Determination of the six major human herpesviruses in cerebrospinal fluid and blood specimens of children

Abstract

Aim: To detect and differentiate six major human herpesviruses in the cerebrospinal fluid (CSF) and blood of children by polymerase chain reaction restriction fragment length polymorphism (PCR‐RFLP). Methods: We synthesized two pairs of primers in the well‐conserved regions of the DNA polymerase gene in human herpesviruses. One pair was designed to amplify cytomegalovirus (CMV), Epstein‐Barr virus (EBV), herpes simplex virus type 1 (HSV‐1) and type 2 (HSV‐2), and the other pair to amplify varicella‐zoster virus (VZV) and human herpesvirus 6 (HHV‐6) by PCR. Virus species identification was achieved by restriction enzyme digestion with BamHI and BstUI. Ninety‐eight CSF and 75 blood specimens were analysed by this technique. At the same time, all blood specimens were also examined by enzyme‐linked immunosorbent assay (ELISA). Results: Thirteen (13.3%) of 98 CSF specimens and 26 (34.7%) of 75 blood specimens were positive for herpesvirus DNA in this PCR assay. Only 10 (13.3%) of the blood specimens were positive in ELISA for virus‐IgM antibody. Sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of PCR in detecting herpesvirus infections compared with ELISA were 100% (10/10), 75.4% (49/65), 38.5% (10/26) and 100% (49/49), respectively. These results indicate that the positive rate of PCR was significantly higher than that of ELISA (p<0.05). The herpesvirus type of these positive specimens was rapidly detected using restriction enzyme digestion with BamHI and BstUI. Conclusions: PCR‐RFLP is a specific, sensitive and accurate technique for the identification of herpesvirus infections in the CSF and blood of children.

Fourth and Sixth Nerve Palsies Due to Herpes Simplex 1 Infection

“…PCR has become the gold standard for the diagnosis of HSV1 infections even in the absence of serum IgM antibodies. In part, this is because the rise of IgM antibodies occurs in a narrow time window and shows a lower sensitivity than PCR (13)(14)(15). Serum examination in our patient was performed 12 days after symptom onset, beyond the optimal detection period for IgM antibodies.…”

Rapid diagnosis of herpetic encephalitis in children by PCR-microarray technology for simultaneous detection of seven human herpes viruses

“…Thereby, we simultaneously detected and genotyped the seven herpes viruses. There are also PCR-restriction fragment length polymorphism analysis technology can detect multiple human herpes viruses simultaneously [1,2], but the following-up process of this method is cumbersome and easy to contaminate, it is not suitable for large-scale application in clinical detection.…”

Detection of the four major human herpesviruses simultaneously in whole blood and cerebrospinal fluid samples by the fluorescence polarization assay

“…However, conventional detection methods for HSV-1/-2, CMV, and EBV are not used in routine practice due to the complex experimental process and high costs. [2][3][4][5][6] A few fluorescence polarization (FP) assays have been used as the diagnostic method for the detection of particular organisms. 7,8 The FP assay is simple as it can directly detect the hybridization between the fluorescent-labeled probes and the complementary single-stranded DNA in polymerase chain reaction (PCR) products.…”