Differential Sensitivity to Boscalid in Conidia and Ascospores of Didymella bryoniae and Frequency of Boscalid-Insensitive Isolates in South Carolina

Since 2003, a 2:1 mixture of the fungicides boscalid and pyraclostrobin (Pristine) has been used widely on watermelon and other cucurbits, primarily to control gummy stem blight caused by Didymella bryoniae. Several isolates of D. bryoniae that were insensitive to boscalid at 10 mg/liter were found in a watermelon research plot in South Carolina in 2008. In total, 201 isolates collected between 1998 and 2009 were tested for sensitivity to boscalid by determining percentage germination of ascospores and conidia on media amended with boscalid at 0.01 to 10.0 mg/liter. All 31 isolates collected in 1998, 2002, or 2005 were sensitive to boscalid. Of the 170 isolates collected in or after 2006, 84.7% were insensitive to boscalid, including 19 of 30 isolates recovered from greenhouse-grown seedlings. The oldest insensitive isolates were obtained in 2006 from a greenhouse and in 2008 from a commercial field. Ascospores were less sensitive to boscalid than conidia. With boscalid at 1.0 mg/liter, 22.4% of ascospores but only 4.1% of conidia of 31 sensitive isolates germinated. Similarly, a mean of 68.6% of the ascospores and 54.1% of the conidia of 120 insensitive isolates germinated at 1.0 and 10.0 mg/liter. The 50% effective concentration (EC50) values based on ascospore germination were two to three times higher than values based on conidia germination. The significance of miscalculating EC50 values by considering only conidia was demonstrated in a greenhouse experiment. Twelve isolates that were sensitive, moderately insensitive, or highly insensitive based on conidia germination did not differ in relative virulence on boscalid-treated muskmelon seedlings when inoculum suspensions comprised ascospores alone or ascospores and conidia. This is the first report of differential sensitivity to a fungicide between conidia and ascospores in D. bryoniae. Because D. bryoniae produces conidia and ascospores on diseased hosts, both spore types should be used when calculating EC50 values for boscalid.

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First Report of Benomyl Resistance in Didymella bryoniae in Delaware and Maryland

Plant Disease ◽

10.1094/pdis.1999.83.3.304b ◽

1999 ◽

Vol 83 (3) ◽

pp. 304-304 ◽

Cited By ~ 11

Author(s):

K. L. Everts

Keyword(s):

New York ◽

South Carolina ◽

Citrullus Lanatus ◽

New York State ◽

Didymella Bryoniae ◽

Potato Dextrose Agar Medium ◽

First Report ◽

Stem Blight ◽

Colony Diameter ◽

Gummy Stem Blight

Gummy stem blight, caused by Didymella bryoniae (Auersw.) Rehm, is the most severe foliar disease of watermelon, Citrullus lanatus (Thunb.) Matsum. & Nakai, in eastern Maryland and southern Delaware. The fungicide benomyl is used in combination with chlorothalonil to manage gummy stem blight. Under conducive environmental conditions, yield losses are high even when fields are sprayed weekly. Resistance of D. bryoniae to benomyl has been reported in New York State and South Carolina (1). Gummy stem blight-infected leaves and stems were collected from nine and three fields in Wicomico County, MD, and Sussex County, DE, respectively, in 1996. Infected tissue was also collected from two Wicomico County fields in 1997. One single-spore subculture was obtained to represent each field. Agar plugs were taken from actively growing subcultures and inverted on a 25% (quarter strength) potato dextrose agar medium amended with 0 and 33.1 mg of benomyl per liter, the concentration of benomyl that reduced relative colony diameter of four resistant isolates in New York and South Carolina by 50% (1). Two replicate plates were used per experiment and each experiment was repeated once. After 6 days of growth at 21°C in the dark, the colony diameter was measured. Isolates were classified as sensitive if they were unable to grow, moderately sensitive if colony diameter was reduced 40 to 60%, and resistant if colony diameter was reduced less than 10% on the benomyl-amended media, compared with unamended media. Isolates that had previously been tested were used as sensitive (W03) and moderately sensitive (NY1) standards (1). In 1996, two isolates were sensitive, four isolates were moderately sensitive, and six isolates were resistant to benomyl. One isolate from 1997 was resistant and the other was moderately sensitive. This is the first report of resistance to benomyl within the D. bryoniae population in eastern Maryland and southern Delaware. Reference: (1) A. P. Keinath and T. A. Zitter. Plant Dis. 82:479, 1998.

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Boscalid Insensitivity Documented in Didymella bryoniae Isolated from Watermelon in Florida and North Carolina

Plant Health Progress ◽

10.1094/php-2012-0518-01-br ◽

2012 ◽

Vol 13 (1) ◽

pp. 42

Author(s):

Katherine L. Stevenson ◽

Anthony P. Keinath ◽

Anna Thomas ◽

David B. Langston ◽

Pamela D. Roberts ◽

...

Keyword(s):

North Carolina ◽

South Carolina ◽

High Frequency ◽

In Vitro Assays ◽

Didymella Bryoniae ◽

Triazole Fungicides ◽

Stem Blight ◽

Gummy Stem Blight ◽

Commercial Mixture

The fungicide Pristine, a commercial mixture of pyraclostrobin and boscalid, has been used widely on watermelon and other cucurbits to control gummy stem blight, caused by the fungus Didymella bryoniae. Since 2007, isolates of D. bryoniae insensitive to boscalid have been found in Georgia, Indiana, and South Carolina. Most isolates of D. bryoniae obtained in 2009 and 2010 from diseased watermelon leaves collected in several counties in Florida and North Carolina were found to be insensitive to boscalid using in vitro assays. Gummy stem blight would not be effectively managed with Pristine in any of the counties where samples were collected due to the high frequency of insensitivity to boscalid. Fortunately growers can instead use several triazole fungicides registered for use on cucurbits since 2009. Accepted for publication 30 March 2012. Published 18 May 2012.

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Baseline Sensitivity of Didymella bryoniae to Cyprodinil and Fludioxonil and Field Efficacy of these Fungicides Against Isolates Resistant to Pyraclostrobin and Boscalid

Plant Disease ◽

10.1094/pdis-09-14-0993-re ◽

2015 ◽

Vol 99 (6) ◽

pp. 815-822 ◽

Cited By ~ 13

Author(s):

Anthony P. Keinath

Keyword(s):

South Carolina ◽

Citrullus Lanatus ◽

Control Treatment ◽

Didymella Bryoniae ◽

Water Control ◽

Baseline Sensitivity ◽

Qoi Fungicides ◽

Stem Blight ◽

Gummy Stem Blight ◽

Development Of Resistance

To prevent yield reductions from gummy stem blight, fungicides often must be applied to watermelon (Citrullus lanatus) and muskmelon (Cucumis melo). Didymella bryoniae, the ascomycete fungus that causes gummy stem blight, is resistant to thiophanate-methyl, quinone-outside inhibitors (QoI), boscalid, and penthiopyrad. In place of these fungicides, premixtures of cyprodinil and fludioxonil (Switch 62.5WG) or cyprodinil and difenoconazole (Inspire Super 2.82SC) are used. The objectives of this study were to examine baseline isolates of D. bryoniae for sensitivity to cyprodinil and fludioxonil and to determine the efficacy of cyprodinil-fludioxonil and cyprodinil-difenoconazole against isolates resistant to QoI fungicides and boscalid. Colony diameters of 146 isolates of D. bryoniae collected in South Carolina and other U.S. states prior to 2008 were measured on glucose minimal medium amended with cyprodinil or fludioxonil. Mean effective concentration values that reduced relative colony diameter by 50% were 0.052 and 0.099 mg/liter cyprodinil and fludioxonil, respectively. In autumn 2008, 2009, and 2011, field-grown watermelon inoculated with isolates resistant to QoI fungicides and boscalid was treated with boscalid-pyraclostrobin alternated with chlorothalonil, cyprodinil-fludioxonil alternated with chlorothalonil, cyprodinil-difenoconazole alternated with chlorothalonil, tebuconazole alternated with chlorothalonil, chlorothalonil, or water. In 2008 and 2011, both cyprodinil treatments reduced disease severity compared with the water control treatment and chlorothalonil alone. In 2008 and 2009, cyprodinil-fludioxonil reduced severity compared with boscalid-pyraclostrobin and, in 2008, cyprodinil-difenoconazole and tebuconazole also did. Use of cyprodinil-fludioxonil should control gummy stem blight effectively and may delay development of resistance to cyprodinil and fludioxonil in D. bryoniae. However, because Botrytis cinerea became resistant to both cyprodinil and fludioxonil after multiple applications of cyprodinil-fludioxonil per season, prudent fungicide rotations should be followed when using cyprodinil-containing fungicides against D. bryoniae.

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First Report of Pycnidial Stage of Didymella bryoniae, the Causal Fungus of Gummy Stem Blight on Cantaloupe (Cucumis melo var cantalupensis) in Egypt

Egyptian Journal of Phytopathology ◽

10.21608/ejp.2016.92386 ◽

2016 ◽

Vol 44 (1) ◽

pp. 231-232

Author(s):

Abdel-Fattah El-Wakil ◽

Amal Khalil

Keyword(s):

Cucumis Melo ◽

Didymella Bryoniae ◽

First Report ◽

Stem Blight ◽

Gummy Stem Blight ◽

Causal Fungus

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Survival of Didymella bryoniae in Buried Watermelon Vines in South Carolina

Plant Disease ◽

10.1094/pdis.2002.86.1.32 ◽

2002 ◽

Vol 86 (1) ◽

pp. 32-38 ◽

Cited By ~ 21

Author(s):

Anthony P. Keinath

Keyword(s):

South Carolina ◽

Crop Residue ◽

Inoculum Density ◽

Didymella Bryoniae ◽

Gummy Stem Blight ◽

Infection Assay ◽

Nylon Mesh ◽

Mesh Bags ◽

Number Of Segments ◽

Over Time

Sections of dead, dried watermelon vines naturally infected with Didymella bryoniae were placed in nylon mesh bags, covered with soil, and placed in a field at 0-, 12.5-, or 25-cm depths in the November or December prior to 1997, 1998, and 1999. The percentage of 1-cm vine segments that yielded D. bryoniae and the number of segments retrieved intact declined over time. Vine segments cultured on semiselective medium yielded D. bryoniae for 30, 24, and 21 weeks after placement in 1997, 1998, and 1999, respectively. D. bryoniae was not recovered 32 and 29 weeks after placement at any depth in 1998 and 1999, respectively. A hypocotyl-infection assay using watermelon seedlings was developed to detect D. bryoniae conidia produced on recovered vine sections. Using known concentrations of conidia, the percentage of seedlings exhibiting gummy stem blight symptoms increased with the logarithm of the inoculum density. Viable conidia were produced on retrieved vine sections for up to 32 weeks after burial, based on culturing on semiselective agar, but conidia caused disease for only 16 weeks after burial in the hypocotyl-infection assay. Conidia of D. bryoniae were recovered 8 weeks longer at 0 cm than at lower depths in 1999, but not in 1998. Incorporating infested crop residue into soil reduced the number of infective propagules and survival of D. bryoniae.

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First report of gummy stem blight caused by Didymella bryoniae on muskmelon (Cucumis melo) in India

Plant Pathology ◽

10.1111/j.1365-3059.2004.01031.x ◽

2004 ◽

Vol 53 (4) ◽

pp. 533-533 ◽

Cited By ~ 5

Author(s):

J. Sudisha ◽

T. Vasanth Kumar ◽

S. R. Niranjana ◽

H. Shekar Shetty

Keyword(s):

Cucumis Melo ◽

Didymella Bryoniae ◽

First Report ◽

Stem Blight ◽

Gummy Stem Blight

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Didymella bryoniae Isolates from Watermelon Seedlings in Florida Transplant Houses and Their Sensitivity to Boscalid

Plant Health Progress ◽

10.1094/php-br-14-0047 ◽

2015 ◽

Vol 16 (1) ◽

pp. 31-33 ◽

Cited By ~ 1

Author(s):

Katherine E. Hendricks ◽

Pamela D. Roberts

Keyword(s):

South Carolina ◽

Molecular Mechanism ◽

Previous History ◽

Didymella Bryoniae ◽

Stem Blight ◽

Gummy Stem Blight ◽

History Of ◽

North And South

The purpose of this study was to examine why gummy stem blight was not being controlled in watermelon fields without a previous history of boscalid use and whether boscalid resistance could have originated from the transplants. The study evaluated Didymella bryoniae isolates collected from three Florida transplant houses for resistance to boscalid and compared the molecular mechanism of this resistance to that in the literature on isolates from North and South Carolina and Georgia. Accepted by publication 16 February 2015. Published 25 February 2015.

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First Report of Gummy Stem Blight Caused by Didymella bryoniae on Watermelon and Confirmation of the Disease on Pumpkin in Tanzania

Plant Disease ◽

10.1094/pdis-01-11-0044 ◽

2011 ◽

Vol 95 (6) ◽

pp. 768-768 ◽

Cited By ~ 7

Author(s):

B. D. Jensen ◽

A. Massawe ◽

I. S. Swai

Keyword(s):

Citrullus Lanatus ◽

Morphological Characteristics ◽

Pathogenic Fungi ◽

Weather Conditions ◽

Fruit Rot ◽

Didymella Bryoniae ◽

First Report ◽

Stem Blight ◽

Gummy Stem Blight ◽

Specific Pcr

Foliar, stem, and fruit lesions were observed on watermelon (Citrullus lanatus (Thunb.) Matsum. & Nakai) and pumpkin (Cucurbita maxima Duchesne) in two separate research fields in the district of Arusha, Tanzania during the warm, rainy season from February to April 2010. Similar symptoms were observed in commercial watermelon fields and intercropped pumpkin fields in Same and Moshi districts with as much as 100% fruit loss in watermelon. Disease symptoms on watermelon were dark brown, V-shaped leaf lesions. On pumpkin, V-shaped leaf lesions were light brown. On both hosts, stems showed water-soaked lesions after rain, which dried up and cracked. On pumpkin, a gummy, amber exudate was seen after rain on stem and fruit lesions. Flowers and fruits of both hosts developed black rot spots and aborted. Isolation of the causal agent on potato dextrose agar (PDA) from leaf and stem pieces of watermelon and pumpkin plants in Arusha showed white-to-olivaceous green mycelium. Pycnidia formed on one-quarter-strength PDA and produced hyaline, oblong conidia mainly with two guttules, nonseptate, 5 to 11 × 3 to 5 μm. Pathogenicity was tested with three isolates from watermelon and one from pumpkin on four 1-month-old plants per watermelon cvs. Sugar Baby and Charleston Grey and pumpkin cv. Small Sugar per isolate. The test was repeated on the watermelon cultivars. One site on the main stem and two leaves per plant were misted, pricked with a scalpel, inoculated with 3-day-old mycelial plugs (5 × 5 mm), and kept humid at 20 to 30°C in cellophane bags for 3 days. All plants developed leaf and/or stem lesions. Detached, misted leaves were also laid on 2% water agar and inoculated as above. Water-soaked lesions developed around inoculation sites and microscopy of infected tissue revealed pycnidia with conidia as described above. All isolates infected both hosts. A set of control plants and detached leaves, mock inoculated with agar plugs, remained symptomless. The fungus was reisolated from infected leaves and stems of both hosts. On the basis of the morphological characteristics, the fungus was identified as Didymella bryoniae (Auersw.) Rehm (anamorph Phoma cucurbitacearum (Fr.:Fr.) Sacc.) (1,3) and this was confirmed by amplification of species-specific PCR products. The isolates from both hosts were cultured in liquid medium, and DNA was extracted using a DNeasy Plant Mini Kit (Qiagen, Valencia, CA). PCR and multiplex PCR involving D. bryoniae-unique primer sequences D6 and D7S, in combination with primer UNLO28S22, produced the expected band sizes (2). To our knowledge, this is the first report of gummy stem blight and black fruit rot of watermelon caused by D. bryoniae in Tanzania, which confirms a previous report of leaf spot on pumpkin (4), and the first report of black fruit rot on pumpkin. The disease was previously an unidentified problem in watermelon and the severe outbreak was associated with favorable weather conditions. References: (1) A. P. Keinath et al. Phytopathology 85:364, 1995. (2) C. A. Koch and R. S. Utkhede. Can. J. Plant Pathol. 26:291, 2004. (3) E. Punithalingam and P. Holliday. No. 332 in: Descriptions of Pathogenic Fungi and Bacteria. CMI, Kew, Surrey, UK, 1972. (4) E. A. Riley. Mycol. Pap. 75:1, 1960.

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