Foot-and-mouth disease (FMD) affects the livestock industry and socio-economic sustainability of many African countries. The success of FMD control programs in Africa depends largely on understanding the dynamics of FMD virus (FMDV) spread. In light of the recent outbreaks of FMD that affected the North-Western African countries in 2018 and 2019, we investigated the evolutionary phylodynamics of the causative serotype O viral strains all belonging to the East-Africa 3 topotype (O/EA-3). We analyzed a total of 489 sequences encoding the FMDV VP1 genome region generated from samples collected from 25 African and Western Asian countries between 1974 and 2019. Using Bayesian evolutionary models on genomic and epidemiological data, we inferred the routes of introduction and migration of the FMDV O/EA-3 topotype at the inter-regional scale. We inferred a mean substitution rate of 6.64 10(-3) nt/site/year and we predicted that the most recent common ancestor (MRCA) for our panel of samples circulated between February 1967 and November 1973 in Yemen, likely reflecting the epidemiological situation in under sampled cattle-exporting East African countries. Our study also reinforces the role previously described of Sudan and South Sudan as a frequent source of FMDVs spread. In particular, we identified two transboundary routes of O/EA-3 diffusion: the first from Sudan to North-East Africa, and from the latter into Israel and Palestine AT; a second from Sudan to Nigeria, Cameroon, and from there to further into West and North-West Africa. This study highlights the necessity to reinforce surveillance at an inter-regional scale in Africa and Western Asia, in particular along the identified migration routes for the implementation of efficient control measures in the fight against FMD.

Duck enteritis virus (DEV) and Pasteurella multocida, the causative agent of duck plague and fowl cholera, are acute contagious diseases and leading causes of morbidity and mortality in duck. The NHEJ-CRISPR/Cas9-mediated gene editing strategy, accompanied with the Cre–Lox system, have been employed in the present study to show that two new sites at UL55-LORF11 and UL44-44.5 loci in the genome of the attenuated Jansen strain of DEV can be used for the stable expression of the outer membrane protein H (ompH) gene of P. multocida that could be used as a bivalent vaccine candidate with the potential of protecting ducks simultaneously against major viral and bacterial pathogens. The two recombinant viruses, DEV-OmpH-V5-UL55-LORF11 and DEV-OmpH-V5-UL44-44.5, with the insertion of ompH-V5 gene at the UL55-LORF11 and UL44-44.5 loci respectively, showed similar growth kinetics and plaque size, compared to the wildtype virus, confirming that the insertion of the foreign gene into these did not have any detrimental effects on DEV. This is the first time the CRISPR/Cas9 system has been applied to insert a highly immunogenic gene from bacteria into the DEV genome rapidly and efficiently. This approach offers an efficient way to introduce other antigens into the DEV genome for multivalent vector. 

In this study, we developed a new recombinant virus rHVT-F using a Turkey herpesvirus (HVT) vector, expressing the fusion (F) protein of the genotype XII Newcastle disease virus (NDV) circulating in Peru. We evaluated the viral shedding and efficacy against the NDV genotype XII challenge in specific pathogen-free (SPF) chickens. The F protein expression cassette was inserted in the unique long (UL) UL45–UL46 intergenic locus of the HVT genome by utilizing a clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 gene-editing technology via a non-homologous end joining (NHEJ) repair pathway. The rHVT-F virus, which expressed the F protein stably in vitro and in vivo, showed similar growth kinetics to the wild-type HVT (wtHVT) virus. The F protein expression of the rHVT-F virus was detected by an indirect immunofluorescence assay (IFA), Western blotting, and a flow cytometry assay. The presence of an NDV-specific IgY antibody was detected in serum samples by an enzyme-linked immunosorbent assay (ELISA) in SPF chickens vaccinated with the rHVT-F virus. In the challenge experiment, the rHVT-F vaccine fully protects a high, and significantly reduced, virus shedding in oral at 5 days post-challenge (dpc). In conclusion, this new rHVT-F vaccine candidate is capable of fully protecting SPF chickens against the genotype XII challenge.

Lumpy skin disease virus (LSDV) is an emerging poxviral pathogen of cattle that is currently spreading throughout Asia. The disease situation is of high importance for farmers and policy makers in Asia. In October 2020, feral cattle in Hong Kong developed multi-focal cutaneous nodules consistent with lumpy skin disease (LSD). Gross and histological pathology further supported the diagnosis and samples were sent to the OIE Reference Laboratory at The Pirbright Institute for confirmatory testing. LSDV was detected using quantitative polymerase chain reaction (qPCR) and additional molecular analyses. This is the first report of LSD in Hong Kong. Whole genome sequencing (WGS) of the strain LSDV/Hong Kong/2020 and phylogenetic analysis were carried out in order to identify connections to previous outbreaks of LSD, and better understand the drivers of LSDV emergence. Analysis of the 90 core poxvirus genes revealed LSDV/Hong Kong/2020 was a novel strain most closely related to the live-attenuated Neethling vaccine strains of LSDV and more distantly related to wildtype LSDV isolates from Africa, the Middle East and Europe. Analysis of the more variable regions located towards the termini of the poxvirus genome revealed genes in LSDV/Hong Kong/2020 with different patterns of grouping when compared to previously published wildtype and vaccine strains of LSDV. This work reveals that the LSD outbreak in Hong Kong in 2020 was caused by a different strain of LSDV than the LSD epidemic in the Middle East and Europe in 2015–2018. The use of WGS is highly recommended when investigating LSDV disease outbreaks.

In recent months, several outbreaks with clinical signs of MDV-1 were reported in Iranian parent and laying hen farms, in addition to backyard chickens. Several meq gene sequences from these outbreaks were amplified and molecularly characterised.The meq protein sequences revealed three different sizes, namely the standard 339 aa, a shorter form of 338 aa lacking a proline residue at position 191, and a very short (vs) size of 265 aa. Based on sequence and size, the 265 aa meq has never been reported from international research groups before. The protein has only one PPPP repeat motif suggesting it belongs to a highly virulent strain.The standard meq sequences showed 100% BLAST identity to the vv+ isolate Polen5. However, the 338 aa form clustered to the clade usually reported from North America.This is the first report on genetic analysis of MDV-1 from Iran, but further study is required to obtain a better picture of the diversity and prevalence of different MDV-1 strains circulating in the country's farms, backyard poultry and/or other bird species.

African swine fever virus (ASFV) has a major global economic impact. With a case fatality in domestic pigs approaching 100%, it currently presents the largest threat to animal farming. Although genomic differences between attenuated and highly virulent ASFV strains have been identified, the molecular determinants for virulence at the level of gene expression have remained opaque. Here we characterise the transcriptome of ASFV genotype II Georgia 2007/1 (GRG) during infection of the physiologically relevant host cells, porcine macrophages. In this study we applied Cap Analysis Gene Expression sequencing (CAGE-seq) to map the 5' ends of viral mRNAs at 5 and 16 hours post-infection. A bioinformatics analysis of the sequence context surrounding the transcription start sites (TSSs) enabled us to characterise the global early and late promoter landscape of GRG. We compared transcriptome maps of the GRG isolate and the lab-attenuated BA71V strain that highlighted GRG virulent-specific transcripts belonging to multigene families, including two predicted MGF 100 genes I7L and I8L. In parallel, we monitored transcriptome changes in the infected host macrophage cells. Of the 9,384 macrophage genes studied, transcripts for 652 host genes were differentially regulated between 5 and 16 hours-post-infection compared with only 25 between uninfected cells and 5 hours post-infection. NF-kB activated genes and lysosome components like S100 were upregulated, and chemokines such as CCL24, CXCL2, CXCL5 and CXCL8 downregulated. African swine fever virus (ASFV) causes haemorrhagic fever in domestic pigs with case fatality rates approaching 100%, and no approved vaccines or antivirals. The highly-virulent ASFV Georgia 2007/1 strain (GRG) was the first isolated when ASFV spread from Africa to the Caucasus region in 2007. Then spreading through Eastern Europe, and more recently across Asia. We used an RNA-based next generation sequencing technique called CAGE-seq to map the starts of viral genes across the GRG DNA genome. This has allowed us to investigate which viral genes are expressed during early or late stages of infection and how this is controlled, comparing their expression to the non-virulent ASFV-BA71V strain to identify key genes that play a role in virulence. In parallel we investigated how host cells respond to infection, which revealed how the ASFV suppresses components of the host immune response to ultimately win the arms race against its porcine host.

Tabanids (Diptera: Tabanidae) are large haematophagous flies that cause both direct (by biting nuisance) and indirect (primarily by mechanical transmission of diseases) damage to host species. Research studies on this family have received little attention in some parts of Europe. Our aims were to characterise the species richness, abundance, and peak of activity of tabanid fly species in a region of Northern Spain. Home-made canopy traps, sweep nets, and Malaise traps were employed for the collection of tabanids across four cattle farms, two equestrian centres, and two golf courses during a 3-month period in the summer of 2020. A total of 300 specimens of 27 tabanid species belonging to eight genera were identified. The most prevalent species were Haematopota pluvialis (23.3%), Tabanus eggeri (20.0%), and Tabanus bromius (8.0%). The former species was recorded biting humans and therefore should be considered of relevance to public health. Tabanids were more diverse and abundant in scrubland and grazing pastures [relative abundance (RA) =  > 10%; species richness (S) = 8–12; Shannon-Index (H´) = 1.5 − 2.1] compared to crop landscapes (RA =  < 1%; S = 0–1; H´ = 0) according to canopy traps. The tabanid population dynamics was determined to be short, with the greatest abundance and diversity concentrated in mid-late July. This study updates the checklist of this Diptera group in the Northern Spain from nine known extant species to 31 species, providing the first data on the summer peaks of activity of tabanids for this region.

In the light of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic, we have developed a porcine respiratory coronavirus (PRCV) model for in depth mechanistic evaluation of the pathogenesis, virology and immune responses of this important family of viruses. Pigs are a large animal with similar physiology and immunology to humans and are a natural host for PRCV. Four PRCV strains were investigated and shown to induce different degrees of lung pathology. Importantly, although all four strains replicated equally well in porcine cell lines in vitro and in the upper respiratory tract in vivo, PRCV strains causing more severe lung pathology were also able to replicate in ex vivo tracheal organ cultures as well as in vivo in the trachea and lung. The time course of infection of PRCV 135, which caused the most severe pulmonary pathology, was investigated. Virus was shed from the upper respiratory tract until day 10 post infection, with infection of the respiratory mucosa, as well as olfactory and sustentacular cells, providing an excellent model to study upper respiratory tract disease in addition to the commonly known lower respiratory tract disease from PRCV. Infected animals made antibody and T cell responses that cross reacted with the four PRCV strains and Transmissible Gastroenteritis Virus. The antibody response was reproduced in vitro in organ cultures. Comparison of mechanisms of infection and immune control in pigs infected with PRCVs of differing pathogenicity with human data from SARS-CoV-2 infection and from our in vitro organ cultures, will enable key events in coronavirus infection and disease pathogenesis to be identified.