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ARS Home » Southeast Area » Dawson, Georgia » National Peanut Research Laboratory » Research » Research Project #435624

Research Project: Integration of Traditional Methods and Novel Molecular Strategies for Improving Disease Resistance and Input-use Efficiency in Peanut

Location: National Peanut Research Laboratory

2021 Annual Report

Objective 1. Characterize peanut pathogens, host responses, and host-plant interactions, including diversity of plant invasion and plant health genes, and use genomic and transcriptomic knowledge for discovery and development of novel methods or technologies to control diseases. Objective 2. Identify, characterize, and evaluate peanut genes involved in disease resistance and drought tolerance, including discovery and elucidation of agriculturally-relevant candidate genes, and work with breeders to facilitate implementation into breeding programs. Objective 3. In collaboration with Auburn University partners, develop and release superior peanut cultivars and improved germplasm with disease resistance and input-use efficiency. Objective 4. Determine the physiological mechanisms that link Aspergillus infection with aflatoxin contamination (NP 303, C2, PS 2A). Objective 5. Understanding the pathway in aspergillus invasion (collaborative effort with FVSU utilizing their scanning electron microscope abilities to understand changes in hull structure under varying environmental conditions), determine impact of Laccase enzymes on hull degradation (NP 303, C2, PS 2A). Objective 6. Work with breeders to develop varieties with resistance to aflatoxin (NP 301 C1, PS 1A).

Double strand RNA (dsRNA) that targets aflatoxin synthesis can be used as a therapeutic control of mycotoxins in peanut without genetic transformation. Knowing the genetic makeup of peanut pathogens (Cercospora arachidicola, Cercosporidium personatum, Thecaphora frezzi, Aspergillus (A.) niger, A. flavus and A. parasiticus) allows for a better disease management and longer effectiveness of control. Identification and validation of molecular markers associated with biotic (early and late leaf spot, peanut smut, crown rot disease, mycotoxin producing fungi), and abiotic (drought) stress resistance in wild peanuts and land races will accelerate breeding programs. Analysis and non-GMO manipulation of gene expression, physiology, microRNA expression, and changes in methylation patterns, both of the plant seed as of Aspergillus during the process of infection, can point to resistance genes or other desirable seed response relevant to the control of aflatoxins. Aspergillus Laccase enzymes are a potentially important factor in pathogenicity, studying the activity of these enzymes on hull degradation/penetration in relation to the varying composition of hulls depending on cultivar and maturity, can lead to develop tools to interfere/block the action of laccases and hence seed invasion by Aspergillus, this work will incorporate the assistance of Scientists in our Unit who have expertise in mechanical resistance of hulls. Collaborating with other research projects in our Unit, we will provide the infrastructure support for large scale screening of germplasm to be incorporated in our pre-breeding program.

Progress Report
ARS scientists at Dawson, Georgia studied and published the local production of small ribonucleic acids (RNAs) via RNA interference (RNAi) after exogenous application of nucleic acids to peanut leaves; basic research required to understand RNAi in peanut plants and develop strategies for the control of aflatoxins/Aspergillus fungi. In molecular work ARS scientists at Dawson, Georgia in collaboration with scientists from Uganda and Ethiopia, identified the predominant genotypes of Aspergillus fungi invading peanuts in those countries; essential work to develop technologies for the control of Aspergillus and reduce aflatoxin contamination. ARS scientists at Dawson, Georgia continue surveying hundreds of isolates of Aspergillus niger from Georgia peanuts, fungus that causes crown-rot in peanut, the goal is to determine potential resistance to six common fungicides used in the peanut crop. Molecular work led and performed by ARS scientists at Dawson, Georgia, in collaboration with scientists from Argentina, identified important regions in the peanut genome that confer resistance to smut disease. In 2021, ARS scientists from Dawson,Georgia published molecular markers as well as two peanut germplasm sources of peanut-smut resistance. Peanut smut, though not present in the United States, causes up to 50 % losses in its endemic region. In molecular work ARS scientists at Dawson, Georgia in collaboration with scientists from Brazil, 19 molecular markers demonstrated that the fungus that causes sugarcane “rust” has several genotypes and discovered that some of those genotypes can cause rust disease on cultivars previously considered “resistant”. ARS scientists at Dawson, Georgia also reported that 3 of the molecular markers are sufficient to distinguish those genotypes. This is very useful information for sugarcane growers in the United States, since rust spores can be carried by wind for hundreds of miles. For leaf spot studies, genomic DNAs were isolated from the 45 advanced peanut breeding lines and other varieties. A total of 24 resistance (R) gene candidates that showed differential expression in response to drought stress were analyzed by standard polymerase chain reaction. The experiments were performed by ARS scientists at Dawson, Georgia to evaluate functionality of primers to separate resistant and susceptible genotypes. Polymerase chain reaction products were isolated, cloned and sequenced to verify allelic differences. Genotyping primers were designed based on sequencing results to separate single nucleotide differences of the candidate R-genes. For drought studies, newly developed first generation (F1) crosses were planted in May, 2021 for the evaluation for middle season drought tolerance utilizing environmentally controlled rainout shelters. The experiment is in progress. In addition, ARS scientists planted 50 advanced breeding lines used for leaf spot resistance evaluation in a randomized complete block design with 3 replicates at E.V. Smith Station, Auburn University. No fungicide-treated plots were used to facilitate disease. Phenotypic data were integrated into the PCR screening of candidate R-genes associated to leaf spot resistance. ARS scientists at Dawson, Georgia found that breeding lines AU20-35, AU20-25, AU20-29, AU20-43, and AU20-40 have high resistance to leaf spot. In 2021, 52 advanced breeding lines are being tested in field for leaf spot resistance. In 2020 ARS scientists had screened 18 peanut cultivars of known drought tolerance and sensitivity to determine their drought tolerance mechanisms. Lines 8 and TifNV-High Oleic had drought tolerance, possibly explained by a higher WUE and lower stomatal conductance shown by carbon isotope discrimination. ARS scientists at Dawson, Georgia also identified that lines 50 and 49 had apparently a different drought tolerance mechanism related to an elevated soil water extraction capability (showed by high carbon isotope discrimination) resulting in high photosynthesis, transpiration, and higher yields.

1. Identification of predominant Aspergillus genotypes in Ethiopia and Uganda peanuts. ARS scientists at Dawson, Georgia, identified the main genotypes of Aspergillus fungi colonizing seeds in the peanut growing areas of Uganda and Ethiopia; information that is essential to develop management practices.

2. Effective generation of small RNAs within peanut leaves. ARS scientists at Dawson, Georgia, demonstrated that small RNAs can effectively be produced by exogenous application of double strand nucleic acids to the peanut leaves, also found that double-strand DNA was more active than double-strand RNA in producing those small RNA molecules. This is the first step to understand the effect of topical application of nucleic acids to induce RNA interference in peanut plants.

3. Discovery of multiple genotypes of the sugarcane-rust pathogen. ARS scientists at Dawson, Georgia, in collaboration with scientists from Brazil, identified 4 main groups of genotypes of the fungus that causes sugarcane “rust”, determined that certain fungal genotypes can overcome rust resistance in sugarcane cultivars, and provided 3 molecular markers that can distinguish those genotypes. The molecular markers could be used. ARS scientists at Dawson, Georgia, reported 3 molecular markers that can distinguish those genotypes. This is very useful information for sugarcane growers in the United States since rust spores can be carried by wind for hundreds of miles.

4. Identification of two new drought tolerant peanut lines. Field drought evaluation used by ARS scientists at Dawson, Georgia, have verified that the two newly identified peanut lines were drought tolerant and will be incorporated into breeding program. Potential drought responsive genes were identified by ARS scientists at Dawson, Georgia and will be utilized to develop genetic markers in breeding program.

5. Peanut lines with tolerance to leaf spot. ARS scientists at Dawson, Georgia, have found breeding lines of AU20-35, AU20-25 and AU20-29, AU20-43, and AU20-40 have strong resistance to leaf spot. Line 8 and TifNV-High Oleic showed drought tolerance with different mechanisms.

Review Publications
Dang, P.M., Lamb, M.C., Chen, C.Y. 2021. Association of differentially expressed R-gene candidates with leaf spot resistance in peanut (Arachis hypogaea L.). Molecular Biology Reports.
Zhang, H., Wang, M.L., Dang, P.M., Jiang, T., Zhao, S., Lamb, M.C., Chen, C.Y. 2020. Identification of potential QTLs and genes associated with seed composition traits in peanut (Arachis hypogaea L.) using GWAS and RNA-Seq analysis. Gene. 769:145215.
Wang, X., Yang, X., Feng, Y., Dang, P.M., Wang, W., Graze, R., Clevenger, J., Chu, Y., Ozias-Akins, P., Holbrook Jr, C.C., Chen, C. 2021. Transcriptome profile reveals drought induced genes preferentially expressed in response to water deficit in cultivated peanut (Arachis hypogaea L.). Frontiers in Plant Science. 12:645291.
De Blas, F.J., Bruno, C.I., Arias De Ares, R.S., Ballen-Taborda, C., Mamani, E., Oddino, C., Rosso, M., Costero, B., Bressano, M., Soave, J.H., Soave, S.J., Buteler, M.I., Seijo, G.J., Massa, A.N. 2021. Genetic mapping and QTL analysis for peanut smut resistance. Biomed Central (BMC) Plant Biology. 21:312.
Massa, A.N., Bressano, M., Soave, J.H., Buteler, M.I., Seijo, G., Sobolev, V., Orner, V.A., Oddino, C., Soave, S.J., Faustinelli, P.C., De Blas, F.J., Lamb, M.C., Arias De Ares, R.S. 2021. Genotyping tools and resources to assess peanut germplasm: smut-resistant landraces as a case study. PeerJ.
Power, I.L., Faustinelli, P.C., Orner, V.A., Sobolev, V., Arias De Ares, R.S. 2020. Analysis of small RNA populations generated in peanut leaves after exogenous application of dsRNA and dsDNA targeting aflatoxin synthesis genes. PLoS One. 10:13820.
Urashima, A.S., Mistura, T.F., Sakuno, C., Austin, P.D., Arias De Ares, R.S. 2020. Genetic diversity of Puccinia kuehnii, the causal agent of orange rust of sugarcane, from Brazil. Journal of Phytopathology.
Amos, A., Arias De Ares, R.S., Steven, O., Samuel, T., Joseph, S., Dennis, M., John, A., Stephen, B., Andrew, K. 2020. Genetic diversity of aflatoxin-producing Aspergillus flavus isolated from selected groundnut growing agro-ecological zones of Uganda.. BMC Microbiology. 20:252.
Butts, C.L., Dean, L.L., Hendrix, K., Arias De Ares, R.S., Sorensen, R.B., Lamb, M.C. 2021. Hermetic storage of shelled peanut using the purdue improved crop storage bags. Peanut Science. 48(1):22-32.
Copes, W.E., Ibarra Caballero, J., Babiker, E.M., Stewart, J.E., Orner, V.A., Windham, A.S., Arias De Ares, R.S. 2020. Draft genome assembly of Passalora sequoiae a needle blight pathogen on leyland cypress. BMC Research Notes. 13:505.