Molecular Identification and Genetic Diversity of Aspergillus flavus by Using SSR Marker from Chili and Maize

Aspergillus species are major contributors to post-harvest losses and mycotoxin contamination in
agricultural commodities worldwide. This study investigated the morphological and genetic diversity
of Aspergillus flavus isolates from maize (Zea mays L.) and chili (Capsicum annuum L.) in
Bangladesh using combined microbiological and molecular approaches. A total of 38 isolates
collected from Khulna, Jashore, Kushtia, Satkhira, Jhinaidah, and Chuadanga districts were
morphologically characterized under four growth conditions and genotypically analyzed with 13 SSR
markers. Morphological assessment revealed time-dependent colony color patterns, with green
morphotypes predominating under standard conditions and lighter pigmentation under restricted
conditions. Shannon’s Diversity Index indicated the highest morphological diversity in reverse views
after 30 days of culture (R_V_30D). Chi-square tests showed no significant association between
morphological and genetic variation (p > 0.05), indicating that observed morphological differences
were primarily influenced by environmental factors rather than genetic determinants. K-means
clustering and UPGMA analysis identified three morphological clusters based on early colony traits.
Genetic analysis revealed high polymorphism across markers, with allele frequencies ranging from
0.342 (AF48) to 1.000 (four monomorphic markers). Markers AF18, AF48, and AF64 showed the
highest Polymorphism Information Content (PIC ≥ 0.450), with AF64 exhibiting the greatest genetic
diversity (Shannon Index = 0.982). Population differentiation was high (F_ST = 0.630–0.981), with
low gene flow indicating isolation of gene pools between host populations. Linkage disequilibrium
analysis confirmed largely independent loci, except for a nominal association between AF10 and
AF16, which did not withstand Bonferroni correction. STRUCTURE analysis supported a best-fit of
K = 2, revealing strong host-related genetic structuring with low admixture. Phylogenetic analyses
(UPGMA and Neighbor-Joining) and Jaccard distance/network analyses further confirmed that
isolates were more genetically conserved within hosts than between hosts, with some clonal lineages
observed. These findings demonstrate clear host-associated genetic differentiation of A. flavus
populations in Bangladesh, alongside environmentally driven morphological plasticity, providing
insights for epidemiology, mycotoxin risk assessment, and host-specific management strategies to
mitigate post-harvest contamination.

• Supervision