JAC Advance Access originally published online on May 4, 2007
Journal of Antimicrobial Chemotherapy 2007 60(1):182-183; doi:10.1093/jac/dkm140
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Correspondence |
Experience with the Roche LightCycler VRE detection kit during a large outbreak of vanB2/B3 vancomycin-resistant Enterococcus faecium
1 Department of Pathology, Singapore General Hospital, Outram Road, 169608, Singapore 2 Department of Laboratory Medicine, National University Hospital, Singapore
* Corresponding author. Tel: +65-6321-4505; Fax: +65-6222-6826; E-mail: koh.tse.hsien{at}sgh.com.sg
Keywords: real-time PCR , diagnostics , vancomycin-resistant E. faecium
We read with interest the recent correspondence by Young et al.1 describing the poor specificity of the Roche LightCycler vanA/B detection assay to indicate vanB vancomycin-resistant enterococcal carriage in archived stool samples. We would like to relate our recent experience with this test on fresh stool specimens during a large outbreak of vanB2/B3 vancomycin-resistant Enterococcus faecium (VRE) in Singapore.2
A large hospital-wide screening effort was initiated to define the extent of the outbreak. Fresh stool was inoculated into Enterococcosel broth with 8 mg/L vancomycin. DNA was extracted from broths that changed colour after 24 h using the automated MagNA Pure LC (Roche Diagnostics, Mannheim, Germany) or QIAamp® DNA Minikit (Qiagen GmbH, Germany) and tested for the presence of van genes by real-time PCR using the Roche LightCycler VRE detection kit (Roche). If the real-time PCR was positive, the broth was subcultured onto blood agar and Enterococcosel agar containing 6 mg/L vancomycin.
During the month of April 2005, 695 broth extracts were screened by real-time PCR. Of these, 53 were vanB2/B3 positive. vanB2/B3 VRE was successfully subcultured from 19 broths. A proportion of the broths whose extracts were negative by real-time PCR were also subcultured. All 194 of these failed to grow VRE on subculture. Because not every real-time PCR-negative broth was subcultured, we could not calculate an unbiased value for the sensitivity, specificity and negative predictive value of this test. However, we were able to determine that the positive predictive value (probability that vanB2/B3 enterococci would be successfully subcultured from a broth whose extract was real-time PCR positive) in our study sample was only 36%.
Failure to subculture could be because the original load of VRE was below the threshold for successful subculture (unlikely because of enrichment) or the inadvertent detection of vanB elements in other gut bacteria.3 The latter implies that there will always be an inherent lack of specificity if the test is used in this manner. It has to be mentioned that this test was originally devised to be used on pure cultures rather than inoculated broths or directly on patient samples.
The poor positive predictive value of real-time PCR for vanB2/B3 VRE means that infection control resources may be unnecessarily expended on cases which ultimately prove to be VRE negative. The utility of the Roche LightCycler VRE detection kit on broth culture extracts will therefore depend on local infection control policies and the predominant type of van gene. In our practice, a negative real-time PCR result was used to exclude VRE colonization and all real-time PCR positive cases required confirmation by subculture before aggressive infection control measures were undertaken.
None to declare.
Acknowledgements
We thank Goh Hoei Thein, Michelle Tan and Tiong Swee Choo for their technical assistance.
References
1
Young HL, Ballard SA, Roffey P, et al. Direct detection of vanB2 using the Roche LightCycler vanA/B detection assay to indicate vancomycin-resistant enterococcal carriage—sensitive but not specific. J Antimicrob Chemother (2007) 59:809–10.
2 Koh TH, Hsu LY, Chiu LL, et al. Emergence of epidemic clones of vancomycin-resistant Enterococcus faecium in Singapore. J Hosp Infect (2006) 63:234–6.[CrossRef][Web of Science][Medline]
3
Ballard SA, Grabsch EA, Johnson PDR, et al. Comparison of three PCR primer sets for the identification of vanB gene carriage in feces and correlation with carriage of vancomycin-resistant enterococci: interference by vanB-containing anaerobic bacilli. Antimicrob Agents Chemother (2005) 49:77–81.
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