Seroepidemiology and detection of human group C rotaviruses

James, Vivienne Lesley Anderson (1998) Seroepidemiology and detection of human group C rotaviruses. University of Southampton, Doctoral Thesis.

Abstract

The gene coding for the major inner capsid protein VP6 of human group C rotavirus was expressed in insect cells. VP6 was expressed at high level in High Five^TM cells and was exported to the cell culture medium. The recombinant protein formed virus like particles and honeycomb structures. Purified VP6 was used to immunise rabbits and laying hens. Rabbit serum, antibodies extracted from hens eggs and recombinant VP6 were used to develop and optimise ELISAs to detect human group C rotavirus antigens and antibodies. The ELISAs were used to determine the extent of infection due to human group C rotaviruses. Faecal samples (573) from cases of acute gastro-enteritis were tested in the antigen detection ELISA; 1.6% were positive, comparable to the incidence of both astroviruses and classical caliciviruses. Positive results were confirmed by EM, PAGE of dsRNA or RT-PCR developed specifically to detect the human group C rotavirus VP6 gene. The ELISA was more sensitive than EM and PAGE. The ELISA to detect IgG antibodies in serum was shown to be specific for group C rotavirus. In a local seroepidemiological survey of 1000 sera grouped by age 43% of samples were found to have antibodies to human group C rotavirus with the highest proportion (59%) in the 61+ age group. These results suggest that infection with human group C rotaviruses is common.

Genes 6 and 7 of the 'Bristol' strain of human group C rotavirus were sequenced from M13 clones containing full length inserts. Sequence data were confirmed by direct sequencing of PCR-amplified cDNA from a faecal sample containing human group C rotavirus. This work completed the nucleotide sequence determination and coding assignments of human group C rotavirus.

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