The long-term control strategy of SARS-CoV-2 and other major respiratory viruses needs to include antivirals to treat acute infections, in addition to the judicious use of effective vaccines. Whilst COVID-19 vaccines are being rolled out for mass vaccination, the modest number of antivirals in use or development for any disease bears testament to the challenges of antiviral development. We recently showed that non-cytotoxic levels of thapsigargin (TG), an inhibitor of the sarcoplasmic/endoplasmic reticulum (ER) Ca²⁺ ATPase pump, induces a potent host innate immune antiviral response that blocks influenza A virus replication. Here we show that TG is also highly effective in blocking the replication of respiratory syncytial virus (RSV), common cold coronavirus OC43, SARS-CoV-2 and influenza A virus in immortalized or primary human cells. TG’s antiviral performance was significantly better than remdesivir and ribavirin in their respective inhibition of OC43 and RSV. Notably, TG was just as inhibitory to coronaviruses (OC43 and SARS-CoV-2) and influenza viruses (USSR H1N1 and pdm 2009 H1N1) in separate infections as in co-infections. Post-infection oral gavage of acid-stable TG protected mice against a lethal influenza virus challenge. Together with its ability to inhibit the different viruses before or during active infection, and with an antiviral duration of at least 48 h post-TG exposure, we propose that TG (or its derivatives) is a promising broad-spectrum inhibitor against SARS-CoV-2, OC43, RSV and influenza virus.

African swine fever (ASF) is a severe haemorrhagic disease of domestic and wild pigs caused by the African swine fever virus (ASFV). In recent years, ASF has steadily spread towards new geographical areas, reaching Europe and Asia. On January 15th, 2019, Mongolia reported its first ASF outbreak to the World Organization for Animal Health (OIE), becoming, after China, the second country in the region affected by the disease. Following an event of unusual mortality in domestic pigs in Bulgan Province, a field team visited four farms and a meat market in the region to conduct an outbreak investigation and collect samples for laboratory analysis. Different organs were examined for ASF associated lesions, and total nucleic acid was extracted for real-time PCR, virus isolation and molecular characterization. The real-time PCR results confirmed ASFV DNA in 10 out of 10 samples and ASFV was isolated. Phylogenetic analysis established that ASFVs from Mongolia belong to genotype II and serogroup 8. The viruses were identical to each other, and to domestic pig isolates identified in China and Russia, based on the comparison of five genomic targets. Our results suggest a cross-border spread of ASFV, without indicating the source of infection.

Arboviruses such as bluetongue virus (BTV) replicate in arthropod vectors involved in their transmission between susceptible vertebrate-hosts. The “classical” BTV strains infect and replicate effectively in cells of their insect-vectors (Culicoides biting-midges), as well as in those of their mammalian-hosts (ruminants). However, in the last decade, some “atypical” BTV strains, belonging to additional serotypes (e.g., BTV-26), have been found to replicate efficiently only in mammalian cells, while their replication is severely restricted in Culicoides cells. Importantly, there is evidence that these atypical BTV are transmitted by direct-contact between their mammalian hosts. Here, the viral determinants and mechanisms restricting viral replication in Culicoides were investigated using a classical BTV-1, an “atypical” BTV-26 and a BTV-1/BTV-26 reassortant virus, derived by reverse genetics. Viruses containing the capsid of BTV-26 showed a reduced ability to attach to Culicoides cells, blocking early steps of the replication cycle, while attachment and replication in mammalian cells was not restricted. The replication of BTV-26 was also severely reduced in other arthropod cells, derived from mosquitoes or ticks. The data presented identifies mechanisms and potential barriers to infection and transmission by the newly emerged “atypical” BTV strains in Culicoides.

The avifauna endemic to islands is particularly susceptible to population declines and extinctions resulting from the introduction of non-native pathogens. Three pathogens of concern are the avian malaria parasites, the avian poxviruses, and West Nile virus— each of which can be transmitted by Culex quinquefasciatus, a highly adaptive and invasive mosquito. Culex quinquefasciatus has dramatically expanded its range in recent centuries and is now established throughout much of the tropics and sub-tropics, including on many islands that are remote from mainland landmasses and where this geographic separation historically protected island species from mosquito-borne diseases. The potential for ecological disruption by Cx. quinquefasciatus has been particularly striking in the Hawaiian Islands, where the introduction and transmission of avian malaria and avian poxvirus led to the extinction of several endemic bird species, with many more at risk. With Cx. quinquefasciatus now present in many insular communities and global trade and tourism increasing links between these areas, both to each other and to mainlands, there is growing concern that patterns of avian decline in Hawai‘i may be played out in other insular ecosystems. The implementation of traditional methods for Cx. quinquefasciatus control, including larval source management, is often impractical at large scale and when breeding sites are numerous and difficult to locate—typical issues associated with invasive species removal. One alternative approach would be the utilisation of genetic control methods, several of which have been successfully developed in other mosquitos such as Aedes aegypti and the malaria vector Anopheles gambiae. However, the development of similar tools for Cx. quinquefasciatus has been comparatively limited. Here we review the threat that Cx. quinquefasciatus poses as a vector of avian pathogens to island avifauna and discuss specific examples of at-risk bird populations on the islands of Hawai‘i, New Zealand and Galápagos. We also review the major options for the deployment of genetic control tools against Cx. quinquefasciatus, and discuss the current state of the field with a focus on radiation-based sterilisation, transgenic methods, and transinfections using the bacterial endosymbiont Wolbachia.

In order to better understand differences in the outcome of infectious bursal disease virus (IBDV) infection, we inoculated a very virulent (vv) strain into White Leghorn chickens of inbred line W that was previously reported to experience over 24% flock mortality, and three inbred lines (15I, C.B4 and 0) that were previously reported to display no mortality. Within each experimental group, some individuals experienced more severe disease than others but line 15I birds experienced milder disease based on average clinical scores, percentage of birds with gross pathology, average bursal lesion scores and average peak bursal virus titre. RNA-Seq analysis revealed that more severe disease in line W was associated with significant up-regulation of pathways involved in inflammation, cytoskeletal regulation by Rho GTPases, nicotinic acetylcholine receptor signaling, and Wnt signaling in the bursa compared to line 15I. Primary bursal cell populations isolated from uninfected line W birds contained a significantly greater percentage of KUL01+ macrophages than cells isolated from line 15I birds (p < 0.01) and, when stimulated ex vivo with LPS, showed more rapid up-regulation of pro-inflammatory gene expression than those from line 15I birds. We hypothesize that a more rapid induction of pro-inflammatory cytokine responses in bursal cells following IBDV infection leads to more severe disease in line W birds than in line 15I.

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.

In recent years, the CRISPR/Cas9-based gene-editing techniques have been well developed and applied widely in several aspects of research in the biological sciences, in many species, including humans, animals, plants, and even in viruses. Modification of the viral genome is crucial for revealing gene function, virus pathogenesis, gene therapy, genetic engineering, and vaccine development. Herein, we have provided a brief review of the different technologies for the modification of the viral genomes. Particularly, we have focused on the recently developed CRISPR/Cas9-based gene-editing system, detailing its origin, functional principles, and touching on its latest achievements in virology research and applications in vaccine development, especially in large DNA viruses of humans and animals. Future prospects of CRISPR/Cas9-based gene-editing technology in virology research, including the potential shortcomings, are also discussed.

The Animal Welfare Assessment Grid (AWAG) is a method for assessing quality of life, originally designed for experimental primates. This study adapts the AWAG for use in cattle and pigs, by adapting the factors included for these species and including data which had been collected previously as the standard approach to monitoring these species in research. The intention is that the results presented here will allow the future data collected for experimental cattle and pigs to be optimised for inclusion in an AWAG. Data were collected from two vaccine assessment studies at the Pirbright Institute. Factors were scored for every recorded event using retrospective data and CCTV clips. There was a lack of behavioural data recorded in both studies, which limited the accuracy of assessing each animal's welfare. This paper emphasises the importance of including behavioural information when assessing welfare and not simply relying on assessment of physical condition. Scores peaked following an exponential rise as animals reached set humane end points. This demonstrated the potential of using the AWAG to aid the decision-making of when euthanasia should be performed. Our study shows the AWAG to be a useful tool for assessing welfare, which can be used in harm:benefit assessment.