Development of a diagnostic assay for race differentiation of Podosphaera macularis

Hop powdery mildew (caused by Podosphaera macularis) was confirmed in the Pacific Northwest in 1996. Before 2012, the most common race of P. macularis was able to infect plants that possessed powdery mildew resistance based on the R-genes Rb, R3, and R5. After 2012, two additional races of P. macularis were discovered that can overcome the resistance gene R6 and the partial resistance found in the cultivar Cascade. These three races now occur throughout the region, which can complicate management and research efforts because of uncertainty on which race(s) may be present in the region and able to infect susceptible hop genotypes. Current methods for determining the races of P. macularis are labor intensive, costly, and typically require more than 14 days to obtain results. We sought to develop a molecular assay to differentiate races of the fungus possessing virulence on plants with R6, referred to as V6-virulent, from other races. The transcriptomes of 46 isolates of P. macularis were sequenced to identify loci and variants unique to V6-isolates. Fourteen primer pairs were designed for 10 candidate loci that contained single nucleotide polymorphisms (SNP) and short insertion-deletion polymorphisms. Two differentially-labeled locked nucleic acid probes were designed for a contig that contained a conserved SNP associated with V6-virulence. The resulting duplexed real-time PCR assay was validated against 46 V6 and 54 non-V6 P. macularis isolates collected from the United States and Europe. The assay had perfect discrimination of V6-virulence among isolates of P. macularis originating from the western U.S. but failed to predict V6-virulence in three isolates collected from Europe. The specificity of the assay was tested with different species of powdery mildew fungi and other microorganisms associated with hop. Weak non-specific amplification occurred with powdery mildew fungi collected from Vitis vinifera, Fragaria sp., and Zinnia sp.; however, non-specification amplification is not a concern when differentiating pathogen race from colonies on hop. The assay has practical applications in hop breeding, epidemiological studies, and other settings where rapid confirmation of pathogen race is needed.

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Erysiphe trifolii Causing Powdery Mildew of Lentil (Lens culinaris)

Plant Disease ◽

10.1094/pdis-93-8-0797 ◽

2009 ◽

Vol 93 (8) ◽

pp. 797-803 ◽

Cited By ~ 22

Author(s):

Renuka N. Attanayake ◽

Dean A. Glawe ◽

Frank M. Dugan ◽

Weidong Chen

Keyword(s):

Powdery Mildew ◽

Pacific Northwest ◽

The United States ◽

Morphological Characters ◽

Grain Legumes ◽

Its Sequences ◽

Powdery Mildew Fungus ◽

The Pacific ◽

Soybean Genotypes ◽

Erysiphe Trifolii

The taxonomy of the powdery mildew fungus infecting lentil in the Pacific Northwest (PNW) of the United States was investigated on the basis of morphology and rDNA internal transcribed spacer (ITS) sequences. Anamorphic characters were in close agreement with descriptions of Erysiphe trifolii. However, teleomorphs formed chasmothecial appendages with highly branched apices, whereas E. trifolii has been described as producing flexuous or sometimes loosely branched appendages. Branched appendages have been described in Erysiphe diffusa, a fungus reported from species of Lens, Glycine, and Sophora, raising the possibility that the PNW fungus could be E. diffusa. Examination of morphological characters of an authentic specimen of E. trifolii from Austria determined that it included chasmothecial appendages resembling those seen in PNW specimens. Furthermore, ITS sequences from five powdery mildew samples collected from lentils in PNW greenhouses and fields from 2006 to 2008 were identical to one another, and exhibited higher similarity to sequences of E. trifolii (99%) than to those of any other Erysiphe spp. available in GenBank. Parsimony analysis grouped the lentil powdery mildew into a clade with Erysiphe baeumleri, E. trifolii, and E. trifolii–like Oidium sp., but indicated a more distant relationship to E. diffusa. In greenhouse inoculation studies, the lentil powdery mildew fungus did not infect soybean genotypes known to be susceptible to E. diffusa. The pathogenicity of E. trifolii on lentil was confirmed using modified Koch's postulates. This is the first report of E. trifolii infecting lentil. E. diffusa and E. trifolii have different host ranges, so the discovery of E. trifolii on lentil has implications both for determining species of powdery mildews on cool-season grain legumes, and in disease management.

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Distribution and Characterization of Podosphaera macularis Virulent on Hop Cultivars Possessing R6-Based Resistance to Powdery Mildew

Plant Disease ◽

10.1094/pdis-12-15-1449-re ◽

2016 ◽

Vol 100 (6) ◽

pp. 1212-1221 ◽

Cited By ~ 12

Author(s):

Sierra N. Wolfenbarger ◽

Stephen T. Massie ◽

Cynthia Ocamb ◽

Emily B. Eck ◽

Gary G. Grove ◽

...

Keyword(s):

United States ◽

Powdery Mildew ◽

Mating Type ◽

Resistance Genes ◽

Severe Disease ◽

The United States ◽

Fitness Cost ◽

Eastern United States ◽

Host Genotype ◽

Podosphaera Macularis

Host resistance, both quantitative and qualitative, is the preferred long-term approach for disease management in many pathosystems, including powdery mildew of hop (Podosphaera macularis). In 2012, an epidemic of powdery mildew occurred in Washington and Idaho on previously resistant cultivars whose resistance was putatively based on the gene designated R6. In 2013, isolates capable of causing severe disease on cultivars with R6-based resistance were confirmed in Oregon and became widespread during 2014. Surveys of commercial hop yards during 2012 to 2014 documented that powdery mildew is now widespread on cultivars possessing R6 resistance in Washington and Oregon, and the incidence of disease is progressively increasing. Pathogenic fitness, race, and mating type of R6-virulent isolates were compared with isolates of P. macularis lacking R6 virulence. All isolates were positive for the mating type idiomorph MAT1-1 and were able to overcome resistance genes Rb, R3, and R5 but not R1 or R2. In addition, R6-virulent isolates were shown to infect differential cultivars reported to possess the R6 gene and also the R4 gene, although R4 has not yet been broadly deployed in the United States. R6-virulent isolates were not detected from the eastern United States during 2012 to 2015. In growth chamber studies, R6-virulent isolates of P. macularis had a significantly longer latent period and produced fewer lesions on plants with R6 as compared with plants lacking R6, indicating a fitness cost to the fungus. R6-virulent isolates also produced fewer conidia when compared with isolates lacking R6 virulence, independent of whether the isolates were grown on a plant with or without R6. Thus, it is possible that the fitness cost of R6 virulence occurs regardless of host genotype. In field studies, powdery mildew was suppressed by at least 50% on plants possessing R6 as compared with those without R6 when coinoculated with R6-virulent and avirulent isolates. R6 virulence in P. macularis appears to be race specific and, at this time, imposes a measurable fitness penalty on the fungus. Resistance genes R1 and R2 appear to remain effective against R6-virulent isolates of P. macularis in the U.S. Pacific Northwest.

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Perpetuation of Powdery Mildew Infection and Identification of Erysiphe australiana as the Crape Myrtle Pathogen in Mid-Tennessee

Plant Disease ◽

10.1094/pd-90-1098 ◽

2006 ◽

Vol 90 (8) ◽

pp. 1098-1101 ◽

Cited By ~ 11

Author(s):

Ainong Shi ◽

Margaret T. Mmbaga

Keyword(s):

United States ◽

Powdery Mildew ◽

The United States ◽

Diagnostic Tools ◽

Specific Primers ◽

Standard Polymerase Chain Reaction ◽

Common Lilac ◽

Pcr Products ◽

Powdery Mildew Fungi ◽

Powdery Mildew Pathogen

The fungus Erysiphe lagerstroemiae is commonly known as the powdery mildew pathogen in crape myrtle (Lagerstroemiae indica) in the United States, and Erysiphe australiana is the powdery mildew pathogen reported in Japan, China, and Australia. The teleomorph often used to identify powdery mildew fungi rarely develops in crape myrtle, and in our observations, ascocarps never formed. Our study showed that the crape myrtle pathogen overwintered as mycelia on dormant buds. The internal transcribed spacer (ITS) regions of rDNA and the intervening 5.8S rRNA gene were amplified using standard polymerase chain reaction (PCR) protocols and the universal primer pairs ITS1 and ITS4. PCR products were analyzed by electrophoresis in a 1.5% agarose gel and sequenced, and the ITS PCR product was 666 bp from ITS1/ITS4 and 704 bp from ITS1-F/ITS4. BLAST analysis of the sequence of the PCR products showed identical similarity with E. australiana reported in Japan, China, and Australia. Comparison of ITS sequences with information in the GenBank on other powdery mildew fungi showed a closest alignment (93% similarity) to Erysiphe juglandis that infects walnut. Specific primers for E. australiana were developed and evaluated for use as diagnostic tools. Out of 12 specific primer pairs evaluated, four primer pairs and four double primer pairs were highly specific to E. australiana and did not amplify Erysiphe pulchra of dogwood, Erysiphe syringae of common lilac, Erysiphe circinata of maple, or Phyllactinia guttata of oak. The E. australiana-specific primers amplified 16 samples of crape myrtle powdery mildew collected from diverse locations in mid-Tennessee. These results clearly showed that the crape myrtle powdery mildew in mid-Tennessee was caused by E. australiana. Specific primers reported in this article provide a diagnostic tool and may be used to confirm the identity of crape myrtle powdery mildew pathogen in other areas in the United States and wherever the disease occurs.

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Responding to an Introduced Pathogen: Podosphaera macularis (Hop Powdery Mildew) in the Pacific Northwest

Plant Health Progress ◽

10.1094/php-2003-1113-07-rv ◽

2003 ◽

Vol 4 (1) ◽

pp. 21 ◽

Cited By ~ 10

Author(s):

Walter F. Mahaffee ◽

Carla S. Thomas ◽

William W. Turechek ◽

Cynthia M. Ocamb ◽

Mark E. Nelson ◽

...

Keyword(s):

Powdery Mildew ◽

Pacific Northwest ◽

Management Practices ◽

Risk Index ◽

Lessons Learned ◽

Infection Model ◽

The Pacific ◽

Sphaerotheca Macularis ◽

The Cost ◽

Podosphaera Macularis

Powdery mildew of hop (Humulus lupus L.), which is caused by Podosphaera macularis (formerly Sphaerotheca macularis) was found in the Yakima Valley, WA in 1996 and subsequently spread to the growing regions in Oregon and northern and southern Idaho. To rapidly assist growers in reducing the cost associated with the preventive fungicide program, the Gubler/Thomas grape powdery mildew risk infection model was adapted for hops. In addition, field surveys were utilized to identify other management practices that impacted disease development. Weather networks were established and utilized to deliver daily regional maps indicating the risk index. These maps were posted to the web for daily access. Lessons learned from this experience will be useful in addressing future pathogen introductions. Accepted for publication 28 March 2003. Published 13 November 2003.

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First Report of Powdery Mildew, Caused by an Oidium sp., on Poinsettia in California

Plant Disease ◽

10.1094/pdis.1998.82.1.128a ◽

1998 ◽

Vol 82 (1) ◽

pp. 128-128 ◽

Cited By ~ 3

Author(s):

S. T. Koike ◽

G. S. Saenz

Keyword(s):

United States ◽

Powdery Mildew ◽

Pacific Northwest ◽

The United States ◽

Euphorbia Pulcherrima ◽

American Phytopathological Society ◽

First Report ◽

The Pacific ◽

Uninoculated Control ◽

Alternative Identification

In December 1996 and January 1997, powdery mildew was observed on potted poinsettia (Euphorbia pulcherrima Willd. ex Klotzsch) plants in Monterey County, CA. Mycelia were observed on stems, petioles, mature and immature leaves, and bracts. Severely diseased leaves became twisted and bent and senesced prematurely. The white mycelia were conspicuous, epiphytic, and amphigenous; hyphae measured 4.6 to 6.9 μm in diameter. Growth initially was in patches but eventually became effused. Appressoria were slightly lobed to lobed and sometimes opposite. Conidiophore foot cells were cylindrical, sometimes bent at the base, and slightly flexuous to flexuous. Foot cells measured 30.0 to 46.2 μm × 5.8 to 6.9 μm and were followed by one to two shorter cells. Conidia were cylindrical to slightly doliform and measured 25.4 to 32.3 μm × 11.6 to 18.5 μm. The length-to-width ratios of conidia generally were greater than 2.0. Conidia were produced singly, placing the fungus in the Pseudoidium-type powdery mildew group. Conidia germinated at the ends, and no fibrosin bodies were observed. Cleistothecia were not found. The fungus was identified as an Oidium species. Pathogenicity was demonstrated by gently pressing infected leaves having abundant sporulation onto leaves of potted poinsettia plants (cvs. Freedom Red, Peter Star Marble, and Nutcracker White), incubating the plants in a moist chamber for 48 h, and then maintaining plants in a greenhouse. After 12 to 14 days, powdery mildew colonies developed on the inoculated plants, and the pathogen was morphologically identical to the original isolates. Uninoculated control plants did not develop powdery mildew. This is the first report of powdery mildew on poinsettia in California. This fungus appears similar to Microsphaera euphorbiae but has longer, slightly flexuous foot cells that do not match the description for M. euphorbiae (1,2). An alternative identification would be Erysiphe euphorbiae; however, there are no available mitosporic descriptions for morphological comparisons (1,2). In the United States, powdery mildew of poinsettia previously has been reported in various states in the Pacific Northwest, Midwest, and Northeast. References: (1) U. Braun. Beih. Nova Hedwigia 89:1, 1987. (2) D. F. Farr et al. 1989. Fungi on Plants and Plant Products in the United States. American Phytopathological Society, St. Paul, MN.

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Transcriptome-Derived Amplicon Sequencing Markers Elucidate the U.S. Podosphaera macularis Population Structure Across Feral and Commercial Plantings of Humulus lupulus

Phytopathology ◽

10.1094/phyto-07-20-0299-fi ◽

2020 ◽

pp. PHYTO-07-20-029

Author(s):

William A. Weldon ◽

Brian J. Knaus ◽

Niklaus J. Grünwald ◽

Joshua S. Havill ◽

Mary H. Block ◽

...

Keyword(s):

United States ◽

Pacific Northwest ◽

Mating Type ◽

High Throughput ◽

Plant Pathogens ◽

Amplicon Sequencing ◽

The United States ◽

Humulus Lupulus ◽

Geographical Regions ◽

Podosphaera Macularis

Obligately biotrophic plant pathogens pose challenges in population genetic studies due to their genomic complexities and elaborate culturing requirements with limited biomass. Hop powdery mildew (Podosphaera macularis) is an obligately biotrophic ascomycete that threatens sustainable hop production. P. macularis populations of the Pacific Northwest (PNW) United States differ from those of the Midwest and Northeastern United States, lacking one of two mating types needed for sexual recombination and harboring two strains that are differentially aggressive on the cultivar Cascade and able to overcome the Humulus lupulus R-gene R6 (V6), respectively. To develop a high-throughput marker platform for tracking the flow of genotypes across the United States and internationally, we used an existing transcriptome of diverse P. macularis isolates to design a multiplex of 54 amplicon sequencing markers, validated across a panel of 391 U.S. samples and 123 international samples. The results suggest that P. macularis from U.S. commercial hop yards form one population closely related to P. macularis of the United Kingdom, while P. macularis from U.S. feral hop locations grouped with P. macularis of Eastern Europe. Included in this multiplex was a marker that successfully tracked V6-virulence in 65 of 66 samples with a confirmed V6-phenotype. A new qPCR assay for high-throughput genotyping of P. macularis mating type generated the highest resolution distribution map of P. macularis mating type to date. Together, these genotyping strategies enable the high-throughput and inexpensive tracking of pathogen spread among geographical regions from single-colony samples and provide a roadmap to develop markers for other obligate biotrophs.

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First Report of Powdery Mildew Caused by an Oidium sp. on Torenia fournieri

Plant Disease ◽

10.1094/pdis.1999.83.9.878b ◽

1999 ◽

Vol 83 (9) ◽

pp. 878-878 ◽

Cited By ~ 2

Author(s):

G. E. Holcomb

Keyword(s):

Powdery Mildew ◽

Infected Plant ◽

Hyphal Growth ◽

The United States ◽

Plant Leaves ◽

Torenia Fournieri ◽

Bedding Plant ◽

Leaf Surfaces ◽

Powdery Mildew Fungi ◽

Leaf Distortion

Torenia fournieri Lind. ex Fourn. (wishbone flower, bluewings) is a popular summer bedding plant in Louisiana. Clown Mixture cultivars are available in garden centers in March and April. Transplants of cultivar Clown Rose were purchased, transplanted to larger pots, and maintained in a greenhouse. A powdery mildew was observed on these plants in March and all plants (six) were severely diseased by May. Symptoms included leaf distortion and yellowing. Powdery mildew was not present on transplants and none was found in later checks of garden centers. An Oidium sp. was observed sporulating on both leaf surfaces of infected plants. Conidia were ellipsoid, produced in chains, lacked fibrosin bodies, and averaged 41 × 22 μm in dimensions. No sexual stage was observed. Healthy plants of Clown Mixture cultivars were obtained and inoculated by brushing conidia from infected plant leaves to leaves of healthy plants. Plants were maintained in a greenhouse where temperatures ranged from 16 to 26°C. Hyphal growth appeared on inoculated plants after 5 days and the reproductive structures formed later appeared the same as those on originally infected plants. Uninoculated plants remained healthy. No previous reports of powdery mildew diseases of T. fournieri in the United States were found. Other powdery mildew pathogens reported on T. fournieri are Sphaerotheca fuliginea (Schlechtend.:Fr.) Pollacci in Finland and Japan and an Erysiphe sp. in Japan (1). Reference: (1) K. Amano. Host Range and Geographical Distribution of the Powdery Mildew Fungi. Japan Scientific Press, Tokyo, 1986.

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Hop Powdery Mildew Control Through Alteration of Spring Pruning Practices

Plant Disease ◽

10.1094/pdis-10-15-1127-re ◽

2016 ◽

Vol 100 (8) ◽

pp. 1599-1605 ◽

Cited By ~ 8

Author(s):

Claudia Probst ◽

Mark E. Nelson ◽

Gary G. Grove ◽

Megan C. Twomey ◽

David H. Gent

Keyword(s):

Powdery Mildew ◽

Pacific Northwest ◽

Low Cost ◽

Specific Effect ◽

Disease Development ◽

Quality Factors ◽

Early Season ◽

Yield And Quality ◽

The Pacific ◽

Podosphaera Macularis

Podosphaera macularis, the causal agent of hop powdery mildew, is a recurrent threat to hops in the Pacific Northwest because of the potential to reduce cone yield and quality. Early-season pruning is a common practice in hop production for horticultural reasons. Studies were conducted over a 3-year period in a commercial hop yard to quantify the effect of pruning method and timing on disease development, yield, and cone quality factors. A 4-week delay in pruning reduced the incidence of leaves with powdery mildew from 46 to 10% and cones from 9 to 1%, with the specific effect being season dependent. Pruning using chemical desiccants rather than by mechanical means had similar effects on disease levels on leaves. On cones, though, chemical pruning had a small but significant reduction in the incidence of powdery mildew compared with mechanical pruning. Cone yield, levels of bittering-acids, and color were not negatively affected in any individual year or cumulatively over three seasons when pruning treatments were applied repeatedly to the same plots during the study period. Delayed pruning may offer a low-cost means of reducing both the incidence of powdery mildew and early-season fungicide inputs in certain cultivars.

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Sclerotinia Wilt of Hop (Humulus lupulus) Caused by Sclerotinia sclerotiorum in the Pacific Northwest United States

Plant Disease ◽

10.1094/pdis-12-11-1039 ◽

2012 ◽

Vol 96 (4) ◽

pp. 583-583

Author(s):

S. M. Kropf ◽

M. L. Putnam ◽

M. Serdani ◽

M. C. Twomey ◽

J. L. Woods ◽

...

Keyword(s):

United States ◽

Pacific Northwest ◽

Sclerotinia Sclerotiorum ◽

Pcr Amplification ◽

Soil Surface ◽

The United States ◽

Morphological Characters ◽

Nucleotide Polymorphisms ◽

Foliar Symptoms ◽

The Pacific

In June 2009, wilted hop bines were observed in a yard in Marion County, OR. The wilt was associated with a stem rot that occurred ~1 m from the ground near the point where bines are tied together for horticultural purposes. Samples of affected stems were submitted to the Oregon State University Plant Clinic. White hyphae and large, black sclerotia were present on the stems, with a clear delineation between healthy and diseased tissue. The pathogen was identified as Sclerotinia sclerotiorum based on morphological characters. In June 2011, bine wilting was observed on the same farm but in a different hop yard (cv. Nugget) ~10 km from the 2009 occurrence. Affected plants had upward curled leaves with necrotic margins or wilted bines that were severed at the soil line. Wilted bines tended to have smaller diameters than bines with foliar symptoms only. Of 100 plants examined, 75% displayed some foliar symptoms and 66% had at least one bine that was wilted. Yield loss was estimated at 10 to 20% due to bine wilting before cone development. Unlike the 2009 occurrence, wilted bines did not display aerial signs of S. sclerotiorum. Rather, water-soaked lesions covered in white, cottony mycelium were apparent on affected stems 2.5 to 5 cm below the soil surface, some bearing large, irregularly shaped sclerotia. Isolations made onto potato dextrose agar yielded isolates with rapid growth rates and morphological characters consistent with S. sclerotiorum (1). DNA was extracted (2) and pathogen identity was confirmed by PCR amplification and sequencing of the internal transcribed spacer regions from isolates SS001 and SS002 as described before (4). The amplicons were sequenced bidirectionally and consensus sequences were 100% similar to S. sclerotiorum (GenBank No. AAGT01000678.1). Two nucleotide polymorphisms were present that differentiated the sequences from those of 12 S. trifoliorum accessions in GenBank that could be aligned (2). Greenhouse assays utilizing a toothpick inoculation procedure (3) were conducted to fulfill Koch's postulates. Stems of five 4-week-old hop plants of cv. Agate were pierced with a toothpick colonized with S. sclerotiorum. Five control plants were similarly inoculated with toothpicks without the fungus. Inoculated plants developed symptoms similar to those observed in the field within 11 days; four of five plants inoculated with isolate SS001 and two of five plants inoculated with isolate SS002 completely wilted. S. sclerotiorum was reisolated from all inoculated plants but not the control plants. To our knowledge, this is the first report of Sclerotinia wilt on hop in Oregon or the Pacific Northwest (1), where nearly all commercial hop production occurs in the United States. The disease appears to be localized to a limited number of yards, although given the widespread distribution and host range of S. sclerotiorum, it is plausible that the disease may occur in other yards. Recurrent outbreaks and spread of the disease among yards on the affected farm suggests that Sclerotinia wilt has the potential to become a perennial problem on hop and efforts to limit the introduction of S. sclerotiorum into other yards are warranted. References: (1) D. H. Gent. Page 32 in: Compendium of Hop Diseases and Pests. The American Phytopathological Society, St. Paul, MN, 2009. (2) E. N. Njambere et al. Plant Dis. 92:917, 2008. (3) M. L. Putnam. Plant Pathol. 53:252, 2004. (4) T. J. White et al. PCR Protocols: A Guide to Methods and Applications. Academic Press, San Diego, 1990.

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Characterization of Resistance to Powdery Mildew in the Hop Cultivars Newport and Comet

Plant Health Progress ◽

10.1094/php-br-13-0129 ◽

2014 ◽

Vol 15 (2) ◽

pp. 55-56 ◽

Cited By ~ 8

Author(s):

Sierra N. Wolfenbarger ◽

Emily B. Eck ◽

David H. Gent

Keyword(s):

Powdery Mildew ◽

Pacific Northwest ◽

Host Resistance ◽

Resistant Cultivars ◽

Virulent Strains ◽

The Pacific ◽

Potential Sources ◽

Important Disease ◽

Podosphaera Macularis

Hop powdery mildew, caused by Podosphaera macularis, is an important disease in the Pacific Northwest. Resistant cultivars of hop have been developed and have provided field immunity to the disease until virulent strains of P. macularis emerged. Due to the increase of powdery mildew on formerly resistant cultivars, studies were conducted to characterize potential sources of host resistance. Accepted for publication 5 February 2014. Published 27 March 2014.

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