| Summary: | Bacteria of the genus <i>Cutibacterium</i> are Gram-positive commensals and opportunistic pathogens that represent a major challenge in the diagnosis and treatment of implant-associated infections (IAIs). This study provides insight into the distribution of different sequence types (STs) of <i>C. acnes</i>, and the presence of virulence factors (VFs) in 64 <i>Cutibacterium</i> spp. isolates from suspected or confirmed IAIs obtained during routine microbiological diagnostics. Fifty-three <i>C. acnes</i>, six <i>C. avidum</i>, four <i>C. granulosum</i>, and one <i>C. namnetense</i> isolate, collected from different anatomical sites, were included in our study. Using whole-genome sequencing and a single-locus sequencing typing scheme, we successfully characterized all <i>C. acnes</i> strains and revealed the substantial diversity of STs, with the discovery of six previously unidentified STs. Phylotype IA<sub>1</sub>, previously associated with both healthy skin microbiome and infections, was the most prevalent, with ST A1 being the most common. Some minor differences in STs’ distribution were observed in correlation with anatomical location and association with infection. A genomic analysis of 40 investigated VFs among 64 selected strains showed no significant differences between different STs, anatomical sites, or infection-related and infection undetermined/unlikely groups of strains. Most differences in VF distribution were found between strains of different <i>Cutibacterium</i> spp., subspecies, and phylotypes, with CAMP factors, biofilm-related VFs, lipases, and heat shock proteins identified in all analyzed <i>Cutibacterium</i> spp.
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