Table BS for corresponding height and width measurements). There was, however, significant delay in flowering of these cultivars. DISCUSSION While each cultivar set was screened by the same procedure, comparisions between the screenings are not valid since each screening was conducted at a different time. The apparently increased plant mortality due to disease caused by P. ultimum in cultivar set 4 may be due to environmental factors. For example, cultivar set 4 experienced temperatures ranging from 20°C to 32°C whereas the previous 0 o 3 screenings were conducted in temperatures of 16 C-22 C. Greenhouse environments are more precisely controlled during 107 Table 9. Average size of selected geranium cultivars grown 1" non-infested (4’10? WEE-infested (+P) medium. Plant volume (cm3)z . % Cultivar -P +P Reduction Sprinter Scarlet 697 505 28 *' Heidi 1086 607 44 ** Quix 585 311 47 ** Experimental Rose 1059 476 55 ** Ringleader Red 754 307 59 ** Hollywood Salmon 751 290 61 ** PAC Adretta 1393 508 64 ** Ringo Scarlet 1679 605 64 ** Sitta 868 300 65 ** Pinwheel Salmon 1316 424 68 ** Scarlet Diamond 1361 408 70 ** Pink Orbit 616 178 71 ** Ringo Dolly 1356 381 72 ** Cherry Diamond 1136 300 74 ** Hollywood Red 717 175 76 ** Ringleader Pink 698 130 81 ** Ringleader Salmon 694 129 81 ** Appleblossom Orbit Improved 642 103 84 ** Source D.F. M.S. F Cv. 17 2312483.60 30. 92 Hr" Pyt. 1 71179843. 82 951.61 M Cxp 17 713857.37 9.5499 zMeasurements taken 67 days after seeding and 37 days following Pythium infestation. yDifferences between -P and +P significant at 1% (**) or 5% (*) level based on T-test. x1% (**) or 5% (*) significance level. ._o>op ooeeoac_ea_m Lev an to Area u_x .umouru no woman ~m>mp Atv um Lo Arav mp an acmupwpcmmm u+ can a- cmwzuma moucogaCCFQx .cowuoumowcp Enacuxu mcwzoppom mama me new mcwvomm condo mama mop coxo» mucosogammmx~ 108 oom~.~ o~.m- up axu armo.¢ om.-~mmm up nxu romm.mm mm.~om~ p .uxm «www.cmN me._m¢mmane p .u»: orcm.¢— mo.oupp up .>u xrtm~.m— mm.enpmopm up .>u u .m.: .m.o ousaom m .m.: .m.o mogaom m are.mop m.om _w arnmv mpmu umm Locum—ucpm mm o.o~p p.~m on swam ope. um>oan~ uvngo Eammo—noPna< m o.mo— m.mm we «rmme ommp com noozap—oz mp «rm.mpp m.mm mo ravme ovmp xcvm Loved—mcpm ep or~.om p.mm mo arena mmmm appoc omcpm e ~.¢m m.om mm «mom ~mmp omom poacoe_goaxu p P.mn ¢.- om aroma Nmop encamvo aopcmum mp rso.opp «.mm me «scum «mop venue sewn m m.mm p.em Ne «cum comp ecosopo accuse mp ra~.~pp m.mm cc ammo “pup cospum gouaopmcwm N m.mm m.om mm «rwmmp mmmm auuocu< u a Lw3OPm OH was NAMEUV g=FO> HEMFQ LM>WHP=U .esaeoe Aa+v eeamocee-e=_esxa to Aa-v eesmoce_-eoe cw czocm mgo>wu~3o azwcogom umuumpom we szopw on ammo can oNPm mungm>< .op «pomp 109 the winter (experiments 1-3) than during spring and early summer (experiment 4). Pythium ultimum Trow has historically been considered a cool season pathogen (3,12,14), although some evidence indicates otherwise (9). Greater water fluctuations would naturally accompany the higher temperatures experienced during the fourth screening. Periods of high water moisture followed by a very dry period has been connected to increased disease due to Pythium spp. (19). Results show a cultivar difference in mortality response to crown and root rot disease caused by P. ultimum. Surviving geraniums in f. ultimum-infested medium were significantly stunted in comparision to control plants but did not show any other root rot symptoms. Cultivar set 2, showed insignificant growth differences between geraniums grown in non-infested or E. ultimum- infested medium. Plant mortality was minimal. This substantiates previous studies which suggested that stunted geranium growth is often a symptom of root rot preceding the black stem lesions of crown rot leading to subsequent mortality (10). Resistance to P. ultimum was not identified within selected cultivars screened. However, tolerance to disease caused by P. ultimum appeared to be present in varying degrees as measured by plant growth and time to flower. It is possible that a cultivar grown in P. ultimum-infested medium which shows decreased mortality yet has greatly 110 stunted growth may be showing more tolerance than a cultivar which has high mortality but shows size reduction. STS treatment increased mortality due to P. ultimum crown and root rot. Geranium cultivars resistant to disease caused by this P. ultimum/STS interaction cannot be confidently identified. The geraniums 'Showgirl' and 'Smash Hit' in cultivar set 2 showed 0% mortality when grown in P. ultimum-infested medium and treated with STS. treatments. The overall low mortality in this group, however, suggests factors other than resistance to disease caused by P. ultimum are involved. In cultivar set 3, 'Ringo Dolly' and 'Ringo Scarlet' show no plant loss when grown in P. ultimum-infested medium and sprayed with STS. However, in cultivar set 4, these same cultivars showed 69 and 64% mortality respectively. A variety of crops have been shown to have genetic resistance to disease caused by P. ultimum (12) including bean cultivars (Phaseolus vulgaris) (1). However, resistance to P. ultimum was not found among species and varieties of cotton (15). Another study showed cotton and corn to be more resistant to Pythium spp. than tomato, bean and rye (16). Genetic resistance to disease caused by Pythium spp. has not been thoroughly investigated in the seed propagated hybrid geranium. Stephens et al. incuded two geranium cultivars in their search for P. ultimum disease resistance in bedding plant crops. Neither cultivar showed resistance 111 (Personal communication, Dr. Christine T. Stephens; Department of Botany and Plant Pathology, Michigan State University). The genetic recombination possible in the seed propagated geranium offers an opportunity for possible identification and subsequent development of Pythium resistance. Baker and Linderman (4) address the special problems involved with breeding for disease resistance in ornamentals. They point out that the success of newly introduced cultivars is often dependent on horticultural qualities rather than disease resistance. A disease tolerant cultivar will not be grown if the cultivar is not also horticulturally advantageous. They conclude that disease loss must be great enough to make breeding and subsequent development of disease resistant or tolerant cultivars that are also horticulturally acceptable cost effective. The large plant losses due to P. ultimum of geraniums grown in P. ultimum-infested medium and treated with STS may make this a worthwhile situation to explore for disease resistance or tolerance. Findings show that plant stunting may be the only symptom of P. ultimum root rot disease in otherwise healthy appearing geraniums. This provides little warning for the grower who may be unaware of plant stunting symptoms unless non-infected plants are available for comparision. It also emphasizes the importance of identifying cultivars which show tolerance to P. ultimum. LITERATURE CITED 1. Adegbola, M.O.K. and D.J. Hagedorn. 1970. Host resistance and pathogen virulence in Pythium blight of bean. Phytopathology 60:1477-1479. 2. Armitage, A.M. 1978. Seed geranium: timing, growth regulators and environmental problems. Pro. XI Intern. Bedding Plant Conf. pp. 149-151. ' 3. Arndt, C.H. 1943. Pythium ultimum and the damping- off of cotton seedlings. Phytopathology 33:607-611. 4. Baker, K.F. and R.G. Linderman. 1979. Unique features of the pathology of ornamental plants. Ann. Rev. Phytopathol. 17:253-277. 5. Ball, G.V. 1982. Growing geraniums from seed. Pages 183-192 in: Geranium III. J.W. Mastalerz and E.J. Holcomb, eds. Pennsylvania Flower Growers. 410 pp. 6. Cameron, A.C. and M.S. Reid. 1981. The use of silver thiosulphate anionic complex as a foliar spray. II. Prevention of shattering in potted geraniums. HortScience 16:405. 7. Craig, R. 1983. Geraniums for the 'BOs. Florist Review 173: 21-24. 8. Farthing, J. and F. Chappell. 1982. Controlling petal shatter in Pelargoniums. Grower Feb. 18:32. 9. Halpin, J.E., E.W. Hanson, and J.C. Dickson. 1952. Relative pathogenicity of seven species of Pythium on red clover seedlings. Phytopathology 42:10 (A5sEr.). 10. Hausbeck, M.K. 1985. Verification of induced Pythium ultimum mortality in 'Ringo Scarlet' geraniums when treated‘wfth silver thiosulphate. Plant Disease. In Review. 11. Heins, R.D., H.N., Fonda, and A. Cameron. 1984. Mixing and storage of silver thiosulphate. BPI News 15:1-2. 112 113 12. Hendrix, F.F., Jr. and W.A. Campbell. 1973. Pythiums as plant pathogens. Annu. Rev. Phytopathology 13. K0, W. and F.K. Hora. 1971. A selective medium for the quantitative determination of Rhizoctonia solani in soil. Phytopathology 61:707-710. 14. Leach, L.D. 1947. Growth rates of host and pathogen as factors determining the severity of pre-emergence damping-off. J. Agr. Res. 75:161-179. 15. Mathre, D.E. and J.D. Otta. 1967. Sources of resistance in the genus Gossypium to several soilborne pathogens. Plant Disease'Reporter 51:864-866. 16. McCarter, S.M. and R.H. Littrell. 1970. Comparative pathogenicity of Pythium aphanidermatum and Pythium myriotylum to twere plant species and intraspecific variation in virulence. Phytopathology 60:264-268. 17. Miranda, R.M. 1981. Studies on petal abscission in hybrid geranium. Ph.D. Thesis, Mich. State Univ., East Lansing. 18. Reid, M.S., J.L. Paul, M.B. Faroomand, A.M. Kofranek and G.L. Staby. 1980. Pulse treatments with silver thiosulphate complex extend the vase life of cut carnations. J. Amer. Soc. Hort. Sci. 105:25-27. 19. Sleeth, B. 1953. Winter Haven decline of citrus. Plant Disease Reporter 37:425-426. 20. Stephens, C.T. and C.C. Powell. 1982. Pythium species causing damping-off of seedling bedding plants in Ohio greenhouses. Plant Disease 66:731-733. APPENDIX A ._osop ooe.o.c.eu.n .... an .ouvupmcam sumo cvsuvz com.cugeou coauuu.—na_uuoamos :uuu so; ucpvuom Locus when cc so um .cn .9” x ..oo.m_ a~._~ o. and atom. ~_ as. A o. oonm.n on.. m eo.aao._oa< ..Nn.~ oo.. m eo.».o._eo< soon.” o..m ~ oo.o.ue=u o~._ oo._ N oo.o.ae=a a .m.: .n.a unison a .m.: .n.a outsom n.. . .um. on: so. . 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