Phenotypic and genotypic methods for the detection of herpes simplex virus serotypes.
Typing of herpes simplex virus (HSV) into its serotypes plays a major role in epidemiology and management of reactivation. To develop and evaluate a polymerase chain reaction (PCR)-based restriction fragment length polymorphism (RFLP) was employed using Hae III and Taq I against neutralization test, allele-specific PCR and DNA sequencing for the detection of HSV serotypes. Neutralization test, allele-specific PCR, DNA sequencing and PCR-based RFLP were applied simultaneously to 2 standard strains (HSV-1 and HSV-2) and 23 clinical isolates. PCR-based RFLP was applied further to 20 culture negative PCR positive clinical specimens. The 179 bp product of the clinical isolates and specimens amplified using the type-common primers of HSV was subjected to DNA sequencing and PCR-based RFLP. Allele-specific PCR was absolutely specific and highly sensitive. All the typing methods differentiated concordantly 23 clinical isolates into 12 HSV-1 and 11 HSV-2. DNA sequencing did not reveal any nucleotide variations within the serotypes among the isolates sequenced. PCR-based RFLP typed a further 20 culture negative clinical specimens into 15 HSV-1 and 5 HSV-2. PCR-based RFLP was a reliable, less laborious and cost-effective molecular biological tool for the determination of HSV serotypes both for the clinical isolates and culture negative specimens.
Madhavan
H N
HN
L&T Microbiology Research Centre, Vision Research Foundation, Sankara Nethralaya, 18, College Road, Chennai 600 006, India. drhnm@sankaranethralaya.org
Priya
K
K
Bagyalakshmi
R
R
D000483:Alleles; D001483:Base Sequence; D017931:DNA Primers / Q000235:genetics; D005260:Female; D005838:Genotype; D006561:Herpes Simplex / Q000821:virology; D018259:Herpesvirus 1, Human / Q000145:classification / Q000235:genetics / Q000302:isolation & purification; D018258:Herpesvirus 2, Human / Q000145:classification / Q000235:genetics / Q000302:isolation & purification; D006801:Humans; D008297:Male; D008969:Molecular Sequence Data; D010641:Phenotype; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length; D012689:Sequence Homology, Nucleic Acid; D012703:Serotyping / Q000379:methods; D018139:Simplexvirus / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D014773:Virology / Q000379:methods*
D017931:DNA Primers