Research Project: Molecular and Genetic Approaches to Manage Cotton and Sorghum Diseases

Location: Insect Control and Cotton Disease Research

2023 Annual Report

Objectives
Objective 1. Develop molecular tools to rapidly identify and detect presence of cotton boll rot pathogens in plants and insects. (NP303, C1, PS1A, PS1B) Objective 2. Improve the understanding of insect vector/pathogen interactions and pathogen reservoirs to reduce the spread of cotton diseases. (NP303, C2, PS2C, PS2D) Objective 3. Identify fungal and host genes that can be targeted for disease suppression, and develop cotton germplasm resistant to FOV and nematodes. (NP303, C2, PS2A, PS2B; C3, PS3A, PS3B) Objective 4. Evaluate national and international sorghum germplasm resources, and determine the inheritance and allelic relationships of host plant resistance to anthracnose, grain mold, head smut, and downy mildew diseases. (NP303, C3, PS3A; C2, PS2A) Objective 5. Determine the population structure of diverse anthracnose (Colletotrichum sublineolum) and head smut (Sporisorium reilianum) isolates and phenotypic expression patterns of virulence on host differentials. (NP303, C2, PS2A)

Approach
Boll rots, nematodes, and fungal pathogens continue to pose the greatest threat to U.S. cotton production. To address boll rots, we previously sequenced the genome of several bacterial boll rot pathogens vectored by cotton fleahoppers, stink bugs, and verde plant bugs to identify pathogenicity genes involved in boll rot disease. In this project, we will confirm whether Lygus spp. and stink bugs vector boll rot and Fusarium wilt pathogens, respectively, and identify pathogenicity genes common to all bacterial boll rot pathogens with the long-term goal of developing a PCR-based kit that can be used in the field to rapidly detect insects harboring pathogens. The ability to rapidly detect boll rot pathogens within insects and plants, along with an improved understanding of insect vector and pathogen interactions, are both critically needed to develop sound management strategies for cotton diseases and respective insect vectors. A strain of Fusarium oxysporum f. sp. vasinfectum, known as race 4 (FOV4), was recently detected in several cotton fields in West Texas and New Mexico. This strain was initially detected and confined to cotton fields in California. Unlike other FOVs, race 4 does not require the presence of nematodes to cause severe disease of plants; however, the interaction between root knot nematodes and FOV4 pathogenicity remains unclear. Spread of FOV4 in the United States could be catastrophic because there are currently no feasible control options once this pathogen is established in a field. The development of resistant germplasms is widely deemed the most practical and long-term solution for managing this disease. To this end, we will identify and test sources of resistance to FOV4 and develop respective markers which will be incorporated into previously developed nematode-resistant germplasm to facilitate our breeding efforts to produce cotton lines that are resistant to both FOV4 and nematodes.

Progress Report
Work in FY 2023 provided a clearer understanding of the dynamics of two major cotton diseases in the U.S., boll rot and Fusarium wilt (Fov). Sorghum research led to the discovery of several lines that are resistant to anthracnose, grain mold, head smut, and downy mildew diseases. In work addressing Objective 1, tarnished plant bugs were collected from cotton fields in Texas as well as from fields in Mississippi that showed signs of boll rot; work is underway to determine if these insects harbor boll rot pathogens. Research addressing Objective 2 led to the development of several genetically near-identical (isogenic) cotton lines that are resistant or susceptible to the fungal disease, Fov, caused by race 4 (Fov4). The development of these isogenic sister lines that differ in their susceptibility to Fov4 is critical for establishing the underlying mechanism(s) of Fov4 resistance in cotton and might provide potential sources of Fov4 resistance that can be incorporated in breeding programs. Work under Objective 3 identified several cotton germplasm lines that showed resistance to Fov4 under controlled environmental conditions; the most promising candidates were propagated to increase seed production for field evaluations. In research addressing Objective 4, hundreds of sorghum lines from national and international germplasm resources were screened for resistance to anthracnose, grain mold, head smut, and downy mildew diseases. Based on evaluations in greenhouses, several lines appeared to be highly resistant to one or more of these fungal diseases. Some of these resistant lines were crossed with susceptible lines to provide insight on the inheritance patterns of resistance; this information will be critical in developing new disease-resistant sorghum varieties for use by U.S. farmers. Work under Objective 5 led to the collection and identification of eight anthracnose and head smut isolates with varying levels of virulence; the work revealed the variability and associated phenotypic expression patterns of virulence among anthracnose and head smut isolates. Recognition of this variability will be critical in identifying and evaluating sorghum lines that are resistant to these fungal pathogens.

Accomplishments
1. Registration of cotton germplasm lines resistant to plant-parasitic nematodes. Among cotton pests, plant-parasitic nematodes (e.g., reniform and root knot nematodes) are responsible for the greatest yield losses in cotton. These nematodes can also interact synergistically with root rot pathogens such as Fusarium oxysporum f. sp. vasinfectum (Fov) to exacerbate disease severity. Although repeated applications of pesticides, along with crop rotation, can reduce nematode damage, the development and use of nematode-resistant cultivars is widely considered to be the most effective and economical approach for managing nematodes in cotton. ARS researchers at College Station, Texas, in collaboration with Cotton Incorporated and academic colleagues, developed and registered eight germplasm lines of Upland cotton that are resistant to reniform nematodes and that possess superior performance traits. Four of the lines also possess one or two genes that provide resistance to root-knot nematodes. The registration and availability of these lines will greatly facilitate work by breeders to develop new commercial varieties of Upland cotton that are resistant to nematodes and that will concurrently provide resistance against strains of Fov that require the presence of nematodes for infection.

2. Sorghum lines resistant to fungal diseases. Fungal diseases such as anthracnose, grain mold, head smut, and downy mildew reduce global sorghum yields and seed quality by hundreds of millions of dollars on an annual basis. The development of new disease-resistant sorghum varieties is the most economical and effective method for controlling these fungal diseases. ARS researchers at College Station, Texas, and Mayaguez, Puerto Rico, in collaboration with academic colleagues, screened several hundred sorghum lines from national and international sources for resistance to these fungal diseases. The research identified several lines that are highly resistant to one or more of the diseases, and work is underway to evaluate these lines under field conditions. This accomplishment is significant because it provides sorghum breeders and researchers with critical new germplasm sources that will be foundational in developing new, disease-resistant sorghum varieties.

Review Publications
Prom, L.K. 2023. Frequency of isolation of four fungal species colonizing sorghum grain collected from six lines in an anthracnose-infected field. Journal of Agriculture and Crops. 9(1):137-140. https://doi.org/10.32861/jac.91.137.140.
Prom, L.K., Cuevas, H.E., Ahn, E.J., Isakeit, T. 2022. Evaluation of a subset of Ethiopia sorghum collection germplasm from the National Genetic Resources Program of the United States Department of Agriculture for anthracnose resistance. American Journal of Plant Sciences. 13:1403-1411. https://doi.org/10.4236/ajps.2022.1312095.
Prom, L.K., Medrano, E.G., Liu, J. 2023. In vitro antagonistic action by Bacillus velezensis strain LP16S against cotton wilt pathogens. Journal of Agriculture and Crops. 9(3):372-375. https://doi.org/10.32861/jac.93.372.375.
Prom, L.K., Haougui, A., Ali, B.O., Karimou, I., Abdoulaye, A., Oumarou, O.H., Basso, A., Fall, C., Magill, C. 2023. Incidence, severity, and prevalence of sorghum diseases in the major production regions in Niger. Journal of Plant Studies. 12(1). Article 48. https://doi.org/10.5539/jps.v12n1p48.
Ahn, E.J., Prom, L.K., Magill, C.W. 2023. Multi-trait genome-wide association studies of sorghum bicolor regarding resistance to anthracnose, downy mildew, grain mold and head smut. Pathogens. 12(6). Article 779. https://doi.org/10.3390/pathogens12060779.
Ahn, E.J., Fall, C., Botkin, J., Curtin, S.J., Prom, L.K., Magill, C.W. 2023. Inoculation and screening methods for major sorghum diseases caused by fungal pathogens: Claviceps africana, Colletotrichum sublineola, Sporisorium reilianum, Peronosclerospora sorghi and Macrophomina phaseolina. Plants. 12(9). Article 1906. https://doi.org/10.3390/plants12091906.
Ahn, E.J., Botkin, J., Ellur, V., Lee, Y., Poudel, K., Prom, L.K., Magill, C.W. 2023. Genome-wide association study of seed morphology traits in Senegalese sorghum cultivars. Plants. 12(12). Article 2344. https://doi.org/10.3390/plants12122344.
Wagner, T.A., Bell, A.A., Castles, Z.A., Ali, A., Flores, O., Liu, J. 2022. Detection, genotyping, and virulence characterization of Fusarium wilt race 4 (VCG0114) causing cotton wilt in three Texas fields. Journal of Phytopathology. 170(7-8):492-503. https://doi.org/10.1111/jph.13100.
Medrano, E.G., Glover, J.P., Bell, A.A., Brewer, M.J. 2021. A Serratia marcesens strains involved in cotton (Gossypium hirsutum) boll infection by a prokaryote. Current Microbiology. 12:1565-1578. https://doi.org/10.4236/as.2021.1212100.
Wagner, T.A., Davie, S.S., Mcgill, C., Liu, J. 2022. Interaction of Fusarium wilt race 4 with root-knot nematode increases disease severity in cotton. Plant Disease. https://10.1094/pdis-12-21-2725-sc.
Zhang, L., Chen, J., Liu, J., Sun, Q., Liu, J., Li, H., Wang, P., Chu, Z., Zhang, X., Yuan, Y., Shi, Y., Cai, Y. 2022. lncRNA7 and lncRNA2 modulate cell wall defense genes to regulate cotton resistance to Verticillium wilt. Plant Physiology. Article kiac041. https://doi.org/10.1093/plphys/kiac041.
Medrano, E.G., Haydel, S.E. 2021. Complete genome sequence of the methicillin-resistant Staphylococcus aureus strain SQL1/USA300, used for testing the antimicrobial properties of clay phyllosilicates and customized aluminosilicates. Microbiology Resource Announcements. 10(45). Article e00861-21. https://doi.org/10.1128/MRA.00861-21.
Mays, T.D., Medrano, E.G., Mamoudou, S., Schuster, G.L. 2022. Bacillus thuringiensis derived vegetative insecticidal protein Vip3Aa20 as a potential aflatoxin mitigation tool in maize (Zea mays). Research Journal of Plant Pathology. 5(2:05). https://doi.org/10.36648/iprjpp.22.5.005.
Cuevas, H.E., Cruet-Burgos, C.M., Prom, L.K., Knoll, J.E., Stutts, L.R., Vermerris, W. 2021. The inheritance of anthracnose (Colletotrichum sublineola) resistance response in sorghum differential lines QL3 and IS18760. Scientific Reports. 11. Article 20525. https://doi.org/10.1038/s41598-021-99994-3.
Bell, A.A., Robinson, F., Quintana, J., Hinze, L.L., Harris, J.N., Liu, J., Wagner, T.A., Prom, S., Saladino, V., Zheng, X., Stelly, D., Nichols, R. 2023. Registration of eight germplasm lines of upland cotton resistant to nematodes with elite agronomic performance. Journal of Plant Registrations. 17:536-543. https://doi.org/10.1002/plr2.20290.
Liu, J., Wagner, T. 2021. Detection and genotyping of Fov4 (race 4, VCG0114), the fusarium wilt pathogen of cotton. In: Coleman, J., editor. Fusarium Wilt: Methods and Protocols, Methods in Molecular Biology. Switzerland: Springer Nature. 2391:191-205. https://doi.org/10.1007/978-1-0716-1795-3_16.
Wagner, T.A., Bell, A.A., Puckhaber, L.S., Magill, C., Liu, J. 2023. Effect of RNAi suppression of the gossypol pathway on resistance to necrotrophic fungal seedling disease pathogens in cotton. Journal of Phytopathology. 127. Article 102085. https://doi.org/10.1016/j.pmpp.2023.102085.