Cytomegalovirus Glycoprotein Types and Disease Causation

UoM administered thesis: Phd

  • Authors:
  • Zahrah Buhamad

Abstract

Human Cytomegalovirus (HCMV) is the most common cause of viral congenital infection in the world. Around 5-10% of HCMV infected children are symptomatic at birth, and 50-90% of these develop severe manifestations with a 30% mortality rate. Among the asymptomatic children at birth, 10-15% develop late sequelae. The major cell entry glycoproteins of HCMV form three complexes: gC-I containing gB; gC-II containing gM & gN; and gC-III containing gH, gL, and gO (or UL128-131). These entry glycoproteins are polymorphic, producing different glycoprotein genotypes. The polymorphic nature of the glycoproteins as well as their ability to elicit neutralizing antibodies made them of interest in correlating them with the severity and outcome of the disease. This study aimed to develop a robust system to identify clusters of glycoprotein genotypes and to correlate them with disease manifestation. PCR assays of high sensitivity were used to identify all six glycoproteins. The PCR products were digested using restriction enzymes (RFLP) to identify the glycoprotein genotypes. Available laboratory strains (AD169, Towne, Davis, Toledo, and Merlin) as well as 112 clinical samples were amplified and genotyped using the assay, and their glycoprotein genotype profile was determined. A reliable sensitive assay was successfully developed to identify all glycoprotein genotypes including a novel gM assay using PCR/RFLP. The clinical samples were grouped according to disease manifestation (Group 1: congenital/postnatal patients (subgroup 1A: confirmed congenital patients & subgroup 1B: patients with either congenital or postnatal infection), Group 2: immunocompetent patients, Group 3: immunocompromised patients (subgroup 3A: immunocompromised patients with primary infection, subgroup 3B: immunocompromised patients with recurrent infection & subgroup 3C: immunocompromised patients with unconfirmed primary or recurrent infection)). Genotype gB1 was found predominantly prevalent in congenital/postnatal and immunocompromised patients, while gB3 was the most common genotype in immunocompetent patients. This result along with the phylogenetic analysis performed in this study suggest a relationship between gB genotypes and the immune response of the patients, where gB3 may be positively selected by host immune pressure. The novel gM assay genotyped the highly conserved gene (UL100) into three distinct genotypes; gM3 genotype associated with the congenital/postnatal group; which may provide an insight into understanding viral attachment and spread into the host cell. In congenital/postnatal infection, gH1 (72.7%) and gL4 (65.1%) were the most prevalent genotypes (gH1= 32/44, gL4= 28/43; P=0.000). In immunocompetent patients, mixed gH and mixed gL genotypes significantly correlated with the group, and in the immunocompromised group gH2 and mixed gL genotype were the most common genotypes (51.1% and 46.9% respectively). Glycoproteins gO, gH and gL are components of gC-III complex and gO1 was found to be the most prevalent gO genotype in all infection types (Group 1= 32.1%, Group 2= 85.7%, Group 3= 18.8%; P

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Original languageEnglish
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Award date1 Aug 2018