Location: Mycology and Nematology Genetic Diversity and Biology Laboratory
2023 Annual Report
Objectives
Objective 1. Develop systematic knowledge for identification and classification of fungi and oomycetes that impact plant health. (NP303, C1, PS1A).
Objective 2. Develop diagnostic assays, genomic data, molecular markers, and specimen images to identify and detect plant pathogens. (NP303, C1, PS1A, PS1B).
Approach
Fungi and fungal-like oomycetes are among the most destructive plant pathogens responsible for diseases of agricultural and natural ecosystems worldwide. Conversely, many other fungi are beneficial to agriculture and the environment, providing potent biological control of destructive insect pests and plant pathogens. Early, rapid and accurate identification of fungal pathogens and beneficials is critical to preventing major disease crises. Emerging, insect, and quarantine-significant pathogenic fungi in historically understudied groups important to U.S. agriculture will be studied using cutting edge molecular technologies to determine species boundaries, phylogenetic relationships and gene regions most useful for diagnostic methods development. The objectives of this project are to (1) develop systematic knowledge for identification and classification of fungi and oomycetes that impact plant health and (2) develop diagnostic assays, genomic data, molecular markers, and specimen images to identify and detect plant pathogens. This project will apply a suite of classical and state-of-the-art systematic approaches towards the common goal of characterizing molecular, morphological, biological, and ecological features of plant and insect pathogens. Expected products of this research include new species descriptions and phylogenetic systems that link evolutionary relationships with key pathogen traits such as adaptation, migration, host range, transmission, and pathogenicity. Resources such as pathogen genome sequences, DNA barcodes, molecular markers, voucher specimens, curated culture resources, high-resolution images and illustrations, diagnostic keys, and other diagnostic materials will provide standardized references to facilitate early and accurate pathogen identification and provide the infrastructure needed for the development of pathogen diagnostic tools. The research completed through this project plan will result in enhanced plant disease management, help reduce the time that plants and plant products spend in quarantine, and support more effective use of biocontrol species.
Progress Report
This project replaces 8042-22000-298-000D “Enhancing Plant Protection through Fungal Systematics,” which ended April 22, 2022. This research is relevant to NP 303, C1, PS1A and PS1B. Progress was made under Objective 1, “Develop systematic knowledge for identification and classification of fungi and oomycetes that impact plant health,” in identifying and describing invasive and emerging plant pathogens and characterizing the genotypes of disease-causing fungi and oomycetes. Downy mildew species were characterized and described, including a new pathogen of coneflowers, a popular ornamental and medicinal plant. Species of Bipolaris, Ceratocystis, Colletotrichum, Curvularia, Diaporthe, Macrophomina, Neopestalotiopsis, and Phyllachora pathogens of grain crops, weedy grasses and various specialty crops were also characterized and illustrated. Endophytic fungi from Lippia alba, a plant widely used by traditional medicine practitioners due to their associated bioactive essential oils, were sequenced and characterized phylogenetically. Methods utilizing FTA paper were developed to avoid having to ship living cultures internationally. The most abundant species belonged to the genera Colletotrichum, Corynespora, Fusarium, and Macrophomina. Seventy-one fungal colonies were isolated from Magnolia, an important ornamental tree. The most representative genera isolated include Coniochaeta, Muyocopron, Nemania, Phyllosticta, Ramularia and Xylaria. Fungal isolates determined to belong to the genus Muyocopron were selected to do more in-depth sequencing and analyses to determine if they constitute a new species. Species in this genus are known to produce secondary metabolites with anti-bacterial, anti-cancer, and anti-inflammatory effects. Data collection and annotation for 12 of 63 genomes of the insect pathogen Beauveria bassiana that are now in revision stage of submission to Genbank were completed. Genome assembly and annotation of insect pathogenic Lecanicillium species were commenced as was screening of Beauveria genomes to develop more informative and robust PCR assays for rapid identification, phylotyping, and mating type diagnosis.
Regarding Objective 2, “Develop diagnostic assays, genomic data, molecular markers, and specimen images to identify and detect plant pathogens,”digital images were produced for taxa mentioned above for use in publications and identification tools. Genome data for approximately 15 species of fungal insect and plant pathogenic fungi were produced for use in diagnostic assays and phylogenetic analyses to better distinguish species. Additionally, a collaborative project on a diagnostic assay for tar spot of corn was finalized and submitted for publication. A total of three species of downy mildews were microscopically imaged: downy mildews of coneflower, redbud (only the third time it’s been found), and Potentilla. In additional Sanger and Illumina sequencing was performed for approximately 40 species of plant pathogens including numerous species listed under Objective 1 for eventual use in identification assays. Accurately identifying and genotyping fungal, plant and insect pathogens will enable better methods of control and tracking of disease outbreaks. This will enhance the development of integrated and sustainable pest management strategies, minimizing the need to apply chemical pesticides. A combination of Illumina and Nanopore sequencing was utilized.
Accomplishments
1. New pathogen of purple coneflower was discovered Downy mildew diseases of ornamentals have recently emerged as important and deadly pathogens. ARS scientists in Beltsville, Maryland, in collaboration with university scientists in Germany recently discovered and described a previously unknown pathogen on coneflower, an important ornamental and medicinal plant. The identification of this downy mildew pathogen is also the first step towards developing control measures that can be specifically targeted against the pathogen and prevent millions of dollars of damage to nursery crop growers.
2. The mango sudden decline disease pathogen was characterized with genome data. Mango sudden decline is a potentially deadly disease of mango, which is spread by a bark beetle. While the beetle is known to be present area where mango is grown in the U.S., the disease has not yet been observed. ARS scientists in Beltsville, Maryland, in collaboration with other USDA APHIS scientists, in Beltsville, Maryland, sequenced the genomes of several isolates of the pathogen to determine how to distinguish the species implicated in causing this disease, discovering that multiple species may be involved. This information will be used by quarantine and extension personnel to identify additional species that may have the potential to cause this disease and to prevent its spread.
3. Phylogenomics of the insect biocontrol fungus Beauveria bassiana. The soil-dwelling insect biocontrol fungus Beauveria bassiana is used for the control of a wide range of pest insect species of significance to agriculture, horticulture, domestic animals, and humans. Prior genetic surveys indicate that Beauveria bassiana represents an extensive cryptic species complex whose individual species and isolates vary extensively in virulence with respect to different insect host species. To date, a comprehensive phylogeny of this complex has not been available because generic molecular markers for fungi cannot resolve this species’ complex recent diversification. ARS scientists in Beltsville, Maryland, and Ithaca, New York, have conducted comprehensive comparisons of nearly 100 Beauveria genomes to identify hundreds of conserved single-copy genes that are informative to the evolution of this group that together yield a highly resolved and well supported evolutionary tree of the species complex.
Review Publications
Amaral, A.G., Duarte, I.G., Vieira, W.A., Castlebury, L.A., Camara, M.P. 2023. Colletotrichum karsti causing anthracnose on Dioscorea cayennensis in Brazil. Journal of Plant Pathology. 105:381-382. https://doi.org/10.1007/s42161-022-01298-6.
Crous, P.W., Sandoval-Denis, W., Costa, M.M., Groenewald, J.Z., Van Iperen, A., Starink-Willemse, M., Hernández-Restrepo, M., Salgado-Salazar, C. 2022. Fusarium and allied fusarioid taxa (FUSA). 1. Fungal Systematics and Evolution. 9(1):161-200. https://doi.org/10.3114/fuse.2022.09.08
Duarte, I.G., Veloso, J., Amaral, A.G., Da Silva, A.C., Silva, H.R., Balbino, V.D., Vieira, W.D., Castlebury, L.A., Câmara, M.S. 2022. Colletotrichum siamense causing anthracnose on Etlingera elatior. Crop Protection. 162(1). Article e106092. https://doi.org/10.1016/j.cropro.2022.106092.
Salgado-Salazar, C., Thines, M. 2022. Two new species of plasmopara affecting wild grapes in the United States. Mycological Progress. 21:63.
Geiser, D.M., Al-Hatmi, A., Aoki, T., Arie, T., Balmas, V., Barnes, I., Bergstrom, G.C., Bhattacharyya, M.K., Bloomquist, C.L., Bowden, R.L., Brankovics, B., Brown, D.W., Burgess, L.W., Bushley, K., Busman, M., Cano-Lira, J.F., Carillo, J.D., Chang, H., Chen, C., Chen, W., Chilvers, M., Chulze, S., Coleman, J.J., Cuomo, C.A., De Beer, W., De Hoog, S., Del Castillo-Munera, J., Del Ponte, E.M., Dieguez-Uribeondo, J., Di Pietro, A., Edel-Hermann, V., Elmer, W.H., Epstein, L., Eskalen, A., Esposto, M., Everts, K.L., Fernandez-Pavia, S.P., Ferreira Da Silva, G., Foroud, N.A., Fourie, G., Frandsen, R., Freeman, S., Freitag, M., Frenkel, O., Fuller, K.K., Gagkaeva, T., Gardiner, D.M., Glenn, A.E., Gold, S.E., Gordon, T., Gryzenhout, M., Guarro, J., Gugino, B.K., Gutierrez, S., Kim, H., Kistler, H.C., Lofton, L., Laraba, I., Martin, F.N., Mccormick, S.P., O Donnell, K., Proctor, R., Rehner, S.A., Vaughan, M.M., Ward, T.J. 2021. A monophyletic Fusarium that includes the Fusarium solani species complex is strongly supported by a 19-gene phylogenomic analysis. Phytopathology. https://doi.org/10.1094/PHYTO-08-20-0330-LE.
Matos Franco, G., Chen, Y., Doyle, V.P., Rehner, S.A., Diaz, R. 2022. Mortality of the crapemyrtle bark scale (Hemiptera: Eriococcidae) by commercial biopesticides under greenhouse and field conditions. Biological Control. 175:105061.
Vieira, W.D., Duarte, I.G., Lima, A.F., Amaral, A.G., Da Silva, A.C., Doyle, V.P., Castlebury, L.A., Camara, M.P. 2022. Elucidating the Colletotrichum spp. diversity responsible for papaya anthracnose in Brazil. Fungal Biology. 126(9). https://doi.org/10.1016/j.funbio.2022.08.001.
Fernandez, H.S., Manamgoda, D.S., Udayanga, D., Deshappriyaa, N., Munasinghea, M.S., Castlebury, L.A. 2022. Molecular phylogeny and morphology reveal two new graminicolous species, Bipolaris adikaramae sp. nov and B. petchii sp. nov., with new records of fungi from cultivated rice and weedy grass hosts. Mycological Progress. 21:59. https://doi.org/10.1007/s11557-022-01809-w.
Stalpers, J.A., Redhead, S.A., Rossman, A.Y., Crouch, J.A., Cubeta, M.A., Kirschner, R., Langer, G.J., Larsson, K.H., Mack, J., Marvanová, L., May, T.W., Norvell, L.L., Oberwinkler, F., Papp, V., Roberts, P., Rajchenberg, M., Seifert, K.A., Thorn, G. 2021. Competing sexual-asexual generic names in the Agaricomycotina (Basidiomycota), with recommendations for use. IMA Fungus. 12:22. https://doi.org/10.1186/s43008-021-00061-3.
Matos Franco, G., Chen, Y., Doyle, V.P., Rehner, S.A., Diaz, R. 2022. Will the application of biocontrol fungi disrupt predation of Acanthococcus lagerstroemiae by coccinellids? Journal of Invertebrate Pathology. 193:107789. https://doi.org/10.1016/j.jip.2022.107789.
LeBlanc, N.R., Salgado-Salazar, C. 2023. Genome sequence resources for Verticillium dahliae-inhibiting Streptomyces spp. isolated from agricultural soil. PhytoFrontiers. https://doi.org/10.1094/PHYTOFR-10-22-0116-A.
Rehner, S.A., Gazis, R., Doyle, V.P., Vieira, W.A., Campos, P.M., Shao, J.Y. 2023. Genomic analyses of the Colletotrichum gloeosporioides species complex: 13 tree endophytes from the Neotropics and Paleotropics. Microbiology Resource Announcements. 12(1). https://doi.org/10.1128/mra.01040-22.
Queiroz, C.A., Caniato, F.F., Siqueira, V.K.S., de Moraes Catarino, A., Hanada, R.E., O'Donnell, K., Laraba, I., Rehner, S.A., Sousa, N.R., Silva, G.F. 2023. Population genetic analysis of Fusarium decemcellulare, a guaraná pathogen, reveals high genetic diversity in the Amazonas state, Brazil. Plant Disease. 107(5):1343-1354. https://doi.org/10.1094/PDIS-01-22-0083-RE.
Salgado-Salazar, C., Catstroagudin, V., Leblanc, N.R., Daughtrey, M., Hausbeck, M., Palmer, C., Shishkoff, N., Warfield, C.Y., Crouch, J. 2023. The impatiens downy mildew epidemic in the U.S. is caused by new, introgressed lineages of Plasmopara destructor with prominent genotypic heterogenicity and high evolutionary potential. Plant Disease. https://doi.org/10.1094/PDIS-08-22-1872-RE.
Groth-Helms, D., Rivera, Y., Martin, F.N., Arif, M., Sharma, P., Castlebury, L.A. 2023. Terminology and guidelines for diagnostic assay development and validation: Best practices for molecular tests. PhytoFrontiers. 3(1):23-35. https://doi.org/10.1094/PHYTOFR-05-22-0059-FI.
