EVALUATION OF AN MGB REAL TIME PCR REAGENT FROM NANOGEN FOR HERPES SIMPLEX VIRUS DETECTION AND TYPING

Herpes simplex virus is the most common cause of acquired, sporadic encephalitis in the United States.  Due to its specificity, sensitivity and potential for speed, PCR is now accepted as the diagnostic gold standard for the identification of HSV in cerebrospinal fluid. Besides sensitive target detection, this assay provides the ability to distinguish between HSV-1 and HSV-2.  A variety of studies indicate that these subtypes display differences in mortality, severity of disease and reactivation frequencies.  In spinal fluid, type 2 is typically associated with more serious outcomes in neonates.  In the adult population, detection of type 2 in the CSF is associated with benign recurrent aseptic meningitis (Mollaret's meningitis), while type 1 is the most common cause of Herpes simplex encephalitis, a life threatening disease. 

            Nanogen’s patented minor groove binder (MGB) technology enables laboratories to develop accurate and sensitive real time PCR assays. In addition, their library of proprietary modified bases provides the ability to accommodate various sequence polymorphisms in the target sequence.  We evaluated a prototype MGB HSV Typing assay that allows simultaneous detection of both HSV Type 1 and 2 viruses in a single sample.  Type specific probes, labeled with different fluorescent dyes, are used to detect a common amplicon.  Including primers and probe for detecting an internal control allows the user to monitor sample extraction and PCR inhibition. The post-PCR melt curve is used to identify the HSV type, as well as aiding in the detection of unknown variants.  We evaluated this pre-release reagent for specificity, sensitivity and the ability to make the correct typing call.