2011 Annual Report
1a.Objectives (from AD-416)
Identify and develop corn germplasm with resistance to Aspergillus flavus infection/aflatoxin contamination and ear-feeding insects and release this germplasm together with information on molecular markers and methodology that will expedite its deployment into commercially available corn hybrids. Specific objectives include the following: (1) determine the effects of indigenous fungal species and ear-feeding insects on A. flavus infection and aflatoxin accumulation in corn grain; (2) identify new sources of corn germplasm with resistance to A. flavus infection and aflatoxin accumulation and/or resistance to damage by southwestern corn borer, fall armyworm, and corn earworm; (3) identify quantitative trait loci, genes, and proteins associated with resistance in corn to A. flavus infection, aflatoxin accumulation, and insect damage; and (4) enhance corn germplasm with resistance to A. flavus infection, aflatoxin accumulation, and insect damage and release germplasm lines as sources of resistance.
1b.Approach (from AD-416)
Objective 1. Determine the effects of indigenous fungal species and ear-feeding insects on A. flavus infection and aflatoxin accumulation in corn grain. Colonization of corn grain is rarely by a single fungal species, but rather a mixture of fungi. Fusarium verticillioides (syn. F. moniliforme) is the most commonly reported fungus infecting corn in the USA, and it is frequently found together with A. flavus. Acremonium zeae is a common contaminant of preharvest corn in the Southeast. It has been reported to suppress growth of both A. flavus and F. verticillioides in laboratory experiments. The interactions of these fungi will be investigated to determine whether F. verticillioides and A. zeae affect A. flavus infection of corn grain and the subsequent accumulation of aflatoxin, and if so, whether these fungi are impediments to the identification of aflatoxin-resistant corn germplasm. The association between insect damage and aflatoxin accumulation in different corn genotypes will be investigated and the extent to which resistance to damage by southwestern corn borer, Diatraea grandiosella; fall armyworm, Spodoptera frugiperda; or corn earworm, Helicoverpa zea, reduces aflatoxin contamination will be determined. Objective 2. Identify new sources of corn germplasm with resistance to A. flavus infection and aflatoxin accumulation and/or resistance to damage by southwestern corn borer, fall armyworm, and corn earworm. Corn germplasm from diverse backgrounds will be screened for resistance to A. flavus/aflatoxin, southwestern corn borer, fall armyworm, and corn earworm. Information on the effects of other fungi or insects on A. flavus/aflatoxin accumulation (Objective.
1)will be used to refine and improve techniques for evaluating germplasm for resistance. Newly identified sources of resistance will be used to pursue Objectives 3 and 4. Objective 3. Identify quantitative trait loci, genes, and proteins associated with resistance in corn to A. flavus infection, aflatoxin accumulation, and insect damage. Populations of F2:3 families and recombinant inbred lines derived from crosses between aflatoxin or insect resistant inbred lines and susceptible lines will be used to identify quantitative trait loci (QTL) associated with resistance. Resistant and susceptible corn inbred lines and recombinant inbred lines will be used in complementary investigations to identify candidate genes and proteins associated with resistance. Molecular markers identified in these investigations will be used in developing improved germplasm lines (Objective 4). Objective 4. Enhance corn germplasm with resistance to A. flavus infection, aflatoxin accumulation, and insect damage and release germplasm lines as sources of resistance. Both breeding methods based on phenotypic performance and those based on molecular markers will be used to enhance germplasm with resistance to aflatoxin contamination and insect damage. The effectiveness of molecular markers based on QTL, genes, and proteins identified in Objective 3 in transferring resistance to A. flavus/aflatoxin and insect damage into germplasm lines with desirable agronomic qualities will be determined.
Two corn germplasm lines, Mp718 and Mp719, developed by ARS at Mississippi State will be released as sources of resistance to aflatoxin contamination. These lines, which were selected from Mp715 x Va35, are 2 weeks earlier maturing than Mp715, the resistant parent. Other lines selected from Mp715 x Va35 and Mp313E x Va35 show promise and are being further evaluated for resistance to aflatoxin accumulation and other desirable qualities. Germplasm accessions obtained through the Germplasm Enhancement of Maize (GEM) project and from the International Maize and Wheat Improvement Center (CIMMYT) were evaluated for resistance to aflatoxin accumulation and insect damage. Some lines developed from GEM germplasm over the past several years exhibit high levels of resistance to aflatoxin accumulation and are being evaluated for possible release. A panel of 300 diverse lines from U.S. breeding programs and CIMMYT was chosen for an association mapping project to identify genes associated with reduced aflatoxin accumulation. These lines were crossed with Va35, a susceptible line; the testcrosses were evaluated for Aspergillus flavus infection, aflatoxin accumulation, and corn earworm damage in field tests conducted in two locations in Mississippi and two locations in Texas in 2009 and 2010. Some CIMMYT lines appear to be good sources of resistance to aflatoxin accumulation. The results should be useful in identifying genes associated with resistance to A. flavus/aflatoxin and in determining which lines possess unique genes for resistance. Genotyping via high throughput sequencing is being done in cooperation with USDA-ARS at Cornell University. Further data analysis will use this genotypic information to determine distribution of rare alleles and assess their importance in determining or influencing phenotype. Experiments to identify quantitative trait loci (QTL) for resistance to A. flavus/aflatoxin in Mp313E, Mp715, and Mp717 provided valuable information on molecular markers for resistance to aflatoxin accumulation in corn. Estimates of QTL associated with resistance are being used to develop near-isogenic lines from backcrosses of Mp313E, Mp715, and Mp717 to B73, T173, and Va35. The lines are now in fifth backcross generation. Application of A. flavus strains that do not produce aflatoxin to corn fields as a bio-control method to reduce losses to aflatoxin is gaining acceptance in many areas. When susceptible and resistant corn hybrids were inoculated with a toxin-producing A. flavus strain, aflatoxin contamination was 10× higher in the susceptible hybrids than resistant hybrids. When inoculated with a combination of toxin-producing and non-producing strains, toxin production in both susceptible and resistant hybrids was reduced by about 75%. Aflatoxin accumulation in the susceptible hybrids was still 6x the tolerance level of 20 parts per billion (ppb) established by the FDA; aflatoxin accumulation in resistant hybrids was less than 20 ppb. Using hybrids with genetic resistance in combination with a non-toxin-producing strain of A. flavus to reduce aflatoxin contamination is a promising means of reducing aflatoxin contamination.
Corn germplasm lines with resistance to aflatoxin accumulation released. Two new corn germplasm lines developed by ARS at Mississippi State are being released as additional sources of resistance to aflatoxin contamination. Over seven years of field testing, accumulation was 90% lower in these lines, Mp718 and Mp719, than in susceptible checks. Mp718 and Mp719, along with lines already released by ARS at Mississippi State including Mp313E, Mp715, and Mp717, will be used by commercial companies to develop corn hybrids with genetic resistance to aflatoxin contamination. These germplasm lines exhibit resistance to accumulation of both aflatoxin and Aspergillus (A.) flavus biomass, and in hybrids significant general combining ability for reduced A. flavus infection and reduced aflatoxin accumulation. This indicates that corn hybrids with genetic resistance to A. flavus infection and aflatoxin accumulation can be developed from these lines, and losses to aflatoxin contamination can be reduced by selecting either directly for reduced aflatoxin accumulation or indirectly for reduced fungal infection. Commercial hybrids with genetic resistance are essential to the reduction or elimination of losses to aflatoxin contamination in corn.
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Ankala, A., Bolla, B.K., Shivaji, R., Williams, W.P., Wilkinson, J. 2011. Comparative analysis of the performance of Aspergillus flavus on resistant and susceptible maize genotypes during infection. Fungal Ecology. 4:32-41.
Kelley, R.Y., Gresham, C., Harper, J., Bridges, S.M., Warburton, M.L., Hawkins, L.K., Pechanova, O., Peethambaran, B., Pechan, T., Luthe, D.S., Mylroie, J.E., Ankala, A., Ozkan, S., Henry, W.B., Williams, W.P. 2010. Integrated database for identifying candidate genes for Aspergillus flavus resistance in maize. BMC Bioinformatics. 11(6):S25.
Setter, T.L., Yan, J., Warburton, M.L., Ribaut, J., Xu, Y., Sawkins, M., Buckler, E.S., Zhang, Z., Gore, M.A. 2010. Genetic association mapping identifies single nucleotide polymorphisms in genes that affect abscisic acid levels in maize floral tissues during drought. Journal of Experimental Botany. 62:701-716. doi:10.1093/jxb/erq308.
Williams, W.P., Windham, G.L., Krakowsky, M.D., Scully, B.T., Ni, X. 2010. Aflatoxin accumulation in BT and non-BT maize testcrosses. Journal of Crop Improvement. 24:392-399.
Pechanova, O., Techan, T., Williams, W.P., Luthe, D.S. 2011. Proteomic analysis of the maize rachis: Potential roles of constitutive and induced proteins in resistance to Aspergillus flavus and aflatoxin. Proteomics. 11:114-127.
Williams, W.P., Ozkan, S., Ankala, A., Windham, G.L. 2011. Ear rot, aflatoxin accumulation, and fungal biomass in maize after inoculation with Aspergillus flavus. Field Crops Research. 120:196-200.
Gill, T., Sandoya, G., Williams, W.P., Luthe, D.S. 2011. Belowground resistance to western corn rootworm in Lepidopteran-resistant maize genotypes. Journal of Economic Entomology. 104:299-307.
Warburton, M.L., Brooks, T.D., Windham, G.L., Williams, W.P. 2011. Identification of novel QTL contributing resistance to aflatoxin accumulation in maize. Molecular Breeding. 27:491-499.
Li, Q., Li, L., Yang, X., Warburton, M.L., Bai, G., Dai, J., Li, J., Yan, J. 2010. Relationship, evolutionary fate and function of two maize co-orthologs of rice GW2 associated with kernel size and weight. Biomed Central (BMC) Plant Biology. 10:143-157.
Yan, J., Warburton, M.L., Crouch, J. 2011. Association mapping for enhancing maize genetic improvement. Crop Science. 51:433-449.