Core genome MLST reveals genetic and BafA-associated phenotypic diversities in Bartonella henselae strains

Bartonella henselae is a zoonotic pathogen associated with cat-scratch disease. Although multilocus sequence typing (MLST) has been used for strain classification, its resolution for distinguishing between B. henselae isolates remains limited. We herein developed a B. henselae-specific core genome MLST (cgMLST) scheme based on whole-genome sequencing data and examined the genetic and phenotypic diversities of 80 strains derived from cats, humans, mongooses, and masked palm civets. Using the conventional MLST scheme, the 80 strains were classified into nine sequence types (STs), while cgMLST subdivided them into 72 cgSTs, demonstrating a marked improvement in discriminatory power. The cgMLST scheme comprised 1,183 core genes and showed high applicability across the 80 strains. A phylogenetic analysis revealed that ST1, which has been associated with cat-scratch disease, was further subdivided into three major clusters and two singletons, indicating high genetic heterogeneity within this ST. We also found that the bafA subtypes clustered in a manner that was largely consistent with the cgMLST-based phylogenetic structure, suggesting a close relationship between bafA variations and the genomic background of B. henselae strains. In a human umbilical vein endothelial cell proliferation assay, strains belonging to distinct cgSTs exhibited strain-dependent differences in proliferative capacity, which were associated with the bafA subtype classification. Some strains induced focal cell fragmentation and a reduced cell density at a high multiplicity of infection, indicating strain-dependent differences in endothelial cell injury. Collectively, the present results establish a high-resolution cgMLST framework for B. henselae and demonstrate that genetically distinct strains have diverse endothelial cell phenotypes.