Inactivation of SARS-CoV-2 in chlorinated swimming pool water

SARS-CoV-2 transmission remains a global problem which exerts a significant direct cost to public health. Additionally, other aspects of physical and mental health can be affected by limited access to social and exercise venues as a result of lockdowns in the community or personal reluctance due to safety concerns. Swimming pools have reopened in the UK as of April 12th, but the effect of swimming pool water on inactivation of SARS-CoV-2 has not yet been directly demonstrated. Here we demonstrate that water which adheres to UK swimming pool guidelines is sufficient to reduce SARS-CoV-2 infectious titre by at least 3 orders of magnitude.

Characterisation of a newly isolated SVCV strain in Ukraine

Spring Viraemia of Carp Virus (SVCV) has been well documented since the 1960s. This virus infection generally results in an acute haemorrhagic syndrome with septicaemia and associated high mortality. During a fish health inspection in carp farms in the Eastern region of Ukraine, SVCV was isolated from yearlings of the common carp Cyprinus carpio. For virus isolation and characterisation, methods of cell culture, EM, PCR, and sequencing were used. Preliminary examination of infected fish revealed a range of lesions, particularly in spleen and kidney tissues. The virus grew in fish cell lines of FHM and EPC with infectious titre of 106.2–6.5 and 106.9–7.4 TCID50/ml, respectively. Investigation by electron microscopy demonstrated that ultrastructurally the isolated virus was similar to rhabdoviruses. Virions were non-enveloped with a typical bullet profile, approximately 80 to 180 nm in length and 60 to 90 nm in diameter. In addition, the nucleotide sequence of the glycoprotein gene G fragment in size of 579 base pairs was analysed. The nucleotide sequence was registered at GenBank under the accession number MH043331. The phylogenetic analysis revealed a close relationship of East Ukrainian isolates of SVCV with sequences that represent the strains from genogroup Ib.

Attenuation and Stability of CHIKV-NoLS, a Live-Attenuated Chikungunya Virus Vaccine Candidate

Our previous investigation of the nucleolar localisation sequence (NoLS) of chikungunya virus (CHIKV) capsid protein demonstrated the role of capsid in CHIKV virulence. Mutating the NoLS of capsid in CHIKV led to the development of a unique live-attenuated CHIKV vaccine candidate, termed CHIKV-NoLS. CHIKV-NoLS-immunised mice developed long-term immunity from CHIKV infection after a single dose. To further evaluate CHIKV-NoLS attenuation and suitability as a vaccine, we examined the footpad of inoculated mice for underlying CHIKV-NoLS-induced immunopathology by histological and flow cytometric analysis. In comparison to CHIKV-WT-infected mice, CHIKV-NoLS-inoculated mice exhibited minimal inflammation and tissue damage. To examine the stability of attenuation, the plaque phenotype and replication kinetics of CHIKV-NoLS were determined following extended in vitro passage. The average plaque size of CHIKV-NoLS remained notably smaller than CHIKV-WT after extended passage and attenuated replication was maintained. To examine thermostability, CHIKV-NoLS was stored at 21 °C, 4 °C, −20 °C and −80 °C and infectious CHIKV-NoLS quantified up to 84 days. The infectious titre of CHIKV-NoLS remains stable after 56 days when stored at either −20 °C or −80 °C. Interestingly, unlike CHIKV-WT, the infectious titre of CHIKV-NoLS is not sensitive to freeze thaw cycles. These data further demonstrate preclinical safety and stability of CHIKV-NoLS.

A systematic investigation of production of synthetic prions from recombinant prion protein

According to the protein-only hypothesis, infectious mammalian prions, which exist as distinct strains with discrete biological properties, consist of multichain assemblies of misfolded cellular prion protein (PrP). A critical test would be to produce prion strains synthetically from defined components. Crucially, high-titre ‘synthetic' prions could then be used to determine the structural basis of infectivity and strain diversity at the atomic level. While there have been multiple reports of production of prions from bacterially expressed recombinant PrP using various methods, systematic production of high-titre material in a form suitable for structural analysis remains a key goal. Here, we report a novel high-throughput strategy for exploring a matrix of conditions, additives and potential cofactors that might generate high-titre prions from recombinant mouse PrP, with screening for infectivity using a sensitive automated cell-based bioassay. Overall, approximately 20 000 unique conditions were examined. While some resulted in apparently infected cell cultures, this was transient and not reproducible. We also adapted published methods that reported production of synthetic prions from recombinant hamster PrP, but again did not find evidence of significant infectious titre when using recombinant mouse PrP as substrate. Collectively, our findings are consistent with the formation of prion infectivity from recombinant mouse PrP being a rare stochastic event and we conclude that systematic generation of prions from recombinant PrP may only become possible once the detailed structure of authentic ex vivo prions is solved.

Functional importance of dengue virus maturation: infectious properties of immature virions

Prior to the release of flavivirus particles from infected cells, the viral surface protein prM is cleaved to M by the cellular enzyme furin. For dengue virus (DENV), this maturation process appears to be very inefficient since a high proportion of progeny virions contain uncleaved prM. Furthermore, it has been reported that prM-containing DENV particles are infectious. These observations contradict the general assumption that prM processing is required to render virus particles infectious. Therefore, in this study, we reinvestigated the infectious properties of immature DENV virions. DENV particles were produced in furin-deficient LoVo cells. We observed that DENV-infected LoVo cells secrete high numbers of prM-containing particles. Subsequent analysis of the infectious titre revealed that immature particles lack the ability to infect cells, the infectious unit to particle ratio being 10 000-fold reduced compared with that of wild-type virus. Our results indicate that cleavage of prM to M is required for DENV infectivity.