Mark Welch, J. | Search bioRxiv

bioRxiv2026

maniFasta and the AllOralsDB: simplifying construction of comprehensive reference databases for metaproteomics

Metaproteomics aims to capture a taxonomically comprehensive snapshot of proteins in a sample. Design of reference databases is a key aspect of metaproteomic workflows, as these define what is ultimately seen. Databases tailored to focal biomes offer optimal performance, yet their construction often requires drawing on heterogeneous data sources, posing a challenge to reproducibility and documentation. Here we present maniFasta, a tool enabling users to generate standardized, reproducible, and robustly documented protein reference sets from diverse input sources and datatypes. Users provide information on their desired input types and sources, and the output is an integrated database comprising a protein sequence file (FASTA), with harmonized identifiers and standardized headers, and an associated provenance metadata table (manifest). We highlight the value of maniFasta in the context of salivary metaproteomics, addressing the need for a taxonomically comprehensive reference database. The AllOralsDB resource includes human proteins, as well as proteins from bacteria and archaea, fungi and other microeukaryotes, viruses and viroid-like elements, dietary sources, and common contaminants. Together, this work provides a community resource for oral and salivary metaproteomics (https://www.homd.org/ftp/AllOralsDB/), and a versatile and accessible tool for constructing protein databases for metaproteomics generally (https://github.com/KauffmanLab/maniFasta).

microbiology↗

bioRxiv2026

Site-specialization of human oral Porphyromonas species

Site-specificity within the human oral cavity reflects adaptation mechanisms such as genome divergence and metabolic specialization. Members of the genus Porphyromonas are distributed across oral sites in health and disease, yet the specific distribution of taxa and the functional basis of their site-specificity remain poorly understood. We analyzed 1,242 metagenomes from nine oral sites in healthy individuals and 24 subgingival plaque samples from individuals with periodontitis. Competitive mapping to a dereplicated genus-level pangenome of 84 reference genomes, combined with phylogenomic, gene-level detection, and functional profiling, revealed distinct site-specific distribution patterns, ecotype differentiation, and metabolic specialization across Porphyromonas taxa. Porphyromonas pasteri was the most abundant and widespread taxon in healthy subjects, comprising two ecotypes--one mucosal, one plaque-associated. Porphyromonas gingivalis was rare in healthy subjects but present in periodontal disease, although detected in only half of periodontitis samples. P. gingivalis exhibited the broadest metabolic repertoire, suggestive of a survival strategy adaptive to disparate conditions. In contrast, Porphyromonas catoniae, restricted to healthy dental plaque, lacked biosynthetic pathways for cobalamin, biotin, and serine, implying nutritional dependency on other taxa or the host. Porphyromonas endodontalis, detected in subgingival plaque across both health and disease, also lacked several metabolic pathways. A 44 kb conjugative element identified in P. gingivalis was detected across healthy and periodontitis subgingival plaque microbiomes independently of the P. gingivalis chromosome, indicating horizontal transfer. These findings reveal genomic divergence and complex metabolic specialization among Porphyromonas taxa, refining our understanding of their role in the ecological structure of the human oral microbiome. ImportanceMicrobial specialization within host-associated ecosystems is a key driver of community structure and function. The human oral cavity, with its spatially structured and ecologically diverse niches, offers a powerful model for studying these patterns. Our findings reveal that Porphyromonas taxa are largely restricted to specific oral sites, with Porphyromonas pasteri emerging as the most abundant and ecologically versatile member of the genus, and that large mobile elements shape genomic diversity across oral taxa. This work advances our understanding of microbial biogeography and highlights the ecological and metabolic significance of Porphyromonas taxa within the complex oral community.

microbiology↗