Genetic Variation in the Gene Leading to the BORSA Phenotype in .

BACKGROUND/OBJECTIVES: is a leading cause of bacteraemia in Danish hospitals. Approximately 70% of clinical isolates are penicillin-resistant, which is predominantly due to -mediated β-lactamase production.

METHODS

A collection of 489 strains derived from bacteraemia were cultured and their genomes sequenced.

RESULTS

From this collection, 71% of isolates were methicillin-susceptible (MSSA) harbouring . While most isolates contained the gene belonging to the well-characterised A, B, C and D variants, three strains (1%) produced a BlaZ protein characterised by having threonine residues on both positions 128 and 216 and, therefore, belonged to neither of the established variants. We named this variant, variant F. We report that clinical isolates expressing variant F were resistant to oxacillin. The β-lactamase production phenotype in isolates carrying either of the A, B, C or D variants was only weakly discernible on MIC gradient strip and disk diffusion tests. When the β-lactamases were expressed either from a T7 promoter or from their endogenous promoters in , variant F was significantly better at degrading ampicillin than variant A. We also showed that variant F conferred oxacillin resistance when expressed in an isogenic strain, while variant A did not. Finally, we demonstrated that the F variant threonine 216 played a role in the enzyme's superior activity.

CONCLUSIONS

Our findings demonstrate that the new F variant of BlaZ is sufficient to render a BORSA strain, which is superior in the degradation of common anti-staphylococcal β-lactam antibiotics, such as benzylpenicillin, cloxacillin, and oxacillin. It is sensitive to β-lactamase inhibitors and rapidly degrades nitrocefin. We provide a genetic explanation for the borderline oxacillin-resistant (BORSA) phenotype.