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Preprint in English | bioRxiv | ID: ppbiorxiv-485570


Human rotavirus (RV) vaccines used worldwide have been developed using live attenuated platforms. The recent development of a reverse genetics system for RVs has delivered the possibility of engineering chimeric viruses expressing heterologous peptides from other virus species to generate polyvalent vaccines. We tested the feasibility of this using two approaches. Firstly, we inserted short SARS-CoV-2 spike peptides into the hypervariable region of the simian SA11 RV strain viral protein (VP) 4. Secondly, we fused the receptor binding domain (RBD) of the SARS-CoV-2 spike protein, or the shorter receptor binding motif (RBM) nested within the RBD, to the C-terminus of non-structural protein (NSP) 3 of the bovine RF strain RV, with or without an intervening T2A peptide. Mutating the hypervariable region of SA11 VP4 impeded viral replication, and for these mutants no cross-reactivity with spike antibodies was detected. To rescue NSP3 mutants, we established a plasmid-based reverse genetics system for the bovine RF strain. Except for the RBD mutant, all NSP3 mutants delivered endpoint titres and replication kinetics comparable to that of the WT virus. In ELISAs, cell lysates of an NSP3 mutant expressing the RBD peptide showed cross reactivity with a SARS-CoV-2 RBD antibody. 3D bovine gut enteroids were susceptible to infection by all NSP3 mutants but only RBM mutant showed cross reactivity with SARS-CoV-2 RBD antibody. The tolerability of large peptide insertions in the NSP3 segment highlights the potential for this approach in the development of vaccine vectors targeting multiple enteric pathogens simultaneously. IMPORTANCEWe explored the use of rotaviruses (RVs) to express heterologous peptides, using SARS-CoV-2 as an exemplar. Small SARS-CoV-2 peptide insertion (<34 amino acids) into the hypervariable region of the viral protein 4 (VP4) of RV SA11 strain resulted in reduced viral titre and replication, thus limiting its use as a potential vaccine expression platform. To test RF strain for its tolerance for peptide insertions, we constructed a reverse genetics system. NSP3 was C-terminally tagged with SARS-CoV-2 spike peptides of up to 193 amino acids. With a T2A-separated 193 amino acid tag on NSP3, there was little effect on the viral rescue efficiency, titre and replication. Tagged NSP3 elicited cross-reactivity with SARS-CoV-2 spike antibodies in ELISA. This is the first report describing epitope tagging of VP4, and of a reverse genetics system for the RF strain. We highlight the potential for development of RV vaccine vectors targeting multiple enteric pathogens simultaneously.

Preprint in English | medRxiv | ID: ppmedrxiv-20219642


Identifying linked cases of infection is a key part of the public health response to viral infectious disease. Viral genome sequence data is of great value in this task, but requires careful analysis, and may need to be complemented by additional types of data. The Covid-19 pandemic has highlighted the urgent need for analytical methods which bring together sources of data to inform epidemiological investigations. We here describe A2B-COVID, an approach for the rapid identification of linked cases of coronavirus infection. Our method combines knowledge about infection dynamics, data describing the movements of individuals, and novel approaches to genome sequence data to assess whether or not cases of infection are consistent or inconsistent with linkage via transmission. We apply our method to analyse and compare data collected from two wards at Cambridge University Hospitals, showing qualitatively different patterns of linkage between cases on designated Covid-19 and non-Covid-19 wards. Our method is suitable for the rapid analysis of data from clinical or other potential outbreak settings.

Preprint in English | medRxiv | ID: ppmedrxiv-20182279


COVID-19 poses a major challenge to care homes, as SARS-CoV-2 is readily transmitted and causes disproportionately severe disease in older people. Here, 1,167 residents from 337 care homes were identified from a dataset of 6,600 COVID-19 cases from the East of England. Older age and being a care home resident were associated with increased mortality. SARS-CoV-2 genomes were available for 700 residents from 292 care homes. By integrating genomic and temporal data, 409 viral clusters within the 292 homes were identified, indicating two different patterns - outbreaks among care home residents and independent introductions with limited onward transmission. Approximately 70% of residents in the genomic analysis were admitted to hospital during the study, providing extensive opportunities for transmission between care homes and hospitals. Limiting viral transmission within care homes should be a key target for infection control to reduce COVID-19 mortality in this population. Impact statementSARS-CoV-2 can spread efficiently within care homes causing COVID-19 outbreaks among residents, who are at increased risk of severe disease, emphasising the importance of stringent infection control in this population.