Comparison of two multiplexed PCR assays for the detection of HSV-1, HSV-2, and VZV with extracted and unextracted cutaneous and mucosal specimens.

BACKGROUND

Several analyte specific reagents (ASRs) are available for the detection and differentiation of HSV-1, HSV-2, and VZV in clinical specimens. However, there is limited data on the test performance of these reagents used in multiplexed PCR assays.

OBJECTIVE

This study compared the performance of two multiplexed ASR sets for detection of HSV-1, HSV-2, and VZV in dermal specimens.

STUDY

DESIGN

Two commercially available ASRs were combined to produce multiplexed PCR assays for simultaneous detection of HSV-1, HSV-2, and VZV. Seeded samples were used to determine the limit of detection (LOD) for each assay. Patient samples (n=156) were tested in duplicate and results for each method compared to the reference standard of culture. Both extracted and unextracted specimens were used in the study.

RESULTS

Both multiplexed PCR assays showed similar test performance, with minimal LOD differences observed. The LOD was 10(3) copies/mL for HSV-1 and HSV-2 using the Focus assay compared to 5×10(3) copies/mL and 2×10(4) copies/mL, respectively for the EraGen assay. Both assays showed equal performance for VZV with a LOD of 5×10(3) copies/mL. Analytical specificity testing showed no cross reactivity with other selected DNA viruses. Both assays showed similar performance when clinical samples were tested using both extracted and unextracted specimens.

CONCLUSION

Commercially available ASRs can be successfully multiplexed for the PCR detection of HSV-1, HSV-2, and VZV using dermal specimens. Either direct testing or nucleic acid extracted specimens can be used with similar performance in these assays.