Moderate-Level Resistance to Glycopeptide LY333328 Mediated by Genes of the vanA and vanB Clusters in Enterococci

Abstract

ABSTRACT Three of five natural plasmids carrying a wild-type vanA gene cluster did not confer LY333328 glycopeptide resistance on Enterococcus faecalis JH2-2 (MIC = 2 μg/ml). The two remaining plasmids conferred resistance to the drug (MIC, 8 μg/ml). The vanB gene cluster did not confer resistance to LY333328, since this antibiotic was not an inducer. Mutations in the vanS B sensor gene that allowed induction by teicoplanin or constitutive expression of the vanB cluster led to LY333328 resistance (MIC, 8 to 16 μg/ml). Overproduction of the VanH, VanA, and VanX proteins for d -alanyl- d -lactate ( d -Ala- d -Lac) synthesis and d -Ala- d -Ala hydrolysis was sufficient for resistance to LY333328 (MIC, 16 μg/ml). Mutations in the host d -Ala: d -Ala ligase contributed to LY333328 resistance in certain VanA- and VanB-type strains, but the MICs of the antibiotic did not exceed 16 μg/ml. Addition of d -2-hydroxybutyrate in the culture medium of mutants that did not produce the VanH d -lactate dehydrogenase led to incorporation of this d -2-hydroxy acid at the C-terminal ends of the peptidoglycan precursors and to LY333328 resistance (MIC, 64 μg/ml). The vanZ gene of the vanA cluster conferred resistance to LY333328 (MIC, 8 μg/ml) by an unknown mechanism. These data indicate that VanA- and VanB-type enterococci may acquire moderate-level resistance to LY333328 (MIC ≤ 16 μg/ml) in a single step by various mechanisms.