The aim of the study was to find evidence of SARS-CoV-2 infection in UK cats. Design: Tissue samples were tested for SARS-CoV-2 antigen using immunofluorescence and for viral RNA by in situ hybridisation. A set of 387 oropharyngeal swabs that had been submitted for routine respiratory pathogen testing was tested for SARS-CoV-2 RNA using reverse transcriptase quantitative PCR. Results: Lung tissue collected post-mortem from cat 1 tested positive for both SARS-CoV-2 nucleocapsid antigen and RNA. SARS-CoV-2 RNA was detected in an oropharyngeal swab collected from cat 2 that presented with rhinitis and conjunctivitis. High throughput sequencing of the viral genome revealed five single nucleotide polymorphisms (SNPs) compared to the nearest UK human SARS-CoV-2 sequence, and this human virus contained eight SNPs compared to the original Wuhan-Hu-1 reference sequence. An analysis of the viral genome of cat 2 together with nine other feline-derived SARS-CoV-2 sequences from around the world revealed no shared cat-specific mutations. Conclusions: These findings indicate that human-to-cat transmission of SARS-CoV-2 occurred during the COVID-19 pandemic in the UK, with the infected cats developing mild or severe respiratory disease. Given the ability of the new coronavirus to infect different species, it will be important to monitor for human-to-cat, cat-to-cat and cat-to-human transmission.

Two cats from different COVID-19-infected households in the UK were found to be infected with SARS-CoV-2 from humans, demonstrated by immunofluorescence, in situ hybridisation, reverse transcriptase quantitative PCR and viral genome sequencing. Lung tissue collected post-mortem from cat 1 displayed pathological and histological findings consistent with viral pneumonia and tested positive for SARS-CoV-2 antigens and RNA. SARS-CoV-2 RNA was detected in an oropharyngeal swab collected from cat 2 that presented with rhinitis and conjunctivitis. High throughput sequencing of the virus from cat 2 revealed that the feline viral genome contained five single nucleotide polymorphisms (SNPs) compared to the nearest UK human SARS-CoV-2 sequence, and this human virus contained eight SNPs compared to the original Wuhan-Hu-1 reference. An analysis of the viral genome of cat 2 together with nine other feline-derived SARS-CoV-2 sequences from around the world revealed no shared cat-specific mutations. These findings indicate that human-to-cat transmission of SARS-CoV-2 occurred during the COVID-19 pandemic in the UK, with the infected cats developing mild or severe respiratory disease. Given the versatility of the new coronavirus, it will be important to monitor for human-to-cat, cat-to-cat and cat-to-human transmission.

The pandemic spread of Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2), the etiological agent of Coronavirus Disease 2019 (COVID-19), represents an ongoing international health crisis. A key symptom of SARS-CoV-2 infection is the onset of fever, with a hyperthermic temperature range of 38 to 41°C. Fever is an evolutionarily conserved host response to microbial infection that can influence the outcome of viral pathogenicity and regulation of host innate and adaptive immune responses. However, it remains to be determined what effect elevated temperature has on SARS-CoV-2 replication. Utilizing a three-dimensional (3D) air–liquid interface (ALI) model that closely mimics the natural tissue physiology of SARS-CoV-2 infection in the respiratory airway, we identify tissue temperature to play an important role in the regulation of SARS-CoV-2 infection. Respiratory tissue incubated at 40°C remained permissive to SARS-CoV-2 entry but refractory to viral transcription, leading to significantly reduced levels of viral RNA replication and apical shedding of infectious virus. We identify tissue temperature to play an important role in the differential regulation of epithelial host responses to SARS-CoV-2 infection that impact upon multiple pathways, including intracellular immune regulation, without disruption to general transcription or epithelium integrity. We present the first evidence that febrile temperatures associated with COVID-19 inhibit SARS-CoV-2 replication in respiratory epithelia. Our data identify an important role for tissue temperature in the epithelial restriction of SARS-CoV-2 independently of canonical interferon (IFN)-mediated antiviral immune defenses.

The primary envelopment/de-envelopment of Herpes viruses during nuclear exit is poorly understood. In Herpes simplex virus type-1 (HSV-1), proteins pUL31 and pUL34 are critical, while pUS3 and some others contribute; however, efficient membrane fusion may require additional host proteins. We postulated that vesicle fusion proteins present in the nuclear envelope might facilitate primary envelopment and/or de-envelopment fusion with the outer nuclear membrane. Indeed, a subpopulation of vesicle-associated membrane protein-associated protein B (VAPB), a known vesicle trafficking protein, was present in the nuclear membrane co-locating with pUL34. VAPB knockdown significantly reduced both cell-associated and supernatant virus titers. Moreover, VAPB depletion reduced cytoplasmic accumulation of virus particles and increased levels of nuclear encapsidated viral DNA. These results suggest that VAPB is an important player in the exit of primary enveloped HSV-1 virions from the nucleus. Importantly, VAPB knockdown did not alter pUL34, calnexin or GM-130 localization during infection, arguing against an indirect effect of VAPB on cellular vesicles and trafficking. Immunogold-labelling electron microscopy confirmed VAPB presence in nuclear membranes and moreover associated with primary enveloped HSV-1 particles. These data suggest that VAPB could be a cellular component of a complex that facilitates UL31/UL34/US3-mediated HSV-1 nuclear egress.

The pandemic spread of SARS-CoV-2, the etiological agent of COVID-19, represents a significant and ongoing international health crisis. A key symptom of SARS-CoV-2 infection is the onset of fever, with a hyperthermic temperature range of 38 to 41°C. Fever is an evolutionarily conserved host response to microbial infection and inflammation that can influence the outcome of viral pathogenicity and regulation of host innate and adaptive immune responses. However, it remains to be determined what effect elevated temperature has on SARS-CoV-2 tropism and replication. Utilizing a 3D air-liquid interface (ALI) model that closely mimics the natural tissue physiology and cellular tropism of SARS-CoV-2 infection in the respiratory airway, we identify tissue temperature to play an important role in the regulation of SARS-CoV-2 infection. We show that temperature elevation induces wide-spread transcriptome changes that impact upon the regulation of multiple pathways, including epigenetic regulation and lncRNA expression, without disruption of general cellular transcription or the induction of interferon (IFN)-mediated antiviral immune defences. Respiratory tissue incubated at temperatures >37°C remained permissive to SARS-CoV-2 infection but severely restricted the initiation of viral transcription, leading to significantly reduced levels of intraepithelial viral RNA accumulation and apical shedding of infectious virus. To our knowledge, we present the first evidence that febrile temperatures associated with COVID-19 inhibit SARS-CoV-2 replication. Our data identify an important role for temperature elevation in the epithelial restriction of SARS-CoV-2 that occurs independently of the induction of canonical IFN-mediated antiviral immune defences and interferon-stimulated gene (ISG) expression.

Persistent infection with oncogenic human papillomavirus (HPV) is a prerequisite for cervical disease development, yet data regarding the host immune response to infection at the genotype level are quite limited. We created pseudoviruses bearing the major (L1) and minor (L2) capsid proteins and L1 virus-like particles representing the reference sequence and a consensus of 34 European sequences of HPV51. Despite the formation of similarly sized particles, motifs in the reference L1 and L2 genes had a profound impact on the immunogenicity, antigenicity and infectivity of these antigens. The antibody status of women exhibiting low-grade disease was similar between HPV16 and the consensus HPV51, but both demonstrated discrepancies between binding and neutralizing antibody responses. These data support the use of pseudoviruses as the preferred target antigen in studies of natural HPV infection and the need to consider variation in both the L1 and L2 proteins for the appropriate presentation of antibody epitopes. Received 5 November 2014 Accepted 12 March 2015Persistent infection with one or more of about a dozen human papillomavirus (HPV) genotypes (Bouvard et al., 2009) is associated with the development of cervical cancer, a significant cause of morbidity and mortality in women worldwide (Schiffman et al., 2007). The propensity for certain genotypes to persist longer than others (Rositch et al., 2013) may contribute to the observed HPV genotype distribution differences between low-grade disease and cervical cancer (Guan et al., 2012).The non-enveloped capsid of HPV comprises the major (L1) and minor (L2) capsid proteins. The L1 protein facilitates virus attachment to host cells, whilst the L2 protein is essential for subsequent virus infectivity (Buck et al., 2013;Raff et al., 2013;Wang & Roden, 2013).Current L1 virus-like particle (VLP)-based HPV vaccines target the most prevalent genotypes, HPV16 and HPV18, and elicit type-specific neutralizing antibodies thought to confer the high levels of efficacy observed in clinical trials (Lehtinen & Dillner, 2013;Schiller et al., 2012). A limited number of serological assays are available for measuring vaccine type-specific antibody responses, including an L1 VLP ELISA, a mAb competitive VLP assay and an L1L2 pseudovirus neutralization assay. Despite some discrepancies, overall inter-assay agreements are good (Dessy et al., 2008;Krajden et al., 2014). Natural infection (NI) studies usually compare the HPV DNA status of individuals with their respective antibody status in order to better understand the pathogen-host relationship. Most of these studies have examined HPV16 and/or HPV18 (Castellsagué et al., 2014;Lin et al., 2013;Pastrana et al., 2004; Safaeian et al., 2010;Xi et al., 2002), whilst some have expanded their investigations to evaluate other genotypes including HPV6, HPV11, HPV31, HPV33, HPV35, HPV45, HPV52 and HPV58 (Carter et al., 2000;Ochi et al., 2008; Syrjänen et al., 2009;Wilson et al., 2013). Most of these studies have made use of an immobilized L1-based t...

High-risk human papillomavirus (HPV) is the causative agent of benign, precancerous and cancerous lesions, in both anogenital and oropharyngeal sites. Increased expression of the viral oncoproteins E6 and E7 are responsible for tumour progression. The treatment of these precancerous and cancerous lesions is invasive, painful and with long-term side effects. Localised microwaves have been used successfully in the clinic for the treatment of verrucas, which are caused by low-risk HPV genotypes (>75% success rate versus >33% for cryotherapy). Moreover, local hyperthermia is known to have anti-tumour effects. Ten-second microwave treatment of 3D in vitro-grown cervical tumour tissues (HPV16-positive SiHa cell) resulted in cell death in the treated zone while the tissue integrity was disrupted in the adjacent area. Microwaves induced apoptosis (induction of cleaved caspase 3) and autophagy (induction of LC3) and inhibited cell proliferation (loss of Ki67 and MCM2) in the entire tissue. Furthermore, HPV16 E6 and E7 expression was reduced in cells in the treated and transition zones, with subsequent induction of expression of the apoptosis-regulator, p53 over a 24 hour period following microwave treatment. Thermal stress, identified with the Heat Shock Protein 70 (HSP70) and translational stress identified by G3BP expression, was observed in the transition zone. In conclusion, we demonstrate that the microwave treatment induces cell stress pathways and inhibits HPV oncoprotein expression that causes tumour progression. Induction of apoptosis and reduced cell proliferation suggest a reversal of the cervical tumour phenotype in the 3D tissues.

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