ABSTRACT
It was determined the ratio of viral DNA and DNA from Vero cells using the polymerase chain reaction in real time in Vero cell lysate, infected with L2 strain of the herpes simplex virus type 1. Copy number of the virus reached a maximum after 24 hours of incubation of infection. Total DNA was isolated and sequenced using NGS technology by Ion Torrent device. Nucleotide sequences of the thymidine kinase gene (UL23) and DNA polymerase (UL30) were determined for a population of HSV-1 strain L2. Comparison of the primary structure of these genes with the corresponding nucleotide sequences of known strains of HSV-1 KOS and 17 was conducted. Differences in the structure of genes UL23 and UL30 between strain L2 and reference strains KOS and 17 are not important, because changes are found in non-conservative regions.
Subject(s)
DNA-Directed DNA Polymerase/genetics , Exodeoxyribonucleases/genetics , Herpesvirus 1, Human/genetics , Thymidine Kinase/genetics , Viral Proteins/genetics , Animals , Chlorocebus aethiops , Polymerase Chain Reaction , Sequence Analysis, DNA/methods , Vero Cells/virologyABSTRACT
Cloned laboratory mutants of herpes simplex virus type I resistant to acycloguanosine H-phosphonate have been investigated. For all clones were shown that mutations resulted to increasing of sensitivity to acting of sidofovir. Thymidine kinase of mutant viruses partially preserves the ability to phosphorilate thymidine, but loses the ability to phosphorilate BVDU.
Subject(s)
Acyclovir/analogs & derivatives , DNA-Directed DNA Polymerase/metabolism , Drug Resistance, Viral/drug effects , Herpesvirus 1, Human/drug effects , Herpesvirus 1, Human/enzymology , Organophosphonates/pharmacology , Thymidine Kinase/chemistry , Acyclovir/pharmacology , Animals , DNA-Directed DNA Polymerase/genetics , Humans , Mutation , Thymidine Kinase/genetics , Vero CellsABSTRACT
The primary structures of DNA-polymerase (ul30) and thymidine kinase (ul23) genes from several herpes simplex virus type 1 (HSV-1) clinical isolates which differed in sensitivity for a number of antiherpetic drugs were determined and compared with those for two laboratory HSV-1 strains one of which was ACV-sensitive (L2), while the another was resistant (L2) to ACV. The phylogenetic analysis of the sequences showed that conserved regions of ul30 gene of HSV-1 clinical isolates and L2 strain were homologous with the exception of point mutations and degenerated substitutions. Several new mutations in the HSV-1 DNA-polymerase and thymidine kinase functional domains were established and identified as the substitutions associated with the strain-resistance to ACV and other drugs.