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

Location: Crop Genetics and Breeding Research

2020 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, CIMMYT and US national germplasm center (GRIN database) were evaluated for resistance to whorl and ear feeding insects, and reduced aflatoxin accumulation. In addition, 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 2019, a total of nine sets of breeding crosses at various stages (F1 to F6) and RIL populations were advanced for new germplasm development with insect resistance and reduced aflatoxin accumulation. Ninety-four new hybrid testcrosses were evaluated in a four-replication RCBD for both yield and aflatoxin in 2018 and 2019. Also in 2019 another set of 112 new hybrids was also evaluated for yield and aflatoxin in a four-replicate RCBD experiment. ARS scientists at Tifton, Georgia, also participated in the Georgia state variety trial for corn in 2019, and evaluated 72 commercial corn hybrids for ear-feeding insect resistance in five years, and provided the information for growers to use for hybrid selection, and for the purpose of not only assessing efficacy of new transgenic technology, but also monitoring Bt resistance in natural insect pest populations with existing technology. In multidisciplinary collaborative/cooperative research efforts in our region, ARS scientists at Tifton, Georgia, continue to participate in the South Eastern Regional Aflatoxin Trial (SERAT) with two separate trials for yield and aflatoxin accumulation, respectively. Six hybrids from our team were entered for the trial in 2019 and our location again served as a test location for yield and aflatoxin trials with 47 entries each. ARS scientists at Tifton, Georgia, also work with other scientists from the Aflatoxin Mitigation Center of Excellence (AMCOE) project by screening a set of 43 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 2019, ARS scientists at Tifton, Georgia, advanced 489 S4-S6 selections by selecting for superior agronomic traits with fall armyworm resistance, low aflatoxin accumulation, and superior agronomic traits. Approximately 650 additional selections in various stages of development (F1 – F5) were advanced in 2019. A minimum path set of 41 B73-teosinte introgression lines (ILs) was acquired from the Maize Genetics Stock Center. Seed production on these lines was low in our environment, but enough seed for experiments was recovered for all but one IL. The remaining 40 ILs and B73 were screened for aflatoxin in inoculated experiments over two seasons (RCBD, 4 reps/year). Two ILs had significantly lower aflatoxin than B73. Seed was increased for future experiments to screen for resistance to insects or other diseases. ARS scientists at 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 scientists at Tifton, Georgia, 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 four years (2014-2017) 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. In 2019 and 2020, a collaborator from the University of Georgia collected samples from our G2F field for a microbiome screening experiment. 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 scientists at Tifton, Georgia, have been participating in a five-year area-wide IPM project for sugarcane aphid management since 2016, in addition to participating in the State Variety Test every year. In 2019, 52 grain, 40 silage, and 30 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, ARS scientists at Tifton, Georgia, are working on new germplasm screening and development. A total of 296 mostly grain sorghum germplasm lines from the sorghum association panel (SAP) were screened for sugarcane aphid and anthracnose resistance in 2019. This SAP experiment was also used to help develop a robot-based phenotyping platform in collaboration with Fort Valley State University and Iowa State University. A total of 11 breeding crosses were made in 2015 using sorghum inbreds with aphid and fall armyworm resistant parents, and a total of 1001 segregating accessions, derived from these crosses are being continuously evaluated and advanced in 2019. In 2019, seven sweet sorghum F7 selections were evaluated for sugarcane aphid resistance, lodging, juice brix (sugar content) and other traits (3 rep RCBD). Seed was increased and the four best lines are being evaluated again in 2020 at Tifton,Georgia, Mississippi State, Mississippi, and Lubbock, Texas, by ARS collaborators. In order to map QTL for sugarcane aphid resistance, in 2018 and 2019 ARS scientists at Tifton, Georgia, planted 104 RILs from the cross PI 602951 x SC112 in RCBD experiments in naturally infested fields in Tifton, Georgia. SC112 is highly resistant to sugarcane aphid. In cooperation with an ARS scientist at Mayaguez, Puerto Rico, a major QTL for aphid resistance was found on Chromosome 6. Markers linked to this QTL will be useful for breeding sugarcane aphid resistance into elite sorghum lines. An additional 125 lines were advanced to F4 from the cross SC173 x N98, which will be used for future mapping experiments.

Accomplishments
1. Chemical characterization of sweet sorghum juice and bagasse. Sweet sorghum juice is traditionally used to produce a high-value edible syrup and has potential uses in fermentation to produce fuel ethanol or higher-value bio-based products. However, selecting and breeding sweet sorghum for specific juice chemical properties is slow and laborious. The effects of environment and biotic stresses, such as sugarcane aphid infestation, on juice properties are also largely unknown. Juice and bagasse were collected from different cultivars grown in various environments over three years at Tifton, Georgia. In collaboration with an ARS scientist at New Orleans, Louisiana, rapid analytical techniques were used to screen the sweet sorghum juice and bagasse for various chemical properties. These techniques included cyclic voltammetry and fluorescence excitation emission spectrophotometry with parallel factor analysis (EEM/PARAFAC). Three fluorescent chemical signatures were detected in the juice by EEM/PARAFAC (two protein-like structures and a conjugated aromatic structure), indicative of redox-active compounds. Rapid detection of chemical signatures could be used to pre-screen breeding material for juice quality traits, without tedious or expensive chemical separations. In addition, a correlation between trans-aconitic acid and sugarcane aphid tolerance was observed, suggesting that this compound could play a role in biotic stress tolerance in sorghum.

Review Publications
Kaur, G., Guo, J., Brown, S., Head, G.P., Price, P.A., Paula-Moraes, S., Ni, X., Dimase, M., Huang, F. 2019. Field-evolved resistance of Helicoverpa zea (Boddie) to transgenic maize expressing pyramided Cry1A.105/Cry2Ab2 proteins in northeast Louisiana, the United States. Journal of Invertebrate Pathology. 163:1-20.
Li, S., Hussain, F., Unnithan, G.C., Dong, S., Ulabdin, Z., Gu, S., Mathew, L.G., Fabrick, J.A., Ni, X., Carriere, Y., Tabashnik, B.E., Li, X. 2019. A long non-coding RNA regulates cadherin transcription and susceptibility to Bt toxin Cry1Ac in pink bollworm, Pectinophora gossypiella. Pesticide Biochemistry and Physiology. 158:54-60. https://doi.org/10.1016/j.pestbp.2019.04.007.
Zhao, X., Wang, W., Ni, X., Chu, X., Li, Y., Lu, C. 2019. Utilising near-infrared hyperspectral imaging to detect low-level peanut powder contamination of whole wheat flour. Biosystems Engineering. 184:55-68.
Pekar, J.J., Murray, S.C., Isakeit, T.S., Scully, B.T., Guo, B., Knoll, J.E., Ni, X., Abbas, H.K., Williams, W.P., Xu, W. 2019. Evaluation of elite maize inbred lines for reduced Aspergillus flavus infection, aflatoxin accumulation, and agronomic traits. Crop Science. 59:2562-2571. https://doi.org/10.2135/cropsci2019.04.0206.
Uchimiya, M., Knoll, J.E. 2019. Accumulation of carboxylate and aromatic fluorophores by a pest-resistant sweet sorghum [Sorghum bicolor (L) Moench] genotype. ACS Omega. 4(24):20519-20529. https://doi.org/10.1021/acsomega.9b02267.
Falcon, C.M., Kaeppler, S.M., Spalding, E.P., Miller, N.D., Haase, N., Alkhalifah, N., Bohn, M., Buckler IV, E.S., Campbell, D.A., Ciampitti, I., Coffey, L., Edwards, J.W., Ertl, D., Flint Garcia, S.A., Gore, M.A., Graham, C., Hirsch, C.N., Holland, J.B., Jarquin, D., Knoll, J.E., Lauter, N.C., Lawrence-Dill, C.J., Lee, E.C., Lorenz, A., Lynch, J.P., Murray, S.C., Nelson, R., Romay, M., Rocheford, T., Schnable, P., Scully, B.T., Smith, M.C., Springer, N., Tuinstra, M., Walton, R., Weldekidan, T., Wisser, R.J., Xu, W., De Leon, N. Relative utility of agronomic, phenological, and morphological traits for assessing genotype-by-environment interaction in maize inbreds. Crop Science. 2020; 60:62-81. https://doi.org/10.1002/csc2.20035
Mcfarland, B.A., Alkhalifah, N., Bohn, M., Bubert, J., Buckler IV, E.S., Ciampitti, I., Edwards, J.W., Ertl, D., Gage, J.L., Falcon, C.M., Flint Garcia, S.A., Gore, M., Graham, C., Hirsch, C., Holland, J.B., Hood, E., Hooker, D., Jarquin, D., Kaeppler, S., Knoll, J.E., Kruger, G., Lauter, N.C., Lee, E.C., Lima, D.C., Lorenz, A., Lynch, J.P., Mckay, J., Miller, N.D., Moose, S.P., Murray, S.C., Nelson, R., Poudyal, C., Rocheford, T., Rodriguez, O., Romay, M., Schnable, J.C., Schnable, P.S., Scully, B.T., Sekhon, R., Silverstein, K., Singh, M., Smith, M., Spalding, E.P., Springer, N., Thelen, K., Thomison, P., Tuinstra, M., Wallace, J., Walls, R., Wills, D., Wisser, R.J., Xu, W., Yeh, C., De Leon, N. Maize genomes to fields (G2F): 2014 –2017 field seasons: genotype, phenotype, climatic, soil and inbred ear image datasets. BMC Research Notes. 13,71 (2020). https://doi.org/10.1186/s13104-020-4922-8.
Soni, P., Gangurde, S.S., Ortega-Beltron, A., Kumar, R., Parmar, S., Sudini, H.K., Lei, Y., Ni, X., Huai, D., Fountain, J.C., Njoroge, S., Mahuku, G., Radhakrishnan, T., Zhuang, W., Guo, B., Liao, B., Singam, P., Pandey, M.K., Bandyopadhyay, R., Varshney, R.K. 2020. Functional biology and molecular mechanisms of host-pathogen interactions for aflatoxin contamination in groundnut (Arachis hypogaea L.) and maize (Zea mays L.). Frontiers in Microbiology. 11:Article 227 p. 1-22. https://doi.org/10.3389/fmicb.2020.00227.
Jia, B., Wang, W., Ni, X., Chu, X., Yoon, S.C., Lawrence, K.C. 2020. Detection of mycotoxins and toxigenic fungi in cereal grains using vibrational spectroscopic techniques: A review. World Mycotoxin Journal. 13(2):163-178. https://doi.org/10.3920/WMJ2019.2510.
Harris-Shultz, K.R., Knoll, J.E., Punnuri, S., Niland, E., Ni, X. 2020. Evaluation of strains of Beauveria bassiana and Isaria fumosorosea to control sugarcane aphids on grain sorghum. Agrosystems, Geosciences & Environment. 3:e20047. https://doi.org/10.1002/agg2.20047.
Harris-Shultz, K.R., Punnuri, S., Knoll, J.E., Ni, X., Wang, H. 2020. The sorghum epicuticular wax locus Bloomless2 reduces plant damage in P898012 caused by the sugarcane aphid. Agrosystems, Geosciences & Environment. 3:e20008. https://doi.org/10.1002/agg2.20008.
Lahiri, S., Ni, X., Buntin, G., Toews, M.D. 2020. Parasitism of Melanaphis sacchari (Hemiptera: Aphididae) by Lysiphlebus testaceipes (Hymenoptera: Braconidae) in the greenhouse and field. Journal of Entomological Science. 55:14-24.
Jia, B., Wang, W., Ni, X., Lawrence, K.C., Zhuang, H., Yoon, S.C., Gao, Z. 2020. Essential processing methods of hyperspectral images of agricultural and food products. Chemometrics and Intelligent Laboratory Systems. 198:Article 103936. https://doi.org/10.1016/j.chemolab.2020.103936.
Lu, Y., Wang, W., Huang, M., Ni, X., Chu, X., Li, C. 2020. Evaluation and classification of five cereal fungi on culture medium using Visible/Near-Infrared (Vis/NIR) hyperspectral imaging. Infrared Physics and Technology. 105:Article 103206. https://doi.org/10.1016/j.infrared.2020.103206.