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Continued global outbreaks of H5Nx highly pathogenic avian influenza viruses (HPAIV) have been reported in poultry since the emergence of the Asian Goose/Guangdong lineage of HPAIV H5N1 in 1996. Consequently, vaccines have been developed and employed to protect commercial and non-commercial poultry flocks. However, constant changes in virus immunogenetics makes vaccine candidates that prevent morbidity, mortality, reduce shedding, and prevent transmission a moving target. Here, we tested the effects of disrupting the mRNA splice sites and thus expression of either of the two isoforms of the viral ion channel M2 and M42 expressed by the H5N2 low pathogenic avian influenza virus (LPAIV) progenitor of the 1983 Pennsylvania HPAIV outbreak. Both G52C (ΔM2) and G145A (ΔM42) versions of the A/chicken/Pennsylvania/1/83 (Ck/Penn) virus replicated wellin vitroandin ovo, but showed altered virion morphology, with the G145A virus exhibiting filamentous budding. The G52C and G145A viruses also infected chickens efficiently and stimulated robust immune responses; however, unlike the wild type virus, they did not transmit to naïve-contact birds. In protection studies, vaccination of birds with G52C or G145A viruses protected 100% of birds from lethal challenge with homologous and distant heterologous H5 HPAIV strains from North American and Asian lineages and significantly reduced virus shedding compared to controls. Furthermore, the live virus vaccines decreased virus shedding 10,000-fold more than corresponding inactivated forms of the vaccine virus. Taken together, these studies demonstrate a strategy for developing a non-transmittable but highly protective live attenuated virus for AIV.

Angiotensin converting enzyme 2 (ACE2) is a host cell membrane protein (receptor) that mediates the binding of coronavirus, most notably SARS coronaviruses in the respiratory and gastrointestinal tracts. Although SARS-CoV-2 infection is mainly confined to humans, there have been numerous incidents of spillback (reverse zoonoses) to domestic and captive animals. An absence of information on the spatial distribution of ACE2 in animal tissues limits our understanding of host species susceptibility. Here, we describe the distribution of ACE2 using immunohistochemistry (IHC) on histological sections derived from carnivores, ungulates, primates and chiroptera. Comparison of mink (Neovison vison) and ferret (Mustela putorius furo) respiratory tracts showed substantial differences, demonstrating that ACE2 is present in the lower respiratory tract of mink but not ferrets. The presence of ACE2 in the respiratory tract in some species was much more restricted as indicated by limited immunolabelling in the nasal turbinate, trachea and lungs of cats (Felis catus) and only the nasal turbinate in the golden Syrian hamster (Mesocricetus auratus). In the lungs of other species, ACE2 could be detected on the bronchiolar epithelium of the sheep (Ovis aries), cattle (Bos taurus), European badger (Meles meles), cheetah (Acinonyx jubatus), tiger and lion (Panthera spp.). In addition, ACE2 was present in the nasal mucosa epithelium of the serotine bat (Eptesicus serotinus) but not in pig (Sus scrofa domestica), cattle or sheep. In the intestine, ACE2 immunolabelling was seen on the microvillus of enterocytes (surface of intestine) across various taxa. These results provide anatomical evidence of ACE2expression in a number of species which will enable further understanding of host susceptibility and tissue tropism of ACE2 receptor-mediated viral infection

There is need for effective and affordable vaccines against SARS-CoV-2 to tackle the ongoing pandemic. In this study, we describe a protein nanoparticle vaccine against SARS-CoV-2. The vaccine is based on the display of coronavirus spike glycoprotein receptor-binding domain (RBD) on a synthetic virus-like particle (VLP) platform, SpyCatcher003-mi3, using SpyTag/SpyCatcher technology. Low doses of RBD-SpyVLP in a prime-boost regimen induce a strong neutralising antibody response in mice and pigs that is superior to convalescent human sera. We evaluate antibody quality using ACE2 blocking and neutralisation of cell infection by pseudovirus or wild-type SARS-CoV-2. Using competition assays with a monoclonal antibody panel, we show that RBD-SpyVLP induces a polyclonal antibody response that recognises key epitopes on the RBD, reducing the likelihood of selecting neutralisation-escape mutants. Moreover, RBD-SpyVLP is thermostable and can be lyophilised without losing immunogenicity, to facilitate global distribution and reduce cold-chain dependence. The data suggests that RBD-SpyVLP provides strong potential to address clinical and logistic challenges of the COVID-19 pandemic., Vaccines for SARS-COV-2 are needed in the ongoing pandemic. Here the authors characterize a vaccine candidate that presents the receptor-binding domain (RBD) of SARS-CoV-2 spike protein on a synthetic VLP platform using SpyTag/SpyCatcher technology and show immunogenicity of a prime-boost regimen in mice and pigs.

The Coronavirus Disease 2019 (COVID-19) pandemic, caused by SARS Coronavirus 2 (SARS-CoV-2), continues to cause significant mortality in human populations worldwide. SARS-CoV-2 has high sequence similarity to SARS-CoV and other related coronaviruses circulating in bats. It is still unclear whether transmission occurred directly from bats to humans, or through an intermediate host, bringing into question the broader host range of SARS-CoV-2. Using a combination of low biocontainment entry assays as well as live virus, we explored the receptor usage of SARS-CoV-2 using angiotensin-converting enzyme 2 (ACE2) receptors from 22 different species. We demonstrated that in addition to human ACE2, the Spike of SARS-CoV-2 has broad tropism for other mammalian ACE2s, including dog, cat and cattle. However, comparison of SARS-CoV-2 receptor usage to the related SARS-CoV and bat coronavirus, RaTG13, identified distinct patterns of receptor usage, with the two human viruses being more closely aligned. Finally, using bioinformatics, structure analysis and targeted mutagenesis, we identified key residues at the Spike-ACE2 interface which may have played a pivotal role in the emergence of SARS-CoV-2 in humans, some of which are also mutated in newly circulating variants of the virus. To summarise, the broad tropism of SARS-CoV-2 at the point of viral entry identifies the potential risk of infection of a wide range of companion animals, livestock and wildlife.

Introdução: O Acidente com Material biológico (ATMB) é considerado uma urgência médica, pois existe risco de transmissão de patógenos, como hepatite B (HBV), hepatite C (HCV) e HIV. A profilaxia pós-exposição (PEP) iniciada com brevidade terá uma maior eficácia em evitar um soro conversão para hepatite B e HIV em função disso, a primeira consulta imediatamente após o ATMB tem dois objetivos principais: avaliar o risco e, consequentemente, determinar a conduta imediata para minimizar o risco de contaminação. No entanto, o que todos almejamos é uma redução no número dos acidentes com ATMB. Objetivo: Avaliar a incidência de acidentes com material biológico em profissionais de saúde, durante a pandemia comparada com os anos anteriores. Método: Estudo de coorte retrospectivo, dos profissionais de saúde que foram avaliados com acidentes registrados de 1º de janeiro de 2018 a dezembro 2020 atendidos no Serviço de Medicina Ocupacional. Resultados: Em 2018 ocorreram 197, sendo 112 em funcionários e 85 entre os residentes. Em 2019, 187 registros, sendo 117 em funcionários e 70 entre residentes. Em 2020 foram registrados apenas 158 acidentes, 117 em funcionários e 41 em residentes. Discussão e conclusão: Observamos uma redução significativa no número total de 209 acidentes, sendo maior entre os residentes. Seria resultante do impacto da introdução de materiais com dispositivos de segurança ou apenas resultado da redução dos procedimentos invasivos durante a pandemia. Desta forma mantemos divulgação e capacitação dos profissionais de saúde quanto aos procedimentos de prevenção e exposição com material biológico, repetido regularmente a fim de se formar uma consciência prevencionista

Vaccines against SARS-CoV-2 are urgently required, but early development of vaccines against SARS-CoV-1 resulted in enhanced disease after vaccination. Careful assessment of this phenomena is warranted for vaccine development against SARS CoV-2. Here we report detailed immune profiling after ChAdOx1 nCoV-19 (AZD1222) and subsequent high dose challenge in two animal models of SARS-CoV-2 mediated disease. We demonstrate in rhesus macaques the lung pathology caused by SARS-CoV-2 mediated pneumonia is reduced by prior vaccination with ChAdOx1 nCoV-19 which induced neutralising antibody responses after a single intramuscular administration. In a second animal model, ferrets, ChAdOx1 nCoV-19 reduced both virus shedding and lung pathology. Antibody titre were boosted by a second dose. Data from these challenge models on the absence of enhanced disease and the detailed immune profiling, support the continued clinical evaluation of ChAdOx1 nCoV-19., Lambe, Spencer, Thomas, Gilbert and colleagues report on the detailed immune profile of rhesus macaques and ferrets vaccinated against SARS-CoV-2 under high dose challenge. Their findings indicate that the ChAdOx1 nCoV-19 (AZD1222) the vaccine induces immune responses and reduces disease symptoms in both models, including SARS-CoV-2 mediated pneumonia and virus shedding.

The enveloped morbilliviruses utilise conserved proteinaceous receptors to enter host cells: SLAMF1 or Nectin-4. Receptor binding is initiated by the viral attachment protein Haemagglutinin (H), with the viral Fusion protein (F) driving membrane fusion. Crystal structures of the prototypic morbillivirus measles virus H with either SLAMF1 or Nectin-4 are available and have served as the basis for improved understanding of this interaction. However, whether these interactions remain conserved throughout the morbillivirus genus requires further characterisation. Using a random mutagenesis approach, based on error-prone PCR, we targeted the putative receptor binding site for SLAMF1 interaction on peste des petits ruminants virus (PPRV) H, identifying mutations that inhibited virus-induced cell-cell fusion. These data, combined with structural modelling of the PPRV H and ovine SLAMF1 interaction, indicate this region is functionally conserved across all morbilliviruses. Error-prone PCR provides a powerful tool for functionally characterising functional domains within viral proteins.

Background The spread of SARS-CoV-2 has caused a worldwide pandemic that has affected almost every aspect of human life. The development of an effective COVID-19 vaccine could limit the morbidity and mortality caused by infection and may enable the relaxation of social-distancing measures. Age is one of the most significant risk factors for poor health outcomes after SARS-CoV-2 infection; therefore, it is desirable that any new vaccine candidates elicit a robust immune response in older adults. Methods Here, we use in-depth immunophenotyping to characterize the innate and adaptive immune response induced upon intramuscular administration of the adenoviral vectored ChAdOx1 nCoV-19 (AZD-1222) COVID-19 vaccine candidate in mice. Findings A single vaccination generates spike-specific Th1 cells, Th1-like Foxp3+ regulatory T cells, polyfunctional spike-specific CD8+ T cells. and granzyme-B-producing CD8 effectors. Spike-specific IgG and IgM are generated from both the early extrafollicular antibody response and the T follicular helper cell-supported germinal center reaction, which is associated with the production of virus-neutralizing antibodies. A single dose of this vaccine generated a similar type of immune response in aged mice but of a reduced magnitude than in younger mice. We report that a second dose enhances the immune response to this vaccine in aged mice. Conclusions This study shows that ChAdOx1 nCoV-19 induces both cellular and humoral immunity in adult and aged mice and suggests a prime-boost strategy is a rational approach to enhance immunogenicity in older persons. Funding This study was supported by BBSRC, Lister institute of Preventative Medicine, EPSRC VaxHub, and Innovate UK., Graphical Abstract, Highlights ChAdOx1 nCoV-19 induces spike-specific polyfunctional CD8+ and CD4+ T cells ChAdOx1 nCoV-19 stimulates extrafollicular plasma cell and germinal center formation A single dose of ChAdOx1 nCoV-19 induces cellular and humoral immunity in aged mice A booster dose of ChAdOx1 nCoV-19 enhances immunogenicity in aged mice, Context and Significance Effective COVID-19 vaccines will play a central role in the exit strategy from the worldwide pandemic. However, older persons often do not generate protective immunity upon vaccination due to age-dependent changes in their immune system. Because older people are more likely to have poor clinical outcomes after SARS-CoV-2 infection, vaccine strategies that elicit an optimal immune response in older bodies are urgently required. Researchers from the Babraham Institute , the Jenner Institute, and the Pirbright Institute in the UK performed pre-clinical testing of the COVID-19 vaccine candidate ChAdOx1 nCoV-19 in aged mice to assess how aging influences the immune response to this vaccine. The results show that a “prime-boost” vaccine regime enhances immunogenicity in aged mice, indicating that this approach is a rational strategy for vaccinating older persons., Older persons are more likely to have poor health outcomes after SARS-CoV-2 infection; therefore, they are most in need of effective COVID-19 vaccines. Silva-Cayetano et al. show that a prime-boost strategy of the COVID-19 vaccine ChAdOx1 nCoV-19 enhances immunogenicity in aged mice, suggesting this as a strategy for immunizing older people.

The COVID-19 pandemic, caused by SARS-CoV-2 coronavirus, is a global health issue with unprecedented challenges for public health. SARS-CoV-2 primarily infects cells of the respiratory tract via Spike glycoprotein binding to angiotensin-converting enzyme (ACE2). Circadian rhythms coordinate an organism’s response to its environment and can regulate host susceptibility to virus infection. We demonstrate that silencing the circadian regulator Bmal1 or treating lung epithelial cells with the REV-ERB agonist SR9009 reduce ACE2 expression and inhibit SARS-CoV-2 entry and replication. Importantly, treating infected cells with SR9009 limits SARS-CoV-2 replication and secretion of infectious particles, showing that post-entry steps in the viral life cycle are influenced by the circadian system. Transcriptome analysis revealed that Bmal1 silencing induced interferon stimulated gene transcripts in Calu-3 lung epithelial cells, providing a mechanism for the circadian pathway to limit SARS-CoV-2 infection. Our study highlights alternative approaches to understand and improve therapeutic targeting of SARS-CoV-2., Graphical Abstract

SARS Coronavirus 2 (SARS-CoV-2) emerged in late 2019, leading to the Coronavirus Disease 2019 (COVID-19) pandemic that continues to cause significant global mortality in human populations. Given its sequence similarity to SARS-CoV, as well as related coronaviruses circulating in bats, SARS-CoV-2 is thought to have originated in Chiroptera species in China. However, whether the virus spread directly to humans or through an intermediate host is currently unclear, as is the potential for this virus to infect companion animals, livestock, and wildlife that could act as viral reservoirs. Using a combination of surrogate entry assays and live virus, we demonstrate that, in addition to human angiotensin-converting enzyme 2 (ACE2), the Spike glycoprotein of SARS-CoV-2 has a broad host tropism for mammalian ACE2 receptors, despite divergence in the amino acids at the Spike receptor binding site on these proteins. Of the 22 different hosts we investigated, ACE2 proteins from dog, cat, and cattle were the most permissive to SARS-CoV-2, while bat and bird ACE2 proteins were the least efficiently used receptors. The absence of a significant tropism for any of the 3 genetically distinct bat ACE2 proteins we examined indicates that SARS-CoV-2 receptor usage likely shifted during zoonotic transmission from bats into people, possibly in an intermediate reservoir. Comparison of SARS-CoV-2 receptor usage to the related coronaviruses SARS-CoV and RaTG13 identified distinct tropisms, with the 2 human viruses being more closely aligned. Finally, using bioinformatics, structural data, and targeted mutagenesis, we identified amino acid residues within the Spike–ACE2 interface, which may have played a pivotal role in the emergence of SARS-CoV-2 in humans. The apparently broad tropism of SARS-CoV-2 at the point of viral entry confirms the potential risk of infection to a wide range of companion animals, livestock, and wildlife., A study using a combination of surrogate entry assays and live virus suggests that SARS-CoV-2 may have a broad host-range, revealing that the virus's spike protein can use a broad range of host ACE2 receptors to enter cells and that the sequence of this protein might have changed during the zoonotic jump into humans.

Although enveloped viruses canonically mediate particle entry through virus–cell fusion, certain viruses can spread by cell–cell fusion, brought about by receptor engagement and triggering of membrane-bound, viral-encoded fusion proteins on the surface of cells. The formation of pathogenic syncytia or multinucleated cells is seen in vivo, but their contribution to viral pathogenesis is poorly understood. For the negative-strand paramyxoviruses respiratory syncytial virus (RSV) and Nipah virus (NiV), cell–cell spread is highly efficient because their oligomeric fusion protein complexes are active at neutral pH. The recently emerged severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has also been reported to induce syncytia formation in infected cells, with the spike protein initiating cell–cell fusion. Whilst it is well established that fusion protein-specific antibodies can block particle attachment and/or entry into the cell (canonical virus neutralization), their capacity to inhibit cell–cell fusion and the consequences of this neutralization for the control of infection are not well characterized, in part because of the lack of specific tools to assay and quantify this activity. Using an adapted bimolecular fluorescence complementation assay, based on a split GFP–Renilla luciferase reporter, we have established a micro-fusion inhibition test (mFIT) that allows the identification and quantification of these neutralizing antibodies. This assay has been optimized for high-throughput use and its applicability has been demonstrated by screening monoclonal antibody (mAb)-mediated inhibition of RSV and NiV fusion and, separately, the development of fusion-inhibitory antibodies following NiV vaccine immunization in pigs. In light of the recent emergence of coronavirus disease 2019 (COVID-19), a similar assay was developed for SARS-CoV-2 and used to screen mAbs and convalescent patient plasma for fusion-inhibitory antibodies. Using mFITs to assess antibody responses following natural infection or vaccination is favourable, as this assay can be performed entirely at low biocontainment, without the need for live virus. In addition, the repertoire of antibodies that inhibit cell–cell fusion may be different to those that inhibit particle entry, shedding light on the mechanisms underpinning antibody-mediated neutralization of viral spread.

There is dire need for an effective and affordable vaccine against SARS-CoV-2 to tackle the ongoing pandemic. In this study, we describe a modular virus-like particle vaccine candidate displaying the SARS-CoV-2 spike glycoprotein receptor-binding domain (RBD) using SpyTag/SpyCatcher technology (RBD-SpyVLP). Low doses of RBD-SpyVLP in a prime-boost regimen induced a strong neutralising antibody response in mice and pigs that was superior to convalescent human sera. We evaluated antibody quality using ACE2 blocking and neutralisation of cell infection by pseudovirus or wild-type SARS-CoV-2. Using competition assays with a monoclonal antibody panel, we showed that RBD-SpyVLP induced a polyclonal antibody response that recognised all key epitopes on the RBD, reducing the likelihood of selecting neutralisation-escape mutants. The induction of potent and polyclonal antibody responses by RBD-SpyVLP provides strong potential to address clinical and logistic challenges of the COVID-19 pandemic. Moreover, RBD-SpyVLP is highly resilient, thermostable and can be lyophilised without losing immunogenicity, to facilitate global distribution and reduce cold-chain dependence.

Clinical development of the COVID-19 vaccine candidate ChAdOx1 nCoV-19, a replication-deficient simian adenoviral vector expressing the full-length SARS-CoV-2 spike (S) protein was initiated in April 2020 following non-human primate studies using a single immunisation. Here, we compared the immunogenicity of one or two doses of ChAdOx1 nCoV-19 in both mice and pigs. Whilst a single dose induced antigen-specific antibody and T cells responses, a booster immunisation enhanced antibody responses, particularly in pigs, with a significant increase in SARS-CoV-2 neutralising titres.

There are seven ‘established’ morbilliviruses: rinderpest (RPV), measles (MeV), small ruminant (PPRV), cetacean (CeMV), phocine (PDV), canine (CDV) and feline (FmoV) morbillivirus, the majority of which are thought to have narrow host ranges. The exception is CDV, which has been reported in almost all families of the order Carnivora, as well as non-human primates. Morbilliviruses enter cells through one of two proteinaceous receptors: signalling lymphocyte activation molecule F1 (SLAMF1), present on the surface of many immune cells; or Nectin-4, expressed on the basolateral side of various polarised epithelial cells. Given the close similarity of these viruses and their universal use of related receptors, there are realistic fears morbilliviruses may spill-over into naive hosts, i.e. following eradication of RPV in cattle. This spill-over is not entirely unprecedented; PDV and MeV are thought to share common ancestors with CDV and RPV, respectively. To examine morbillivirus receptor usage and predict future spill-over and zoonotic transmission events, we have developed a high-throughput, quantifiable, cell-cell fusion assay. Effector cells, expressing the viral F and H glycoproteins as well as a split-luciferase reporter, are co-cultured with target cells expressing SLAMF1 or Nectin-4 receptors and the remaining half of the reporter. Upon functional receptor engagement the effector and target cells fuse, allowing cytoplasmic mixing and reconstitution of the reporter, a process which can be quantified. Using these assays we are investigating whether the receptor usage profile of morbilliviruses represents an important factor in spill-over transmission into new hosts.