Screening for colonization with methicillin-resistant Staphylococcus aureus (MRSA) is a key aspect of infection control to limit the nosocomial spread of this organism. Current methods for the detection of MRSA in clinical microbiology laboratories, including molecularly based techniques, require a culture step and the isolation of pure colonies that result in a minimum of 20 to 24 h until a result is known. We describe a qualitative in vitro diagnostic test for the rapid detection of MRSA directly from nasal swab specimens (IDI-MRSA; Infectio Diagnostic, Inc., Sainte-Foy, Québec, Canada), based upon a real-time PCR and direct detection of MRSA via amplicon hybridization with a fluorogenic target-specific molecular beacon probe. Samples from 288 patients were analyzed for the presence of MRSA with the IDI-MRSA assay, compared to detection by either direct plating or enrichment broth selective culture methods. The diagnostic values for this MRSA screening method were 91.7% sensitivity, 93.5% specificity, 82.5% positive predictive value, and 97.1% negative predictive value when compared to culture-based methods. The time from the start of processing of specimen to result was approximately 1.5 h. In our hands, the IDI-MRSA assay is a sensitive and specific test for detection of nasal colonization with MRSA and providing for same-day results, allowing more efficient and effective use of infection control resources to control MRSA in health care facilities.

Surveillance cultures for vancomycin-resistant enterococci (VRE) and subsequent characterization of the isolates can be extremely burdensome and difficult. Therefore, efficient and reliable schemes for the characterization of surveillance isolates are needed. In this study, a commercial agar (bile esculin azide agar with 6 g of vancomycin per ml [BEAA]; Remel, Lenexa, Kans.) was used in the initial screening step to establish relatively rapid (i.e., in <24 h from the time of isolation) phenotype-based and PCR-based schemes for the detection and characterization of VRE. The phenotype-based scheme included Gram staining of growth on BEAA and subculture of cocci on sheep blood agar plates for vancomycin disk diffusion and pyrazinamidase (PYR) testing. For the PCR scheme, inocula for van gene detection were taken directly from the BEAA plates. The phenotypic approach was applied to 378 surveillance cultures that yielded growth on BEAA. Gram staining quickly eliminated gram-positive bacilli from further testing, and a negative PYR test classified 25 additional isolates as probable pediococci. A positive PYR test reliably identified 121 single-patient VRE isolates that included 83 Enterococcus faecium, 33 E. gallinarum, and 5 E. casseliflavus strains. The vancomycin inhibition zone size clearly distinguished VanA and VanB strains from VanC strains within 24 h of BEAA isolation. All VanA and VanB strains failed to produce zones of >6 mm, while only one VanC strain produced a zone of <15 mm. Challenging this phenotypic scheme with 47 stock VRE isolates produced similar findings. In direct PCR analyses, false-negative vanA and vanB results occurred with 12% of the specimens. Many of the false-negative reactions also failed to produce an internal control product, which underscores the need for including control primers when a PCR scheme is used. In summary, the phenotype-and the PCR-based schemes provide efficient methods for characterizing VRE within 24 h of isolation.

The development of reliable and rapid methods for the identification of patients colonized with vancomycin-resistant enterococci (VRE) is central to the containment of this agent within a hospital environment. To this end, we evaluated a prototype chromogenic agar medium (VRE-BMX; bioMérieux, Marcy l'Etoile, France) used to recover VRE from clinical specimens. This medium can also identify isolated colonies as either vancomycin-resistant Enterococcus faecium or Enterococcus faecalis, based on distinct colony colors. We compared the performance of VRE-BMX with bile esculin azide agar supplemented with vancomycin (BEAV). For this study, 147 stool samples were plated on each test medium and examined after 24 and 48 h of incubation. At 24 h, the sensitivity and specificity of each medium were as follows: BEAV, 90.9% and 89.9%, respectively; VRE-BMX, 96.4% and 96.6%, respectively. The positive predictive values (PPV) of VRE-BMX and BEAV at 24 h were 89.8% and 80.7%, respectively. VRE-BMX provided the identification of 10 isolates of vancomycin-resistant E. faecalis and 4 isolates of vancomycin-resistant E. faecium that were not recovered by BEAV. Further, VRE-BMX was capable of identifying patients colonized with both E. faecium and E. faecalis, a feature useful for infection control purposes that is not a function of BEAV. In terms of the recovery of vancomycin-resistant E. faecium and E. faecalis, the sensitivity and PPV were as follows: BEAV, 75.7% and 74.6%, respectively; VRE-BMX, 95.5% and 91.3%, respectively. In this initial evaluation, we found that VRE-BMX provided improved recovery of VRE from stool specimens, with the added advantage of being able to differentiate between vancomycin-resistant E. faecalis and E. faecium. Extending the incubation period beyond 24 h did not significantly improve the recovery of VRE and resulted in decreased specificity.

We report a fluorescence polarization immunoassay (FPIA) for zidovudine (azidothymidine; Retrovir). This assay is accurate and specific over the clinically relevant range of zidovudine concentrations in serum (from 1 to 1,250 ng/ml; from 0.004 to 4.8 ,LM) and is unaffected by potentially interfering compounds in the sera of patients with renal or hepatic failure. Cross-reactivity with structural analogs of zidovudine (including zidovudine glucuronide) is less than 0.05%, except for cross-reactivities of 0.2, 0.3, and 0.4% with 3-methylthymidine, 3',5'-dideoxythymidine, and A22U (the optical isomer of zidovudine), respectively. The FPIA for zidovudine is more sensitive and more specific than high-performance liquid chromatography (HPLC); it requires 50 to 60 or 200 versus 500 ,il of serum and is faster to perform (45 specimens per h with the FPIA versus 3 specimens per h with HPLC). The zidovudine FPIA compares well with the radioimmunoassay. A correlation coefficient of 0.992 was observed with 31 serum specimens examined by both methods. All three assays (FPIA, radioimmunoassay, and HPLC) are unaffected by the heat treatment used to inactivate human immunodeficiency virus. The zidovudine FPIA should be particularly useful for analyzing specimens from large numbers of human immunodeficiency virus-infected patients receiving zidovudine in current clinical trials.

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