A comparison of virus isolation, indirect immunofluorescence and nested multiplex polymerase chain reaction for the diagnosis of primary and recurrent herpes simplex type 1 and type 2 infections

Abstract

134 swabs in viral transport medium were received from 126 patients with suspected clinical HSV-1 and HSV-2 infections. They were tested by (i) nested multiplex polymerase chain reaction NMPCR (strongly positive specimens had visible bands on both rounds of PCR) without prior extraction, (ii) culture in primary rhesus monkey kidney, E6-Vero, RD and HEp-2 cells and (iii) antigen detection by immunofluorescence (IF). Antigen detection employed four novel pools (A–D) of monoclonal antibodies (Mab): A was HSV-1 specific, B was HSV-2 specific while C and D were generic. In comparison to NMPCR the sensitivity and specificity of (i) culture was 59% (22/37) and 100% (134/134), (ii) IF by Pool A was 59% (16/27) and 100% (117/117), (iii) IF by Pool B was 40% (4/10) and 100% (130/130) and (iv) IF by Pools C and D were 60% (18/30) and 100% (96/96). Specimens positive by culture were more likely to be strongly positive by NMPCR (χ² P=0.004). Typing by each method concurred on all occasions. NMPCR was cost effective, easier to perform and was the most sensitive method for HSV detection. It should become the method of choice for HSV diagnosis.

Introduction

Herpes simplex virus types 1 and 2 (HSV-1 and HSV-2) infections are amongst the commonest infections encountered clinically in medical practice and worldwide are the main cause of genital ulceration (Beyrer et al., 1998, do Nascimento et al., 1998, Mertz et al., 1998, Risbud et al., 1999). They present with acute primary and recurrent cutaneous and mucocutaneous ulcers and the more lethal conditions of encephalitis and disseminated neonatal infection. HSV-2 is linked specifically to genital ulceration but increasingly HSV-1 is also so encountered and in Northern Ireland has become the predominant genital type recovered from women (Christie et al., 1998). Culture remains the mainstay of diagnosis for HSV associated ulceration and is more sensitive than direct antigen detection (Kok et al., 1998, Slomka et al., 1998), especially when using a shell vial technique (Reina et al., 1997). An exception is recognised for corneal scrapings where immunofluorescence (IF) can yield superior results to culture (Pramod et al., 1998) and has the potential for in vivo application (Sharma and Shimeld, 1997). Increasingly HSV specific polymerase chain reaction (PCR) has been reported with higher sensitivity to either culture (Beards et al., 1998, do Nascimento et al., 1998, Slomka et al., 1998) or direct antigen detection (Slomka et al., 1998, Risbud et al., 1999). Cost and technical problems limit its routine application (Slomka et al., 1998). A comparison is reported of direct antigen detection with a novel set of monoclonal antibodies (Mab) and a nested multiplex PCR (NMPCR) to virus culture for the routine laboratory diagnosis of HSV associated ulceration.