A Gap-Free Telomere-to-Telomere Genome Assembly of Talaromyces marneffei Strain B33c Provides Insights into Chromosomal Architecture and Comparative Genomics.

Talaromyces marneffei is a thermally dimorphic fungal pathogen endemic to Southeast Asia and southern China, yet chromosome-scale genomic resources from early clinical isolates remain limited. Here, we combined PacBio HiFi sequencing with Oxford Nanopore and Hi-C validation to generate a gap-free, telomere-to-telomere genome assembly of strain B33c, a 1984 clinical isolate from China. The 28.1-Mb assembly comprises eight chromosomes and enabled detailed characterization of chromosomal architecture, including chromosome termini, the repeat landscape, and putative centromeric regions. Using 3314 single-copy orthologs, we reconstructed a time-calibrated phylogeny that recovered two major lineages among the sampled T. marneffei isolates, with B33c clustering closely with the Vietnamese isolates. Comparative synteny analyses showed a largely conserved chromosome-scale backbone within T. marneffei, alongside isolate-specific rearrangement-like patterns. Orthogroup profiling revealed a widely shared gene-family repertoire with isolate-specific variation, as illustrated by differences in the functional enrichment profiles of the ATCC 18224- and B33c-specific orthogroup sets. Overall, the B33c T2T assembly expands genomic resources for T. marneffei and provides additional strain-specific information for comparative studies of genome organization and evolution.