Genomics and phenotypic profiling of Bacillus paralicheniformis MB647 from an orthopaedic implant infection.

A rod-shaped, endospore-forming, Gram-positive, facultatively anaerobic bacterium was isolated from an infected orthopaedic implant in Islamabad, Pakistan. The strain, designated MB647, is motile with optimal growth at 37 °C and pH 7.0. Whole genome sequencing followed by genome-based phylogenetic analysis placed MB647 within the family Bacillaceae and confirmed its species identity as Bacillus paralicheniformis, supported by OrthoANI values exceeding 99% relative to reference genomes. B. paralicheniformis is predominantly regarded as an environmental or food-associated species and has only rarely been implicated in human infections. Genomic analysis identified multiple antibiotic resistance determinants conferring resistance to tetracycline, bacitracin, erythromycin, and β-lactams. We also detected putative virulence factors, including hemolysins, proteolytic enzymes (proteases, gelatinases), and biofilm-associated pathways, suggesting a potential role in implant persistence and pathogenesis. Multilocus sequence typing (MLST) based on seven housekeeping genes assigned MB647 to a novel sequence type, ST48. PubMLST comparative analysis confirmed that this allelic profile is unique, with MB647 representing the first and only isolate of ST48 reported from Pakistan. Divergent alleles in recF highlight previously under-sampled genomic diversity in this region. To our knowledge, this is the first report of a human-associated B. paralicheniformis isolate from Pakistan. This study provides crucial genomic and epidemiological insights into an emerging, clinically relevant lineage with a unique resistance and virulence repertoire. ACCESSION NUMBERS: The complete genome sequence of Bacillus paralicheniformis MB647 has been deposited in GenBank under accession number JABWTN000000000.1.