Identification and Validation of Genomic Regions Associated With Charcoal Rot Resistance in Tropical Maize by Genome-Wide Association and Linkage Mapping

Abstract

Charcoal rot is a post-flowering stalk rot (PFSR) disease of maize caused by the fungal pathogen, Macrophomina phaseolina. It is a serious concern for smallholder maize cultivation, due to significant yield loss and plant lodging at harvest, and this disease is expected to surge with climate change effects like drought and high soil temperature. For identification and validation of genomic variants associated with charcoal rot resistance, a genome-wide association study (GWAS) was conducted on CIMMYT Asia association mapping panel comprising 396 tropical-adapted lines, especially to Asian environments. The panel was phenotyped for disease severity across two locations with high disease prevalence in India. A subset of 296,497 high-quality SNPs filtered from genotyping by sequencing was correcting for population structure and kinship matrices for single locus mixed linear model (MLM) of GWAS analysis. A total of 19 SNPs were identified to be associated with charcoal rot resistance with P-value ranging from 5.88 × 10^-06 to 4.80 × 10^-05. Haplotype regression analysis identified 21 significant haplotypes for the trait with Bonferroni corrected P ≤ 0.05. For validating the associated variants and identifying novel QTLs, QTL mapping was conducted using two F_2:3 populations. Two QTLs with overlapping physical intervals, qMSR6 and qFMSR6 on chromosome 6, identified from two different mapping populations and contributed by two different resistant parents, were co-located with the SNPs and haplotypes identified at 103.51 Mb on chromosome 6. Similarly, several SNPs/haplotypes identified on chromosomes 3, 6 and 8 were also found to be physically co-located within QTL intervals detected in one of the two mapping populations. The study also noted that several SNPs/haplotypes for resistance to charcoal rot were located within physical intervals of previously reported QTLs for Gibberella stalk rot resistance, which opens up a new possibility for common disease resistance mechanisms for multiple stalk rots.

Keywords: GWAS–genome-wide association study; charcoal rot; haplotype analysis; linkage (QTL) mapping; maize.

Figure 1

(A) Inflation depicted by Q–Q plots of observed vs. expected –log₁₀ (P-values) plots for charcoal rot using the naïve association model (G-test), GLM (G + Q), and MLM (G + Q + K); G = genotype (fixed), Q = 10 principal components (fixed), K = kinship matrix (random) for CAAM panel. (B) Highly significant SNPs identified from MLM model using Manhattan plot, plotted with the individual SNPs on the X-axis and –log₁₀ P-value of each SNP on the Y-axis. The horizontal line showed the cutoff P-value, and the vertical line represents the identified QTLs and haplotype blocks in these regions for charcoal resistance.

Figure 2

Plot of LOD scores from quantitative trait loci (QTL) analysis for charcoal rot resistance across 10 maize chromosomes in two F2: 3 biparental populations, (A) MSR and (B) FMSR. MSR and FMSR populations were evaluated under artificial inoculation conditions by Macrophomina phaseolina. The horizontal line represents the threshold LOD value of 2.5.