Pcr Combined With Restriction Digestion Analysis To Identify Herpes Virus Family Of Four Viral Infection,

single pair of oligonucleotide primer selected within a highly conserved region of the DNA polymerase gene of the herpesvirus was designed to amplify related viral genomes, i.e. herpes simplex virus type 1, herpes simplex virus type 2, human cytomegalovirus, Epstein-Barr virus, by the polymerase chain reaction. A simple restriction enzyme analysis(with SmaI and BamHI) of these amplified products allowed accurate characterization of the herpesvirus type. Cervical mucus and urine , and blood samples from 99 patients who were suspected with different hepesvirus infections were tested for the presence and identification of herpesvirus DNA by this approach. The amplifed products were visualized on ethidium bromide -stained gel with expected size: HCMV 594bp, HSV1,2 522bp, EBV 528bp. The HSV1 amplified product was cleaved by SmaI into 478bp and 44bp fragments and remained undigested by BamHI; conversely, the HSV2 amplified product was cleaved by BamHI into 224bp and 298bp fragments and remained undigested by SmaI. The EBV amplified product was cleaved by SmaI and BamHI into two fragments 101 and 427bp, 282 and 246bp, respectively. While HCMV remained undigested by both restriction enzymes. The specificity of the PCR products were confirmed by Southern blot using oligonucleotide probes specific for each of the four hepesvirus amplified products before restriction enzyme analysis. This assay was also compared with the conventional cell culture(Gold standard) combined with in situ hybridization and the specific PCR for each of the four herpesviruses, and the agreement of 85.7%, sensitivity of 100%, specificity of 92.3% , and dia-index number of 192.3% were derived. The results show that this polymerase chain reaction provides a highly sensitive and specific technique for the identification of herpesviruses DNA and should be of value for early and rapid diagnosis.