Noncanonical Vancomycin Resistance Cluster from Desulfitobacterium hafniense Y51

Abstract

ABSTRACT The glycopeptide vancomycin is a drug of last resort for infection with gram-positive organisms, and three genes are vital to resistance: vanH , vanA , and vanX . These genes are found in a vanHAX cluster, which is conserved across pathogenic bacteria, glycopeptide antibiotic producers, and other environmental bacteria. The genome sequence of the anaerobic, gram-positive, dehalogenating bacterium Desulfitobacterium hafniense Y51 revealed a predicted vanA homolog; however, it exists in a vanAWK-murFX cluster, unlike those of other vancomycin-resistant organisms. Using purified recombinant VanA from D. hafniense Y51, we determined its substrate specificity and found it to have a 42-fold preference for d -lactate over d -alanine, confirming its activity as a d -Ala- d -Lac ligase and its annotation as VanA. Furthermore, we showed that D. hafniense Y51 is highly resistant to vancomycin, with a MIC for growth of 64 μg/ml. Finally, vanA Dh is expressed during growth in vancomycin, as demonstrated by reverse transcription-PCR. This finding represents a new glycopeptide antibiotic resistance gene cluster and expands the genetic diversity of resistance to this important class of antibiotic.