A panel of 4 different cell lines was optimized for isolation, identification, and authentication of a varicella zoster virus from a swab sample of an 8-year-old boy suspected to varicella zoster infection. The system enabled highly efficient and rapid isolation of viruses in 33°C by serial sub culturing to more than 25 passages. The technique relies on isolation of viral genes by increasing the number of particles that are statistically represented in cell culture and verified by cell culture infectious dose 50% assay, fluorescence amidites- real time- polymerase chain reaction, and the varicella-zoster virus immediate-early protein 62 antibody in immunofluorescence test, using vaccinal Oka as attenuated varicella zoster virus golden standard. The viral genes (ORF38, ORF54) confirmed the new isolate as varicella zoster virus and revealed the amino acid sequence of viral-encoded proteins after 27 passages, identical with positive control virus, in restriction fragment length polymorphismpolymerase chain reaction test. Utilization of successive serial passages at temperatures lower than the normal body temperature would reduce the virus virulence and directly cause virus attenuation. Finally, when compared with the local isolated virus, these results were strongly confirmed. We recorded plaque forming assay to show phenotypic changing related to attenuation. Although in plaque forming assay, the size of plaque seemed smaller at first glance, the statistical distribution of the plaque size did not show any change between the virus in the first and last passages. Due to lack of access to human fetal lung cells, a new authenticated local foreskin cell substrate was used for virus cultivation. In comparison between two cell substrates, foreskin cell line - optimized and cloned- viruses replicated in vitro with kinetics that was similar to those of the human fetal lung cells. Laboratory animals that were infected with the optimized virus fluid as vaccine showed a good neutralization antibody against local varicella zoster virus isolated as compared to vaccinal Oka. These results demonstrate that the virus isolated was authenticated as varicella zoster virus, and this cold adapted attenuated virus may be an applicable candidate for future plan.
Local, VZV, Isolation, Identification, Immunological, New, Cell substrate