Research Project: Genetic Improvement of Maize and Sorghum for Resistance to Biotic and Abiotic Stresses

Location: Crop Genetics and Breeding Research

2022 Annual Report

Objectives
1. Identify, develop, and release Southeast-adapted maize germplasm with reduced aflatoxin accumulation and resistance to key insect pests. 1A. Evaluate exotic maize germplasm from the Germplasm Enhancement of Maize (GEM) program, International Center for the Improvement of Maize and Wheat (CIMMYT), and the U.S. maize germplasm collection for reduced aflatoxin contamination. 1B. Screen for resistance to ear- and kernel-feeding insects in maize germplasm from the GEM, the CIMMYT, and the U.S. maize germplasm collection. 1C. Develop maize germplasm with reduced aflatoxin accumulation, increased resistance to insects, and enhanced agronomic performance in the southeastern Coastal Plain region. 2. Identify, develop, and release new sorghum germplasm with Southeast-adapted maturity genes and greater resistance to the sugarcane aphid, other key insects, and diseases. 2A. Evaluate sorghum lines from the U.S. germplasm collection for anthracnose resistance. 2B. Screen for foliar-feeding sugarcane aphid and fall armyworm and kernel-feeding sorghum midge resistance in sorghum lines from the U.S. germplasm collection. 2C. Develop sorghum germplasm with improved disease and insect resistance and high yield potential. 3. Develop molecular markers for reduced aflatoxin accumulation, and resistance to insects in maize and resistance to insects and foliar diseases in sorghum, and utilize molecular markers for gene identification and cultivar development. 3A. Develop molecular markers for reduced aflatoxin accumulation, and resistance to insects in maize, and utilize molecular markers for gene identification and cultivar development. 3B. Develop molecular markers for resistance to key insects and foliar diseases in sorghum, and utilize molecular markers for gene identification and cultivar improvement.

Approach
Objective 1: Exotic maize germplasm from the Germplasm Enhancement of Maize (GEM) Program, the International Maize and Wheat Improvement Center (CIMMYT), Mexico, and the U.S. maize germplasm collection will be screened for resistance to multiple insects and diseases, and reduced aflatoxin contamination under the southern climate. Equal priority will be given to the GEM and exotic germplasm, since the GEM germplasm will likely have better agronomic traits while the exotic germplasm may offer better resistance/tolerance to biotic and abiotic stress factors. Such a combination would allow us to develop new germplasm with good yield potential and resistant to multiple insects, diseases, and reduced mycotoxin contaminations. To effectively serve the seed industries, the screenings of maize insect pests will focus on key foliar-, ear- and kernel-feeding insects, in particular, fall armyworm, corn earworm and maize weevil. The genetic and biochemical bases for the biotic stress resistance in these newly identified germplasm lines will be further examined. New maize breeding crosses will be made by recombining germplasm with superior agronomic traits with the newly identified germplasm that confers multiple pest resistance and with reduced mycotoxin contamination. New maize germplasm will be developed by continuously screening and continuous self-pollination of the segregating populations. At the same time, maize recombinant inbred lines (RILs) will also be developed to identify DNA markers for multiple pest resistance. Objective 2: A similar approach is utilized for the screening of sorghum germplasm for resistance to multiple biotic stress factors. Previously identified disease resistant and with agronomically-elite germplasm (with Ex-PVP program) in the U.S. germplasm collection will be screened for resistance to sugarcane aphid, fall armyworm, foliar anthracnose disease, and sorghum midge. The genetic and biochemical bases for insect and disease resistance will be examined. The roles of the secondary metabolites to biotic stress resistance in sorghum will also be examined. New sorghum breeding crosses will also be made using the newly identified sorghum germplasm lines that are resistant to multiple biotic stresses and with good yield potential. The breeding crosses will be continuously screened and selected, and self-pollinated to develop and release new sorghum germplasm lines (B lines, or maintainer lines). The best B lines will also be converted into A lines (or cytoplasmic-nuclear male sterile lines) to serve the seed industries. At the same time, sorghum RIL populations will be developed to identify DNA markers for multiple biotic stress resistance at vegetative and reproductive growth stages, respectively. Objective 3: Development of molecular markers for reduced aflatoxin accumulation, and resistance to multiple pests in maize and sorghum will utilize the newly developed genetic resources (i.e., breeding crosses, RIL populations, and new germplasm lines) described in Objective 1 and Objective 2, respectively. The marker development will be performed by working closely with our collaborators and confirmed in multiple locations.

Progress Report
Research activities have been continuously focused on the genetic improvement of maize and sorghum for yield related traits, resistance to Aspergillus flavus infection and aflatoxin accumulation, and damage caused by disease and insect pests. For maize breeding efforts, inbreds and hybrids from Germplasm Enhancement of Maize (GEM) program from North Carolina and Iowa, International Maize and Wheat Improvement Center (CIMMYT) and U.S. national germplasm center (GRIN database) were evaluated for resistance to whorl and ear feeding insects, and reduced aflatoxin accumulation. In addition, African inbred lines, Ex-PVP and other maize germplasm lines (such as from the GEM program and collaborators) are being evaluated continuously for foliar and ear-feeding insect resistance. New breeding crosses have been made continuously and advanced for new maize germplasm development by screening for insect and disease resistance, low aflatoxin accumulation, heat and drought tolerance, and good agronomic traits (e.g., no lodging and good yield potential). In 2021, 10 sets of breeding crosses at various stages (F1 to F6) and recombinant inbred line (RIL) populations were screened continuously for new germplasm development with insect resistance and reduced aflatoxin accumulation. Eighty-six new hybrid testcrosses were evaluated in a four-replication randomized complete block design (RCBD) for both yield and aflatoxin in 2021. Also in 2021 another set of 124 new hybrids (new ARS lines x Ex-PVP testers) was also evaluated for yield and aflatoxin in a four-replicate RCBD experiment. However, aflatoxin could not be evaluated due to lab occupancy restrictions. In multidisciplinary collaborative/cooperative research efforts in our region, ARS researchers in Tifton, Georgia, continue to participate in the Southeastern Regional Aflatoxin Trial (SERAT) with two separate trials with 50 entries each for yield and aflatoxin accumulation, respectively. Six hybrids from our team were entered for the trial in 2021 and our location again served as a test location for yield and aflatoxin trials with 50 entries each. ARS researchers in Tifton, Georgia, also work with other scientists from the Aflatoxin Mitigation Center of Excellence (AMCOE) project by continuously screening a set of accessions (derived from 4-way and 8-way crosses with low levels of aflatoxin accumulation from Texas A&M) for fall armyworm resistance and superior agronomic traits at Tifton, Georgia. In our breeding nursery of 2021, ARS researchers advanced 114 S5-S6 selections by selecting for superior agronomic traits with fall armyworm resistance, low aflatoxin accumulation, and superior agronomic traits. Approximately 500 additional selections in various stages of development (F1 – F6 and BC1) were advanced in 2021. In addition, a new set of 45 maize inbred lines from Africa with known early maturity, drought and low-nitrogen tolerance was also screened for insect resistance and aflatoxin accumulation. ARS researchers in Tifton, Georgia, are participating in the Genomes to Fields (G2F) project. This is a multi-location G x E study involving scientists from ARS and universities from across the U.S. and Canada. Each year ARS researchers have planted approx. 500 hybrid plots at our location, and recorded yield and other phenotypic data, which are shared with the G2F collaborators. A WatchDog mini weather station is used to log in-field weather conditions, and these data are also shared with the G2F team. The first eight years (2014-2021) of genotypic, phenotypic, and weather data from this project are now available to the public through DOI weblinks, as well as inbred ear images from the first two years. For sorghum research, sugarcane aphid outbreaks caused severe yield and economic losses which requires us to focus on providing growers with short- and long-term solutions for managing this invasive pest. ARS researchers in Tifton, Georgia, have completed a five-year Area-Wide IPM project (2016-2021) for sugarcane aphid management, in addition to participating in the State Variety Test every year. In 2021, 15 grain, 28 silage, and 15 forage sorghum hybrids were evaluated and published in the State Variety Trial Annual Report, which provided growers with short-term solutions on aphid management by identifying the best sorghum hybrids in our region. For a long-term solution on managing the current outbreak, ARS researchers are continuously working on new germplasm screening and development with all types of sorghum (including grain, sweet, forage, silage and bioenergy sorghum). A total of 296 mostly grain sorghum germplasm lines from the sorghum association panel (SAP) were screened for anthracnose in 2017 and for sugarcane aphid in 2019 and 2020. This SAP experiment was also used in developing a robot-based phenotyping platform in collaboration with Fort Valley State University and Iowa State University. Drone-based imagery was also captured several times throughout the summer in collaboration with ARS scientists from the Southeast Watershed Research Laboratory in Tifton, Georgia. In 2021, 475 mostly sweet sorghum selections were advanced. Fifty-nine F6 sweet sorghum lines were evaluated for sugarcane aphid response and other traits. Sugarcane aphid infestation was unusually light in 2021, but some inferior F6 lines were able to be discarded based on lodging and other traits. Three new sugarcane aphid resistant sweet sorghum lines have been developed and released.

Accomplishments
1. Identification of a novel quantitative trait locus (QTL) for sugarcane aphid resistance in sorghum line SC112-14. Since its emergence in 2013, the sugarcane aphid has become the major insect pest of sorghum in North America and the Caribbean. In 2018 and 2019, ARS scientists at Tifton, Georgia, evaluated 104 recombinant inbred lines from the cross between two sorghum inbred lines, PI 602951 and SC112-14, in replicated experiments in naturally infested fields in Tifton, Georgia. SC112 is highly resistant to sugarcane aphid, whereas PI 609251 is susceptible. This same population was used previously to map a quantitative trait locus for resistance to the fungal disease anthracnose on Chromosome 5. In cooperation with an ARS scientist at Mayaguez, Puerto Rico, a major locus for sugarcane aphid resistance was found on Chromosome 6. A closer analysis of single nucleotide polymorphism markers showed that this quantitative trait locus is different from the previously reported locus RMES1 that confers insect resistance on the same chromosome. Markers linked to this quantitative trait locus will be useful for breeding robust sugarcane aphid resistance into elite sorghum lines.

2. Identification of markers for sugarcane aphid resistance in the Sorghum Association Panel. Relatively few markers for sugarcane aphid resistance have been identified in sorghum. In 2019 and 2020, ARS scientists, in cooperation with scientists at Fort Valley State University, evaluated sugarcane aphid damage in a replicated field study at Tifton, Georgia. The study included a set of 283 diverse sorghum varieties, and most of them were from the Sorghum Association Panel, which had been widely used for comprehensive genetic diversity studies of grain sorghum. Drone-based data were also collected in the 2020 experiment. Genome-wide association study identified markers on Chromosomes 8 and 10 for aphid resistance that had not been previously reported. Additional markers were found to be associated with drone-based reflectance traits, and three new highly resistant sorghum lines were identified for further study. These new markers will inform future efforts to breed sugarcane aphid resistant sorghum. This research was funded by an incoming agreement with FVSU, “Developing High-throughput Phenotyping Capacity at Fort Valley State University for Genetic Enhancement of Sugarcane Aphid Resistance in Sorghum,” and a NACA with the Area-Wide Pest Management Project for sugarcane aphid management.

Review Publications
Wang, S., Zhang, M., Huang, J., Li, L., Huang, K., Zhang, Y., Li, Y., Ni, X., Deng, Z., Li, X. 2021. Inductive and synergistic interactions between plant allelochemical flavone and Bt toxin Cry1Ac in Helicoverpa armigera. Insect Science. 28:1756-1765. https://doi.org/10.1111/1744-7917.12897.
Poosapati, S., Poretsky, E., Dressano, K., Ruiz, M., Vazquez, A., Sandoval, E., Estrada-Cardenas, A., Duggal, S., Lim, J., Morris, G., Szczepaniec, A., Walse, S.S., Ni, X., Schmelz, E.A., Huffaker, A. 2022. A sorghum genome-wide association study (GWAS) identifies a WRKY transcription factor as a candidate gene underlying sugarcane aphid (Melanaphis sacchari) resistance. Planta. 255. Article 37. https://doi.org/10.1007/s00425-021-03814-x.
Niu, Y., Oyediran, I., Yu, W., Lin, S., Dimase, M., Brown, S., Reay-Jones, F.P., Cook, D., Reisig, D., Thrash, B., Ni, X., Paula-Moraes, S.V., Zhang, Y., Chen, J., Wen, Z., Huang, F. 2021. Populations of Helicoverpa zea (Boddie) in the southeastern United States are commonly resistant to Cry1Ab, but still susceptible to Vip3Aa20 expressed in MIR 162 corn. Toxins. 13(1):63. https://doi.org/10.3390/toxins13010063.
Deng, Z., Leyao, L., Zhang, Y., Zhang, Y., Xie, X., Zhang, M., Huang, J., Ni, X., Li, X. 2021. Identification and characterization of the masculinizing function of the Helicoverpa armigera masc gene. International Journal of Molecular Sciences. 22(16):8650. https://doi.org/10.3390/ijms22168650.
Li, S., Chen, S., Dong, S., Zhang, M., Deng, Z., Ni, X., Huang, J., Li, X. 2021. Spontaneous transposition of HzSINE1 into CYP321A2 is undetectable in the field populations of Helicoverpa zea. Journal of Asia-Pacific Entomology. 24:882-888. https://doi.org/10.1016/j.aspen.2021.07.015.
Li, L., Wang, S., Huang, K., Zhang, Y., Li, Y., Zhang, M., Huang, J., Deng, Z., Ni, X., Li, X. 2021. Identification and characterization of MicroRNAs in gonads of Helicoverpa armigera (Lepidoptera: Noctuidae). Insects. 12:749. https://doi.org/10.3390/insects12080749.
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.
Yao, L., Beibei, J., Yoon, S.C., Zhuang, H., Ni, X., Guo, B., Gold, S.E., Fountain, J.C., Glenn, A.E., Lawrence, K.C., Zhang, H., Guo, X., Zhang, F., Wang, W. 2022. Spatio-temporal patterns of Aspergillus flavus infection and aflatoxin B1 biosynthesis on maize kernels probed by SWIR hyperspectral imaging and synchrotron FTIR microspectroscopy. Food Chemistry. 382:132340. https://doi.org/10.1016/j.foodchem.2022.132340.
Lin, S., Oyediran, I., Niu, Y., Brown, S., Cook, D., Ni, X., Zhang, Y., Reay-Jones, F.P., Chen, J., Wen, Z., Dimase, M., Huang, F. 2022. Resistance allele frequency to Cry1Ab and Vip3Aa20 in Helicoverpa zea (Boddie) (Lepidoptera: Noctuidae) in Louisiana and three other southeastern U.S. states. Toxins. 14(4):270. https://doi.org/10.3390/toxins14040270.
Harris-Shultz, K.R., Armstrong, J.S., Carvalho, G., Pereira Segundo, J., Ni, X. 2022. Melanaphis sorghi (Hemiptera: Aphididae) clonal diversity in the United States and Brazil. Insects. 13(5):416. https://doi.org/10.3390/insects13050416.
Zhang, H., Jia, B., Lu, Y., Yoon, S.C., Ni, X., Zhuang, H., Guo, X., Le, W., Wang, W. 2022. Detection of aflatoxin B1 in single peanut kernels by combining hyperspectral and microscopic imaging technologies. Sensors. 22(13):4864. https://doi.org/10.3390/s22134864.
Yu, W., Lin, S., Dimase, M., Niu, Y., Brown, S., Head, G.P., Price, P.A., Reay-Jones, F.P., Cook, D., Reisig, D., Thrash, B., Ni, X., Paula-Moraes, S.V., Huang, F. 2021. Extended investigation of field-evolved resistance of the corn earworm Helicoverpa zea (Lepidoptera: Noctuidae) to Bacillus thuringiensis Cry1A.105 and Cry2Ab2 proteins in the southeastern United States. Journal of Invertebrate Pathology. 183:107560. https://doi.org/10.1016/j.jip.2021.107560.
Punnuri, S., Ayele, A., Harris-Shultz, K.R., Knoll, J.E., Coffin, A.W., Tadesse, H.K., Armstrong, J.S., Wiggins, T., Li, H., Sattler, S.E., Wallace, J. 2022. Genome-wide association mapping of resistance to the sorghum aphid in sorghum bicolor. Genomics. 114(4). Article 110408. https://doi.org/10.1016/j.ygeno.2022.110408.