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Abstract
Background:
Accurate and early detection of vancomycin-resistant enterococci (VRE) is critical for controlling nosocomial infection. In this study, we evaluated the usefulness of a selective chromogenic agar medium and of multiplex PCR for detection of VRE, and both these techniques were compared with the conventional culture method for VRE detection.
Methods:
We performed the following 3 methods for detecting VRE infection in stool specimens: the routine culture method, culturing in selective chromogenic agar medium (chromID VRE, bioMérieux, France), and multiplex PCR using the Seeplex® VRE ACE Detection kit (Seegene Inc., Korea) with additional PCR for vanC genes.
Results:
We isolated 109 VRE strains from 100 stool specimens by the routine culture method. In chromID VRE, all the isolates showed purple colonies, including Enterococcus gallinarum and E. raffinosus, which were later identified using the Vitek card. All VRE isolates were identified by the multiplex PCR method; 100 were vanA-positive E. faecium, 8 were vanA- and vanC-1-positive E. gallinarum, and 1 was vanA-positive E. raffinosus.
Conclusions:
For VRE surveillance, culturing the isolates in chromID VRE after broth enrichment appears to be an accurate, rapid, and easy method for routine screening test. Multiplex PCR is relatively expensive and needs skilled techniques for detecting VRE, but it can be an auxiliary tool for rapid detection of genotype during a VRE outbreak.
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Fig. 1.
Agarose gel electrophoresis of PCR products: Lane M; VRE size marker, Lanes 1, 2, 4-6; E. gallinarum, Lane 3; E. raffinosus, Lanes 7-10; E. faecium. (A) PCR products of clinical isolates of this study using Seeplex® VRE ACE Detection kit: only vanA is observed in all the lanes. (B) PCR products with vanC-1 primer.
Abbreviations: VRE, vancomycin-resistant enterococci; E. gallinarum, Enteroccus gallinarum; E. raffinosus, Enteroccus raffinosus; E. faecium, Enteroccus faecium.
Table 1.
Primer sequences that were used in PCR for vancomycin-resistance genotyping
| Amplified gene | Primer sequence (5′-3′) | size (bp) | |
|---|---|---|---|
| vanC-1 | (F) | GGTATCAAGGAAACCTC | 822 |
| (R) | CTTCCGCCATCATAGCT | ||
| vanC-2, vanC-3 | (F) | CTCCTACGATTCTCTTG | 439 |
| (R) | CGAGCAAGACCTTTAAG |
Table 2.
Results of routine culture, chromID, and PCR method of studied isolates
| Routine culture | N of isolates (%) | ChromID (color) | Phenotype (N) | Genotype (by PCR) |
|---|---|---|---|---|
| E. faecium | 100 (91.7) | Purple | VanA (88) VanB (12) | vanA |
| E. gallinarum | 8 (7.3) | Purple | VanA (8) | vanA with vanC-1 |
| E. raffinosus | 1 (0.9) | Purple | VanA (1) | vanA |
| E. casseliflavus 1 (0.9) | NG | NT | NT | |
| Total | 109∗ |
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