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Herpes simplex virus genotyping: multiple optional PCR-based RFLP systems and a non-isotopic single-strand
Martín C Abba1, Carlos D Golijow
1Facultad de Ciencias Veterinarias, Centro de Investigaciones en Genética Básica y Aplicada (CIGEBA), Universidad Nacional de La Plata, Calle 60 y 118 s/n, B1900AVW, La Plata, Buenos Aires, Argentina. mabba777@hotmail.com
Journal of Virological Methods
|May 26, 2004
Summary
Two polymerase chain reaction (PCR) methods, PCR-SSCP and PCR-RFLP, effectively genotype herpes simplex virus (HSV) types I and II. These techniques are reliable for clinical labs and identified HSV-2 as the predominant type in cervical samples.
Area of Science:
- Molecular Biology
- Virology
- Clinical Diagnostics
Background:
- Herpes Simplex Virus (HSV) types I and II are significant human pathogens.
- Accurate genotyping of HSV is crucial for epidemiological studies and clinical management.
- Existing genotyping methods may require optimization for clinical laboratory settings.
Purpose of the Study:
- To evaluate and compare two Polymerase Chain Reaction (PCR)-based genotyping techniques for HSV types I/II: PCR-Single Strand Conformation Polymorphism (PCR-SSCP) and PCR-Restriction Fragment Length Polymorphism (PCR-RFLP).
- To assess the prevalence of HSV types I/II in a cohort of normal cervical samples.
Main Methods:
- Genomic DNA from standard HSV strains (KOS and G) and 79 normal cervical samples were analyzed.
- PCR-SSCP was employed to detect conformational differences in DNA strands.
- PCR-RFLP utilized sequence-specific restriction enzyme digestion of PCR products for genotyping.
Main Results:
- Sequence analysis identified 19 potential RFLP systems for HSV typing; five were validated in this study.
- PCR-SSCP demonstrated distinct patterns for HSV-1 and HSV-2.
- PCR-SSCP and PCR-RFLP showed 100% concordance in genotyping results.
- HSV was detected in 21.5% (17/79) of cervical samples, with HSV-2 being the most prevalent type (71% of infections).
Conclusions:
- Both PCR-SSCP and PCR-RFLP are highly accurate and reliable methods for HSV types I/II genotyping.
- These PCR-based techniques are suitable for routine use in clinical laboratories due to their ease of performance.
- The study highlights the significant prevalence of HSV-2 in the studied population.