Abstract:

Objective To establish a rapid and reliable simultaneous amplification and testing （SAT） for detecting herpes simplex virus Ⅰ（HSV-1）. Methods The specific primers were designed,and the probes were optimized according to RNA sequences in open reading frame Us5 region of HSV-1 from GenBank,and HSV-1 RNA was amplified by M-MLV reverse transcriptase and T7 RNA polymerase. The fluorescence signals were collected and determined by fluorescence quantitation polymerase chain reaction （PCR）. A total of 212 clinical specimens （150 cerebrospinal fluid specimens and 62 plasma specimens） were collected from child patients,who were diagnosed or suspected with viral meningitis,and they were determined. Using nested PCR plus sequencing as reference method,the Kappa consistency analysis was performed on the SAT results. Results The sensitivity of HSV-1 RNA by SAT was 100 copies/μL. A total of 18 specimens were positive by SAT, among which 16 specimens were positive by nested PCR,and 12 specimens were positive by sequencing. The Kappa value between SAT and nested PCR plus sequencing was 0.785,and the sensitivity and specificity were 100.0% and 97.0%,respectively. Conclusions The established SAT is a sensitive,specific,accurate,rapid and reliable method for detecting HSV-1 RNA in cerebrospinal fluid and plasma,and it should be a potential method in rapid diagnosis of herpes simplex encephalitis.

Key words: Herpes simplex virus Ⅰ, Simultaneous amplification and testing, Herpes simplex encephalitis, Specificity, Sensitivity