Species composition and functional characteristics of the human multi-organ microbiome: A metagenomic study

Postmortem microbial communities may provide useful information for forensic microbiology, but species-level and functional profiles across multiple cadaveric anatomical sites remain poorly characterized. Here, shotgun metagenomic sequencing was performed on 144 samples from six anatomical sites, including the oral cavity, nasal cavity, trachea, lung, colon, and anus, collected from 24 human cadavers. A total of 15,301,799,968 raw reads were obtained, and 6565 species were identified, and KEGG pathways were annotated at the L1, L2, and L3 levels. Species-level microbial composition differed significantly among anatomical sites. PERMANOVA with permutations blocked by individual identity showed that anatomical site was the dominant factor explaining microbial community variation (R² = 0.3778, p = 0.001, q = 0.001), whereas postmortem interval did not show a significant independent effect within the 1-38-day interval. KEGG functional profiles also differed significantly among anatomical sites at the L2 and L3 levels, and 182 of 214 L3 pathways showed significant site-associated differences after false-discovery-rate correction. Pathway-level mixed-effect models further indicated that anatomical site remained significantly associated with most L3 pathways after accounting for postmortem interval, age, sex, cause of death, and repeated sampling from the same individual. Species-pathway correlation analysis identified significant taxon-function associations, but these were interpreted as correlative rather than direct evidence of species-specific functional contribution. Low-biomass sensitivity analyses indicated that respiratory-site results, especially lung and tracheal findings, should be interpreted cautiously because of high host DNA proportions and low non-host read counts. Inter-site shared occurrence and intra-site co-occurrence analyses further described distributional associations across anatomical sites. This study establishes a multi-site postmortem metagenomic reference framework for characterizing anatomical-site-specific microbial and functional patterns, offering insights into forensic microbiology and postmortem microbial ecology.