Stenotrophomonas maltophilia in a Rhipicephalus linnaei tick cell culture: detection, antibiotic resistance and multi-locus sequence typing

Ticks harbour diverse microbial communities, but opportunistic and environmentally associated bacteria within these systems remain poorly understood. During attempts to establish primary cell cultures from surface-sterilized embryos of the dog tick Rhipicephalus linnaei, we detected the presence of Stenotrophomonas maltophilia, a multidrug-resistant environmental bacterium and emerging opportunistic pathogen. Microscopic examination of Giemsa-stained smears showed abundant extracellular rod-shaped bacteria closely associated with degenerating tick cells, with no evidence of intracellular infection. Molecular identification based on 16S and 23S rRNA gene sequencing confirmed the bacteria as S. maltophilia, and antibiotic susceptibility testing revealed resistance to amoxicillin-clavulanic acid, imipenem, cefoxitin, and nitrofurantoin, but susceptibility to meropenem. Multi-locus sequence typing (MLST) identified the isolate as sequence type 948 (ST948), which is a single-locus variant of ST408 and a double-locus variant of ST144, both of which are representatives of environmental and clinical S. maltophilia strains from diverse geographic origins. These findings provide the first evidence of S. maltophilia occurring in a tick-derived cell culture system and highlight the need to consider opportunistic bacteria when interpreting tick microbiome and cell culture-based studies, particularly those involving veterinary-relevant tick species.