Dynamics of Culture-Recovered Cotton Root-Associated Fungi under Charcoal Rot Disease and Preventive Management Regimes

Charcoal rot (CRD), caused by Macrophomina phaseolina, is a major soil-borne disease of cotton, and conventional chemical control often provides inconsistent suppression. This study aimed to characterize management-associated patterns among culture-recovered cotton root-associated fungi and to identify fungal interactions relevant to M. phaseolina suppression. Roots were collected from asymptomatic and symptomatic cotton plants at mid-season and end-season in a field subjected to chemical, Trichoderma-based, and integrated CRD management. Root-associated isolates were recovered by culture-dependent methods and identified using molecular markers and phylogenetic analyses. Selected fungal taxa were evaluated in vitro and in a 33-day greenhouse seedling assay, while commercial 6-pentyl-α-pyrone (6-PP) was examined in a targeted antifungal assay. From approximately 300 recovered isolates, 66 representative morphotypes were selected and molecularly identified, including 56 fungal and 10 bacterial isolates. Aspergillus was the most frequently recovered fungal genus at mid-season, whereas Alternaria became predominant at season’s end, particularly in symptomatic plants. The numbers of recovered fungal isolates and taxa increased modestly in asymptomatic plants but more strongly in symptomatic plants, while Trichoderma-based management was associated with markedly lower numbers of recovered fungal isolates and taxa. Commercial 6-PP inhibited M. phaseolina in a concentration- and time-dependent manner (94.5–100% at nominal 400–800 µg mL-¹ through 72 h). Greenhouse assays showed limited effects on early seedling development during the latent disease stage. Nevertheless, qPCR revealed mean reductions of 92%, 94%, and 99% in M. phaseolina DNA following co-inoculation with A. niger, N. solani, and A. alternata, respectively, compared with 47% for R. arrhizus. Only the reduction associated with A. alternata remained significant after multiplicity adjustment. These findings associate biological and integrated management with distinct patterns in culture-recovered root-associated fungi and identify microbial interactions and natural product activity relevant to future CRD management.