Global Comparative Genomics of Stenotrophomonas maltophilia Reveals Cryptic Species Diversity, Resistome Variation, and Population Structure.

Background

Stenotrophomonas maltophilia is an increasingly important multidrug-resistant opportunistic pathogen frequently isolated from clinical, environmental, and plant-associated niches. Despite its medical relevance, the global population structure, species-complex boundaries, and genomic determinants of antimicrobial resistance (AMR) and ecological adaptation remain poorly resolved, partly due to inconsistent annotations and fragmented genomic datasets.

Methods

Approximately 2400 genome assemblies annotated as Stenotrophomonas maltophilia were available in the NCBI Assembly database at the time of query. After pre-download filtering to exclude metagenome-assembled genomes and atypical lineages, 1750 isolate genomes were retrieved and subjected to stringent quality control (completeness ≥ 90%, contamination ≤ 5%, ≤500 contigs, N50 ≥ 10 kb, and ≤1% ambiguous bases), yielding a final curated dataset of 1518 high-quality genomes used for downstream analyses. Genomes were assessed using CheckM, annotated with Prokka, and compared using average nucleotide identity (ANI), pan-genome analysis, core-genome phylogenomics, and functional annotation. AMR genes, mobile genetic elements (MGEs), and metadata (source, host, and geographic origin) were integrated to assess lineage-specific genomic features and ecological distributions.

Results

ANI-based clustering resolved the S. maltophilia complex into multiple distinct genomospecies and revealed extensive misidentification of publicly deposited genomes. The pan-genome was highly open, reflecting strong genomic plasticity driven by accessory gene acquisition. Core-genome phylogeny resolved well-supported clades associated with clinical, environmental, and plant-related niches. Resistome profiling showed widespread intrinsic MDR determinants, with certain lineages enriched for efflux pumps, β-lactamases, and trimethoprim-sulfamethoxazole resistance markers. MGE analysis identified lineage-specific integrative conjugative elements, prophages, and transposases that correlated with source and geographic distribution.

Conclusions

This large-scale analysis provides the most comprehensive genomic overview of the S. maltophilia complex to date. Our findings clarify species boundaries, highlight substantial taxonomic misannotation in public databases, and reveal lineage-specific AMR and mobilome patterns linked to ecological and clinical origins. The curated dataset and evolutionary insights generated here establish a foundation for global genomic surveillance, epidemiological tracking, and future studies on the evolution of antimicrobial resistance in S. maltophilia.