Genotyping of HCMV Glycoprotein O using Molecular Techniques

Background: Cytomegalovirus carries 6 different envelope glycoproteins on its surface all of which are essential for infectivity, spread, and pathogenicity of the virus and all are polymorphic. It has been suggested that the disproportionate effects of the virus in congenital or immunocompromised settings could be the biological effect of a particular dominant viral glycoprotein type. However, studies have not produced strong evidence for association between a particular viral glycoprotein type and disease type or outcome. A weakness of many of the previous studies is the focus on a single glycoprotein or limited combination of glycoproteins and outcome in a patient population. Recent data suggests that occurrence of particular mutations in the viral glycoproteins is driven by the type and level of the patient’s immune response. Perhaps explaining the association, but lack of robust correlation, between glycoprotein type and disease, as any patient population will contain a spectrum of host immune responses. Critical to such analyses is the accuracy of determination of the correct genotype grouping. Further investigation into the role of glycoprotein polymorphism in congenital disease requires more robust tools to investigate full glycoprotein profiles in patient groups. The aim of this study is to present a new and more accurate tool for detecting mutations in glycoprotein O (gO).Method: DNA extracted from five laboratory strains of hCMV; AD169, Towne, Davis, Toledo, and Merlin. Polymorphic sequences from gO were amplified using a novel polymerase chain reaction (PCR), followed by restriction enzyme analysis and sequencing. Results: Previous assays show 4 major supergroups for gO. Using our assay, eight distinct genotypes were identified. Conclusion: The new primer set allowed accurate identification of eight gO genotypes. Improved accuracy of genotyping is an essential pre-requisite of studies considering the role of glycoproteins in hCMV congenital disease.