Multilocus Sequence Typing (MLST) and Phylogenetic Study Reveal Species Delimitation of Colletotrichum spp. Associated with Dieback of Guava

The Colletotrichum species complex comprises a significant group of fungal pathogens that infect numerous host plants worldwide. In this research, we examined the population structure, genetic diversity, and species delimitation within the Colletotrichum community linked to dieback-affected guava plants through multilocus sequence typing (MLST) and phylogenetic analysis. We utilized five housekeeping genes: actin (act), calmodulin (cal), glyceraldehyde 3-phosphate dehydrogenase (gapdh), tubulin (tub2), and the internal transcribed spacer region of rDNA (ITS). Isolate genotyping was based on allelic profiles, revealing that the Glyceraldehyde-3-phosphate dehydrogenase GAPDH locus exhibited the highest typing efficiency in MLST analysis. This shows that GAPDH is a potential DNA barcode marker for distinguishing Colletotrichum species in phylogenetic studies. For species delimitation, a multilocus phylogenetic tree was generated using Bayesian inference analysis, consistent with the coalescent theory-based species tree. This analysis resolved the isolates into distinct evolutionary lineages within the Colletotrichum siamense species complex. The coalescent theory-based species tree further supported these lineages as distinct species within the Colletotrichum siamense complex. Based on morpho-genetic analyses of the 10 studied isolates, eight were identified as two new species within the C. siamense complex: C. parkukiae sp nov. and C. psidium sp nov. Additionally, one isolate was identified as C. gloeosporoides, and another as C. communis.