Brown, JR;
(2017)
Next generation sequencing to understand norovirus in immunocompromised children.
Doctoral thesis , UCL (University College London).
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Abstract
Norovirus is a leading cause of gastroenteritis worldwide, causing self-limited vomiting and diarrhoea in immunocompetent people and chronic infections with significant morbidity in immunocompromised patients. Data presented in this thesis uses deep sequencing to increase our understanding of norovirus in a hospital paediatric population with a large proportion of immunocompromised patients. Real-time PCR reveals that norovirus is the most prevalent gastrointestinal virus in this population, causing infection with a higher viral titre than other gastrointestinal viruses. Norovirus is most common in immunocompromised patients and is the virus most commonly associated with chronic infections, which occur primarily in immunocompromised patients. The performance of a novel method for deep sequencing norovirus full genomes is described; this overcomes the limitations of previously published methods and achieves full genomes with >12000-fold read depth regardless of genotype or viral titre. This method is applied to sequence the complete genomes of every new norovirus case at Great Ormond Street Hospital (GOSH) over a 19 month period. Full genomes reveal a broad range of circulating genotypes, more akin to genotypes circulating in the community than those typically seen in hospitals. Phylogenetic analysis shows that the majority (69%) of cases are not acquired from another patient. This suggests multiple introductions of different norovirus strains, with limited nosocomial transmission. Full genome sequencing of longitudinally collected samples shows that chronic norovirus infections may involve super- or re-infection with a different genotype, although this does not affect the duration of infection. Deep sequencing is used to investigate changes in the norovirus intra-host mutation frequency in chronically infected immunosuppressed patients who were and were not treated with oral ribavirin, revealing a possible role for ribavirin in the treatment of chronic norovirus infections. However interpretation of in vivo data is confounded by fluctuating mutation frequencies observed over time in all patients.
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