Heteroresistance to teicoplanin in Enterococcus faecium harboring the vanA gene.

Hospital-acquired infections caused by vancomycin-resistant enterococci (VRE) are increasing. The resistance of VRE to multiple antibiotics makes them clinically challenging. There are at least six phenotypes of VRE, including VanA, VanB, VanC, VanD, VanE, and VanG (2, 8). These phenotypes correspond with the genotypes vanA, vanB, vanC, vanD, vanE, and vanG. VanA and VanB phenotypes are common among VRE isolates. In general, vanA genotype VRE shows VanA phenotype with high-level resistance to both vancomycin and teicoplanin, whereas vanB genotype VRE shows VanB phenotype characterized by various levels of resistance to vancomycin but susceptibility to teicoplanin (2). Interestingly, vanA genotype VRE strains were recently found to be susceptible to teicoplanin in East Asia (Japan, South Korea, Taiwan, and even mainland China) and named VanB phenotype-vanA genotype VRE (3, 4, 6, 7, 11). Until now, the mechanism for this phenomenon has not been clear. Among these strains, point mutations in the key region of the vanS gene or impairment of vanY or vanZ in the Tn1546-like element were discovered (3, 4, 7, 11). Also, several vancomycin-heteroresistant Enterococcus faecium isolates with the vanA genotype have been reported since 2001 (1, 5). Several novel point mutations in the vanR, vanS, vanH, vanA, vanX, and vanY genes of Tn1546 were also discovered, resulting in amino acid replacements of VanR, VanS, and VanH. It was concluded that independent novel mutations can give rise to polymorphism, heteroresistance, and clonal diversity among VRE strains. In the present study, we isolated two clinical VRE strains of E. faecium, ZP2298 and ZP2171, with a high level of resistance to vancomycin (MICs 256 g/ml) and teicoplanin (MICs of 96 and 256 g/ml, respectively) from sputum samples from patients in a teaching hospital in mainland China. No prior exposure to glycopeptides was found in the isolates. Filter mating was performed using the two clinical VRE strains of E. faecium ZP2298 and ZP2171 as donors and E. faecium BM4105RF (Fus Rif) as the recipient as previously described (9). Transconjugants were selected on brain heart infusion (BHI) agar plates containing vancomycin (32 g/ml) and rifampin (rifampicin) (256 g/ml). After conjugation, the transconjugants of E. faecium ZP2298 and ZP2171, named E. faecium FB2298 and FB2171, respectively, exhibit high-level resistance to vancomycin (MICs 256 g/ml), but subcolonies were within the zone of inhibition around the teicoplanin Etest strip described for teicoplanin-heteroresistant isolates with teicoplanin MICs of 96 and 256 g/ml, respectively (Fig. 1). The subcolonies, named E. faecium FBV2298 and FBV2171, were selected and showed high-level resistance to both vancomycin and teicoplanin. E. faecium FBV2171 maintained high level of resistance to vancomycin (MIC 256 g/ml) and was heteroresistant to teicoplanin after three serial passages on the BHI agar without vancomycin (named FBG2171). However, the resistance profile of E. faecium FBV2298 did not change after three serial passages on the BHI agar without vancomycin (named FBG2298). This is the first report of a teicoplaninheteroresistant E. faecium isolate. Pulsed-field gel electrophoresis (PFGE) of SmaI-digested genomic material from the recipient strain (E. faecium BM4145), FIG. 1. Vancomycin and teicoplanin Etest results for VREF isolates. The two clinical VREF strains of E. faecium, ZP2298 and ZP2171, had a high level of resistance to vancomycin and teicoplanin (MICs 256 g/ml). After conjugation, the transconjugants of these two strains of E. faecium, FB2298 and FB2171, exhibit high-level resistance to vancomycin (MICs 256 g/ml), with resistant subcolonies present in the clear zone of inhibition around the teicoplanin Etest strip, described for teicoplanin-heteroresistant isolates. The resistant subcolonies, E. faecium FBV2298 and FBV2171, had teicoplanin MICs of 96 and 256 g/ml, respectively. E. faecium FBG2298 and FBG2171 strains were generated after three serial passages of E. faecium FBV2298 and FBV2171, respectively.