Seneca Valley Virus (SVV) infection has recently spread to pig farms in Canada, America, and China. Humans, mice, and houseflies have been identified as hosts and reservoirs. Although such cross-species transmission events are often limited, sustained outbreaks in a new mammalian host can occur. To determine if mink are a new mammalian host of SSV, we studied the molecular characteristics of isolated SVV genomes and analyzed challenge, pathology, and immune response data. The study was the first systemic analysis of a newly isolated strain of SVV from pigs. The strain caused an intestinal infection with associated pathologic changes in mink. SVV stimulated the production of a specific neutralizing antibody. The findings highlight the importance of identifying SVV infection in mink and the host to detect mutated SSV that could threaten livestock and pose public health and economic risks.

Livestock industry supports the livelihood of around 1.3 billion people in the world, with swine industry contributing with 30 % of total livestock production worldwide. To maintain and guarantee this production, a pivotal point according to the OIE is addressing potential biohazards. To control them, permanent sero-surveillance is crucial to achieve more focused veterinary public health intervention and prevention strategies, to break the chains of transmission, and to enable fast responses against outbreaks. Within this context, multiplex assays are powerful tools with the potential to simplify surveillance programs, since they reduce time, labour, and variability within analysis. In the present work, we developed a multiplex bead-based assay for the detection of specific antibodies to six relevant pathogens affecting swine: ASFV, CSFV, PRRSV, SIV, TB, and HEV. The most immunogenic target antigen of each pathogen was selected as the target protein to coat different microsphere regions in order to develop this multiplex assay. A total of 1544 serum samples from experimental infections as well as field samples were included in the analysis. The 6plex assay exhibited credible diagnostic parameters with sensitivities ranging from 87.0 % to 97.5 % and specificities ranging from 87.9 % to 100.0 %, demonstrating it to be a potential high throughput tool for surveillance of infectious diseases in swine.

Animals like mink, cats and dogs are susceptible to SARS-CoV-2 infection. In the Netherlands, 69 out of 127 mink farms were infected with SARS-CoV-2 between April and November 2020 and all mink on infected farms were culled after SARS-CoV-2 infection to prevent further spread of the virus. On some farms, (feral) cats and dogs were present. This study provides insight into the prevalence of SARS-CoV-2 positive cats and dogs in ten infected mink farms and their possible role in transmission of the virus. Throat and rectal swabs of 101 cats (12 domestic and 89 feral cats) and 13 dogs of ten farms were tested for SARS-CoV-2 using PCR. Serological assays were performed on serum samples from 62 adult cats and all 13 dogs. Whole Genome Sequencing was performed on one cat sample. Cat-to-mink transmission parameters were estimated using data from all ten farms. This study shows evidence of SARS-CoV-2 infection in twelve feral cats and two dogs. Eleven cats (19%) and two dogs (15%) tested serologically positive. Three feral cats (3%) and one dog (8%) tested PCR-positive. The sequence generated from the cat throat swab clustered with mink sequences from the same farm. The calculated rate of mink-to-cat transmission showed that cats on average had a chance of 12% (95%CI 10% to 18%) of becoming infected by mink, assuming no cat-to-cat transmission. As only feral cats were infected it is most likely that infections in cats were initiated by mink, not by humans. Whether both dogs were infected by mink or humans remains inconclusive. This study presents one of the first reports of interspecies transmission of SARS-CoV-2 that does not involve humans, namely mink-to-cat transmission, which should also be considered as a potential risk for spread of SARS-CoV-2.

A document by Brecht Daams, written on Authorea.

African Swine Fever (ASF) is one of the most devastating infectious diseases affecting domestic pigs and wild boar. The grave socio-economic impact of African Swine Fever infection at a global level makes large-scale rapid and robust diagnosis a critical step towards effective control. However, the nucleic acid purification required in most molecular detection methods is time- and labor-intensive, prone to nucleic acid loss or contamination, and impractical for massive active screening or for use in resource-limited areas. Here we describe multiple-probe-assisted DNA capture and amplification technology (MADCAT) - a novel sensitive, simple, and reliable method for detecting ASFV directly from whole blood or other complex matrices. Through the unique DNA capture method which specifically capture only the target DNA onto the well for subsequent amplification, MADCAT abandons the complicated extraction protocol and achieves ultrafast and high-throughput detection. The sample-to-result time for 96 samples is about 100 min, as compared with the 3 - 4 h time of the standard real time qPCR method. The limit of detection (LOD) is 0.5 copies/μL and is 10 times more sensitive than an OIE-recommended qPCR assay when testing serially diluted whole blood samples. The assay is 100% specific against other common swine pathogens. In clinical diagnosis of 48 field samples, all 22 positive samples were correctly identified with lower Ct values than OIE-recommended qPCR, confirming its high diagnostic sensitivity (100%). Owing to its high-throughput, specific high-sensitivity, and cost-efficient features, MADCAT shows great potential for future use in clinical ASFV active screening.

The fast spread of SARS-CoV-2 has led to a global pandemic, calling for fast and accurate assays to allow infection diagnosis and prevention of transmission. We aimed to develop a molecular beacon (MB)-based detection assay for SARS-CoV-2, designed to, detect the ORF1ab and S genes, proposing a two-stage COVID-19 testing strategy, using MBs to detect the presence of target amplicons by fluorescence analysis. Two MBs were designed, optimized in terms of concentration, fluorescence plateaus of hybridization, reaction temperature and best real-time results. A total of 450 nasopharyngel and throat swab samples (418 positive and 32 negative) were tested with the MB assay and the fluorescence levels compared with the cycle threshold (Ct) values obtained from a commercial RT-PCR test in terms of test duration, sensitivity and specificity. Our results show that the samples with higher fluorescence levels correspond to those with low Ct values, suggesting a correlation between viral load and increased MB fluorescence. The proposed assay represents a fast (total duration of 2 h 20 min including amplification and fluorescence reading stages) and simple way of detecting SARS-CoV-2 in clinical samples from the upper respiratory tract. Our two-stage testing strategy is suitable for further development into a point-of-care assay and potentially scalable to population level.

The total impact of the worldwide COVID-19 pandemic is still emerging, changing all relationships as a result, including those with pet animals. In the infection process, the use of Angiotensin-converting enzyme 2 (ACE2) as a cellular receptor to the spike protein of the new coronavirus is a fundamental step. In this sense, understanding which residue plays what role in the interaction between SARS-CoV-2 spike glycoprotein and ACE2 from cats, dogs, and ferrets is an important guide for helping to choose which animal model can be used to study the pathology of COVID-19 and if there are differences between these interactions and those occurring in the human system. Hence, trying to help to answer these questions, we performed classical molecular dynamics simulations to evaluate, from an atomistic point of view, the interactions in these systems. Our results show that there are significant differences in the interacting residues between the systems from different animal species, and the role of ACE2 key residues are different in each system and can assist in the search for different inhibitors for each animal.

Porcine epidemic diarrhea virus (PEDV) strains have been clarified into two genotypes, G1 and G2, based on the sequence of the spike (S) gene. Amino acid mutations that distinguish the two PEDV genotypes were mostly located in the N-terminal domain (NTD) (aa 1-380) of S protein. The fact of increased outbreaks of G2 subtype PEDV and the failure of G1 subtype PEDV strain (CV777)-based vaccine in China since 2010 suggested that multiple amino acid mutations located in the NTD altered the antigenicity of S protein. To determine the role of the NTD of S protein in the antigenicity difference, the NTD of the CV777 vaccine strain (G1) and CH/ZMDZY/11 strain (G2) was expressed in E. coli, respectively. polyclonal antibodies (PAbs) against genotype-specific S proteins were prepared by immunizing BALB/c mice using purified S proteins. Antigenicity was systematically compared by detection of PAbs against two genotype PEDV strains and purified S proteins using Western blot, indirect enzyme-linked immunosorbent assay (ELISA), indirect immunofluorescence assay (IFA), and serum cross-neutralization assay (SN). Consistent with the multiple amino acid mutations in the NTD of S protein, different antigenic cross-reactivity between the two genotypes was demonstrated. There was six-fold and more than twenty-fold difference in ELISA and SN titer between anti-CV777 S protein antibodies against G1 and G2 subtype strains, respectively. There was twofold and eight-fold difference in ELISA and SN titer between anti-ZMDZY S protein antibodies against G1 and G2 genotype strains, respectively. The results proved that the NTD of S protein contributes to the antigenicity difference between PEDV genotypes G1 and G2, and highlighted a G2 strain should be used to develop a vaccine for providing better protection against prevalent genotype of PEDV.

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Worldwide, wild birds are frequently suspected to be involved in the occurrence of outbreaks in captive-bred birds although proofs are lacking and most of the dedicated studies are insufficiently conclusive to confirm or characterize the roles of wild birds in such outbreaks. The aim of this study was to assess and compare, for the most prevalent peridomestic wild birds, the different exposure routes for Avian Influenza and Newcastle disease viruses in conservation breeding sites of Houbara bustards in the United Arab Emirates. To do so, we considered all of the potential pathways by which captive bustards could be exposed to Avian Influenza and Newcastle disease viruses by wild birds, and ran a comparative study of the likelihood of exposure via each of the pathways considered. We merged data from an ecological study dedicated to local wild bird communities with an analysis of the contacts between wild birds and captive bustards and with a prevalence survey of AIV and NDV in wild bird populations. We also extracted data from an extensive review of the scientific literature and by the elicitation of expert opinion. Overall, this analysis highlighted that captive bustards had a high risk of being exposed to pathogens by wild birds. This risk was higher for Newcastle disease virus than Avian influenza virus, and House sparrows represented the riskiest species for the transmission of both viruses through indirect exposure from consumption of water contaminated from the faeces of an infectious bird that got inside the aviary. Thus, this analysis reveals that wild peridomestic birds may play a role in the transmission of avian pathogens to captive bred birds. These results also reaffirm the need to implement sanitary measures to limit contacts between wild and captive birds and highlight priority targets for a thoughtful and efficient sanitary management strategy.

Background. First vaccines for prevention of Coronavirus disease 2019 (COVID-19) are becoming available but there is a huge and unmet need for specific forms of treatment. In this study we aimed to evaluate the potent anti-SARS-CoV-2 effect of siRNA both in vitro and in vivo. Methods. To identify most effective molecule out of a panel of 15 in silico designed siRNAs, an in vitro screening system based on vectors expressing SARS-CoV-2 genes fused with the firefly luciferase reporter gene and SARS-CoV-2-infected cells was used. The most potent siRNA, siR-7, was modified by Locked nucleic acids (LNAs) to obtain siR-7-EM with increased stability and was formulated with the peptide dendrimer KK-46 for enhancing cellular uptake to allow topical application by inhalation of the final formulation - siR-7-EM/KK-46. Using the Syrian Hamster model for SARS-CoV-2 infection the antiviral capacity of siR-7-EM/KK-46 complex was evaluated. Results. We identified the siRNA, siR-7, targeting SARS-CoV-2 RNA-dependent RNA polymerase (RdRp) as the most efficient siRNA inhibiting viral replication in vitro. Moreover, we have shown that LNA-modification and complexation with the designed peptide dendrimer enhanced the antiviral capacity of siR-7 in vitro. We demonstrated significant reduction of virus titer and total lung inflammation in the animals exposed by inhalation of siR-7-EM/KK-46 in vivo. Conclusions. Thus, we developed a therapeutic strategy for COVID-19 based on inhalation of a modified siRNA-peptide dendrimer formulation.

Genetic characterization may provide useful insights into severe acute respiratory coronavirus 2 (SARS CoV 2) circulating in Bangladesh. Here we analyzed the SARS Cov 2 positive 41 nasopharyngeal samples collected in lysis buffer obtained at different regional national laboratories of Bangladesh during July – December 2020. Full length spike gene was amplified and SARS CoV 2 was confirmed upon Sanger sequencing. Multiple ClustalW alignment and phylogenetic study of 15 strains showed that genetically diverse SARS CoV 2 is circulating in the country where 80 documented and 55 novel substitutions have been observed in different regions of spike glycoprotein. Major mutations and/or deletions was identified at conserved amino acid positions that are functionally linked to host transition, antigenic drift, host surface receptor binding or antibody recognition sites, and viral oligomerization interfaces that may have significant effects on pathogenic capacity and epidemiological signatures.

Ticks are involved in the transmission of various pathogens and some tick-borne diseases cause significant problems for the health of humans and livestock. Despite their obvious importance, the composition of viral communities in ticks, and their interactions with pathogens, is poorly understood, particularly in Eastern Europe that constitutes (via bird migrations for example) a major hub for animal-arthropod vectors exchanges. The aim of this study was first to describe the virome of Dermacentor sp., Rhipicephalus sp. and Haemaphysalis sp. ticks collected from poorly investigated regions of Romania (Iasi and Tulcea counties) located at the intersection of various biotopes, countries and routes of migrations. We then focused the study on viruses that could have potential relevance for human and animal health. More than 500 ticks were collected in 2019 from the environment and from small ruminants and analyzed by high-throughput transcriptome sequencing. Among the viral communities infecting Romanian ticks, viruses belonging to the Flaviviridae, Phenuiviridae and Nairoviridae families were identified and full genomes were derivedPhylogenetic analyses placed them in clades where mammalian isolates are found, suggesting that these viruses could constitute novel arboviruses. We also assessed the bacterial microbiome of the collected ticks. The characterization of these microbial communities increases the knowledge of the diversity of viruses in Eastern Europe and provide a basis for further studies on the relationship between ticks and tick-borne viruses.

Porcine Sapovirus (SaV) was first identified by electron microscopy in the United States in 1980 and has since been reported from both asymptomatic and diarrheic pigs usually in mixed infection with other enteric pathogens. SaV as the sole etiological agent of diarrhea in naturally infected pigs has not previously been reported in the United States. Here, we used four independent lines of evidence including metagenomics analysis, real-time RT-PCR (rRT-PCR), histopathology, and in situ hybridization to confirm porcine SaV genogroup III (GIII) as the sole cause of enteritis and diarrhea in pigs. A highly sensitive and specific rRT-PCR was established to detect porcine SaV GIII. Examination of 184 fecal samples from the outbreak farm showed that pigs with clinical diarrhea had significantly lower Ct values (15.9 ± 0.59) compared to clinically unaffected pigs (35.8 ± 0.71). Further survey of 336 fecal samples from different states in the United States demonstrated that samples from pigs with clinical diarrhea had a comparable positive rate (45.3%) with those from non-clinical pigs (43.1%). However, the SaV-positive pigs with clinical diarrhea had significantly higher viral loads (Ct = 26.0 ± 0.5) than those positive but clinically healthy pigs (Ct = 33.2 ± 0.9). Phylogenetic analysis of 20 field SaVs revealed that all belonged to SaV GIII and recombination analysis indicated that intra-genogroup recombination occurred within the field isolates of SaV GIII. These results suggest that porcine SaV GIII plays an important etiologic role in swine enteritis and diarrhea and rRT-PCR is a reliable method to detect porcine SaV. Our findings provide significant insights to better understand the epidemiology and pathogenicity of porcine SaV.

Emerging data shows pregnant women with COVID-19 are at significantly higher risk of severe outcomes compared to non-pregnant women of similar age. This review discusses the invaluable insight revealed from vaccine clinical trials in women who were vaccinated and inadvertently became pregnant during the trial period. It further explores a number of clinical avenues in their management and proposes a drug development strategy in-line with clinical trials for vaccines and drug treatments for the drug development community. Little is known of the long-term effects of COVID-19 on the mother and the baby. We provide a rationale for our hypothesis that COVID-19 predisposes pregnant women to cardiovascular diseases later in life, in a similar way, to preeclampsia and may increase the risk of preeclampsia in their subsequent pregnancy. This is an ever-evolving landscape and early knowledge for healthcare providers and drug innovators is offered to ensure benefits outweigh the risks.

African swine fever (ASF) is one of the most severe infectious diseases of pigs. In this study, a LAMP assay coupled with the CRISPR Cas12a system was established in one tube for the detection of the ASFV p72 gene. The single-strand DNA-fluorophore-quencher (ssDNA-FQ) reporters and CRISPR-derived RNA (crRNAs) were screened and selected for the CRISPR detection system. In combination with LAMP amplification assay, the detection limit for the LAMP-CRISPR assay can reach 7 copies/μl of p72 gene per reaction. Furthermore, this method displays no cross-reactivity with other porcine DNA or RNA viruses. The performance of the LAMP-CRISPR assay was compared with real-time qPCR tests for clinical samples, a good consistency between the LAMP-CRISPR assay and real-time qPCR was observed. In the current study, a LAMP coupled with the CRISPR detection method was developed. The method shed a light on the convenient, portable, low cost, highly sensitive and specific detection of ASFV, demonstrating a great application potential for monitoring on-site ASFV in the field.

Megalocytivirus is a genus of piscine viruses that belongs to the Iridoviridae family, and this family comprises red seabream iridovirus (RSIV), turbot reddish body iridovirus (TRBIV), and infectious spleen and kidney necrosis virus. RSIV causes high mortality and economic losses in the rock bream (Oplegnathus fasciatus) Korean aquaculture industry. The World Organization for Animal Health’s Office International des Epizooties has provided a manual for RSIV detection. However, detection against TRBIV has not been confirmed. In this study, a multiplex PCR method was established to detect two genotypes of Megalocytivirus, RSIV and TRBIV. New primer pairs were optimized for the PCR reaction. A mixture of one universal and two specific primer pairs could amplify three distinct products targeting three different genes. The sensitivity of the primer pairs was evaluated and results showed a detection limit of 2.0  105 copies for each target gene. Moreover, the primer pairs did not amplify any other viruses. The evaluation of multiplex PCR using 21 RSIV Korean isolates has shown that it can distinguish two genotypes of Megalocytivirus from 21 RSIV Korean isolates. Finally, we describe a new multiplex PCR method to detect different genotypes of Megalocytivirus simultaneously, which makes the diagnosis of viral diseases occurring in the Korean aquaculture industry more convenient.

The hypothesis that feed ingredients could serve as vehicles for the transport and transmission of viral pathogens was first validated under laboratory conditions. To bridge the gap from the laboratory to the field, this current project tested whether three significant viruses of swine could survive in feed ingredients during long-distance commercial transport across the continental US. One-metric ton totes of soybean meal (organic and conventional) and complete feed were spiked with a 10 mL mixture of PRRSV 174, PEDV, and SVA and transported for 23 days in a commercial semi-trailer truck, crossing 29 states, and 10,183 km. Samples were tested for the presence of viral RNA by PCR, and for viable virus in soy-based samples by swine bioassay and in complete feed samples by natural feeding. Viable PRRSV, PEDV, and SVA were detected in both soy products and viable PEDV and SVA in complete feed. These results provide the first evidence that viral pathogens of pigs can survive in representative volumes of feed and feed ingredients during long-distance commercial transport across the continental US.

Rabies is a global viral zoonosis endemic to South Africa, resulting in fatal encephalitis in warm blooded animals, including humans. The loss of human lives and economic losses in rural areas through loss of livestock are substantial. A review was conducted of all confirmed rabies cases in South Africa from 1993 to 2019, with a total of 11 701 cases identified to species level to assess the wildlife plays in the epidemiology of rabies. A spatiotemporal cluster analysis using a discrete Poisson space-time probability model, accounting for underlying estimated dog and livestock densities, identified 13 significant clusters (p<0.05). These included four long-term clusters lasting more than 8 years in duration and seven short term clusters lasting less than 2 years, with the remaining two clusters being of intermediate length. Outside of these endemic clusters, wildlife outbreaks in the remainder of South Africa were often less than one and a half years in duration most likely due to the rapid decline of wildlife vectors, especially jackals associated with rabies infection. Domestic dogs accounted for 59.8% of cases, with domestic cats (3.2%), livestock (21.1%) and wildlife (15.8%) making up the remainder of the cases. Yellow mongoose (Cynictis penicillate) was the most frequently affected wildlife species, followed by bat-eared fox (Otocyon megalotis), black-backed jackal (Canis mesomelas), meerkat (Suricata suricatta) and aardwolf (Proteles cristatus). Rabies in wildlife species followed different spatial distributions: black-backed jackal cases were more common in the north-western parts of South Africa, yellow mongoose cases more frequent in central South Africa, and bat-eared fox and aardwolf cases were more frequent in southern and western South Africa. Clusters often spanned several provinces, showing the importance of coordinated rabies control campaigns across administrative boundaries, and high-risk areas were highlighted for rabies in South Africa.

African Swine Fever Virus (ASFV) causes a deadly disease of pigs which spread through southeast Asia in 2019. We investigated one of the first outbreaks of ASFV in Lao Peoples Democratic Republic amongst smallholder villages of Thapangtong District, Savannakhet Province. In this study, two ASFV affected villages were compared to two unaffected villages. Evidence of ASFV-like clinical signs appeared in pig herds as early as May 2019, with median epidemic days on 1 and 18 June in the two villages, respectively. Using participatory epidemiology mapping techniques, we found statistically significant spatial clustering in both outbreaks (P < 0.001). Villagers reported known risk factors for ASFV transmission  such as free-ranging management systems and wild boar access  in all four villages. The villagers reported increased pig trader activity from Vietnam before the outbreaks; however, the survey did not determine a single outbreak source. The outbreak caused substantial household financial losses with an average of 9 pigs lost to the disease, and Monte Carlo analysis estimated this to be USD 215 per household. ASFV poses a significant threat to food and financial security in smallholder communities such as Thapangtong, where 40.6% of the district’s population are affected by poverty. This study shows ASFV management in the region will require increased local government resources, knowledge of informal trader activity and wild boar monitoring alongside education and support to address intra-village risk factors such as free-ranging, correct waste disposal and swill feeding.

Between late December 2019 to early September 2020, over 10 million people globally were reportedly infected by the severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2), responsible for the coronavirus disease-2019 (COVID-19). In Africa, more than 300,000 infection occurred within the period, from which several viral genetic sequences were generated. Phylogenetic reconstruction of genomic data can provide epidemiological inferences about time of pathogen introduction, epidemic growth rate and temporal-spatial spread of the infection during disease outbreak. In this work, we studied the genetic epidemiology of COVID-19 in Africa. Genetic sequence data of SARS-CoV-2 and metadata from African countries were obtained from open-source sequence database hosted by the GISAID initiative. Whole genome sequences were subjected to multiple sequence alignment, from which Maximum Likelihood phylogenetic tree was constructed based on the general time reversible model. Of the 227 genetic sequences obtained for 9 African countries (DRC=133, Senegal=23, South Africa=20, Ghana=15, Tunisia=6, Algeria=3, Gambia=3, Egypt=2 and Nigeria=2), 220 were whole genome sequences while 7 were partial genome sequences of the surface glycoprotein S. Phylogenetic analysis confirmed multiple introductions of the virus to the continent from multiple external sources prior to local adaptation and spread. The very close alignment of three viruses - Ghana/1659_S14/2020|EPI_ISL_422405, DRC/KN0054/2020|EPI_ISL_417437, and South_Africa/R05475/2020|EPI_ISL_435059 – to the reference Wuhan strain on the time tree, suggests possible introduction and circulation of the virus into the continent much earlier than when the first case was announced on February 15 2020. In conclusion, this study provided evidence to support multiple introductions of SARS-CoV-2 into Africa, and further suggests that the virus may have already been circulating in the continent prior to official reporting of the first case. Also, there is strong impression to infer likely genetic adaptation of the virus in the continent that may account for the close clustering of isolates from different countries.

Foot-and-mouth disease (FMD) is a devastating animal disease. Differentiation of infected from vaccinated animals (DIVA) is very important for confirming suspected cases, evaluating the prevalence of infection, certifying animals for trade and controlling the disease. In this study, a competitive chemiluminescence immunoassay (3B-cCLIA) for DIVA was developed for the rapid detection of antibodies against non-structural proteins (NSPs) in different species of livestock animals using monoclonal antibody 9E2 as a competitive antibody that recognizes NSP 3B, which only needs to be washed once and takes 15 min. The cut-off (50%), diagnosis sensitivity (97.20%, 95.71%, and 96.15%) and diagnosis specificity (99.51%, 99.43%, and 98.36) of the assay were estimated by testing a panel of known background sera from swine, cattle and sheep. The accuracy rate of 3B-cCLIA was also validated and compared with that of two commercial diagnostic kits. The early diagnostic performance showed that antibodies to NSPs occurred later (approximately 1–2 days) than antibodies to structural proteins. Furthermore, NSP antibodies present in animals vaccinated multiple times (false-positive), especially in cattle and sheep, were confirmed, and the false-positive rate increased with the number of vaccinations. These results indicated that 3B-cCLIA is suitable to rapidly detect antibodies against FMDV NSP 3B in a wide range of species for DIVA.

Atypical porcine pestivirus (APPV), which has been confirmed to be associated with congenital tremor (CT) in pigs, is a newly discovered porcine virus that has been found in the Americas, Europe, and Asia; however, no report of APPV in Japan has been published. We identified an APPV in the central nervous system of Japanese piglets with CT, and firstly determined and analyzed the complete genome sequence. Phylogenetic analysis using the complete genome nucleotide sequence of the Japanese APPV, named Anna/2020, and those of APPVs from the NCBI database showed that APPVs were divided into three genotypes (genotypes 1 to 3), and that Anna/2020 clustered with the genotype 3 APPV strains, but distantly branched from these strains. Pairwise complete coding region nucleotide sequence comparisons revealed that there was 94.0% to 99.7% sequence identity among the genotype 3 strains, while Anna/2020 showed 87.0% to 89.3% identity to those genotype 3 strains, suggesting that Anna/2020 represents a novel APPV lineage within genotype 3. Retrospective examinations using RT-PCR revealed one genotype 1 and two novel genotype 3 APPVs from pigs without CT, and that novel genotype 3 APPVs have been prevalent in Japan since at least 2007.

COVID-19 is a complex disease and many difficulties are faced today especially in the proper choice of pharmacological treatments. The role of antiviral agents for COVID-19 is still being investigated. The evidence for immunomodulatory and anti-inflammatory drugs is quite conflicting, while the use of corticosteroids is supported by robust evidence. The use of heparins in hospitalized critically ill patients is preferred over other anticoagulants. Lastly, conflicting data were found regarding to the use of convalescent plasma and vitamin D. According to data shared by the WHO, many vaccines are under phase 3 clinical trials and some of them already received the marketing approval in EU countries and in the US. In conclusion, drugs repurposing has represented the main approach recently used in the treatment of patients with COVID-19. At this moment, the analysis of efficacy and safety data of drugs and vaccines used in real life context is strongly needed.