Vesicular Disease In Pigs Research Articles

Comparative transcriptomics analysis on Senecavirus A-infected and non-infected cells.

Senecavirus A (SVA) is an emerging virus that causes the vesicular disease in pigs, clinically indistinguishable from other high consequence vesicular diseases. This virus belongs to the genus Senecavirus in the family Picornaviridae. Its genome is a positive-sense, single-stranded RNA, approximately 7,300 nt in length, with a 3' poly(A) tail but without 5'-end capped structure. SVA can efficiently propagate in different cells, including some non-pig-derived cell lines. A wild-type SVA was previously rescued from its cDNA clone using reverse genetics in our laboratory. In the present study, the BSR-T7/5 cell line was inoculated with the passage-5 SVA. At 12 h post-inoculation, SVA-infected and non-infected cells were independently collected for the analysis on comparative transcriptomics. The results totally showed 628 differentially expressed genes, including 565 upregulated and 63 downregulated ones, suggesting that SVA infection significantly stimulated the transcription initiation in cells. GO and KEGG enrichment analyses demonstrated that SVA exerted multiple effects on immunity-related pathways in cells. Furthermore, the RNA sequencing data were subjected to other in-depth analyses, such as the single-nucleotide polymorphism, transcription factors, and protein-protein interactions. The present study, along with our previous proteomics and metabolomics researches, provides a multi-omics insight into the interaction between SVA and its hosts.

CircRNA_8521 promotes Senecavirus A infection by sponging miRNA-324 to regulate LC3A

Senecavirus A (SVA) causes outbreaks of vesicular disease in pigs, which imposes a considerable economic burden on the pork industry. As current SVA prevention measures are ineffective, new strategies for controlling SVA are urgently needed. Circular (circ)RNA is a newly characterized class of widely expressed, endogenous regulatory RNAs, which have been implicated in viral infection; however, whether circRNAs regulate SVA infection remains unknown. To investigate the influence of circRNAs on SVA infection in porcine kidney 15 (PK-15) cells, RNA sequencing technology was used to analyze the circRNA expression profiles of SVA-infected and uninfected PK-15 cells, the interactions between circRNAs, miRNAs, and mRNAs potentially implicated in SVA infection were predicted using bioinformatics tools. The prediction accuracy was verified using quantitative real-time (qRT)-PCR, Western blotting, as well as dual-luciferase reporter and RNA pull-down assays. The results showed that 67 circRNAs were differentially expressed as a result of SVA infection. We found that circ_8521 was significantly upregulated in SVA-infected PK-15 cells and promoted SVA infection. circ_8521 interacted with miR-324. miR-324 bound to LC3A mRNA which inhibited the expression of LC3A. Knockdown of LC3A inhibited SVA infection. However, circ_8521 promoted the expression of LC3A by binding to miR-324, thereby promoting SVA infection. We demonstrated that circ_8521 functioned as an endogenous miR-324 sponge to sequester miR-324, which promoted LC3A expression and ultimately SVA infection.

Construction and characterization of a full-length infectious clone of an emerging senecavirus A strain.

Senecavirus A (SVA) is an emerging virus that causes vesicular disease in pigs. Construction of a full-length SVA cDNA clone is crucial for understanding its replication and pathogenesis. Here, we successfully constructed a CMV-promoter-driven infectious cDNA clone of the SVA isolate SVA/GX/CH/2018, which we named rSVA GX01. Sequence comparison between the pSVA GX01 and the parental isolate (SVA/GX/CH/2018) revealed three single-nucleotide differences. Four-week-old piglets were experimentally infected with either the parental virus or the cloned virus. The results showed that the cloned rSVA GX01 displayed weak pathogenicity in 4-week-old pigs compared to the parental virus SVA CH-GX-01-2018. The infectious clone of SVA will serve as a valuable tool for studying the viral replication cycle and for functional analysis of the viral genome.

Real-time detection of Seneca Valley virus by one-tube RPA-CRISPR/Cas12a assay.

Senecavirus A (SVA) is a highly contagious virus that causes vesicular disease in pigs. At present, laboratory detection methods, such as virus isolation and polymerase chain reaction (PCR), required precision instruments and qualified personnel, making them unsuitable for point-of-care tests (POCT). Fortunately, the emergence of CRISPR/Cas system has provided new opportunities for fast and efficient pathogen detection. This study successfully developed a precise and sensitive detection platform for diagnosing SVA by combining the CRISPR system with recombinase polymerase amplification (RPA). The minimum detection limit of the assay was 10 copies of the SVA genome. Meanwhile, the assay demonstrated high specificity. To validate the effectiveness of this system, we tested 85 swine clinical samples and found that the fluorescence method had a 100% coincidence rate compared to RT-qPCR. Overall, the RPA-CRISPR/Cas12a assay established in our study is a highly effective method for detecting SVA and holds great potential for practical applications in the resource-limited settings.

Development and characterization of a recombinant Senecavirus A expressing enhanced green fluorescent protein.

Senecavirus A (SVA), belonging to the genus Senecavirus in the family Picornaviridae, is an emerging pathogen causing vesicular disease in pigs. The main clinical manifestations of SVA infection include high mortality in neonatal piglets, skin ulceration, and vesicular lesions. So far, there is no commercially available vaccines or drugs against SVA. Construction of SVA infectious clones carrying reporter genes will help understand the characteristics of SVA and promote vaccine development. In this study, we established a reverse genetics system for a local SVA isolate and used it to rescue a recombinant SVA, rSVA-eGFP, expressing the enhanced green fluorescent protein (eGFP) by inserting eGFP, GSG linker and the P2A sequence between 2A and 2B genes. We found that rSVA-eGFP exhibited a high replication efficiency comparable to the parental virus, was able to express the eGFP reporter efficiently and stable in maintaining the reporter gene up to six rounds of serial passages in BHK-21 cells. In mice, rSVA-eGFP also showed similar replication kinetics and pathogenicity to the parental virus, both causing mild lung lesions. In addition, a high-throughput viral neutralization assay was developed using eGFP as a surrogate readout in a fluorescence-based direct titration (FBT) assay based on rSVA-eGFP, facilitating rapid and accurate determination of the neutralizing antibody (nAb) titers. The successful establishment of an SVA reverse genetics system and the rescue of rSVA-eGFP would create a powerful tool for future studies of SVA replication mechanisms and pathogenicity as well as for antiviral development.

Construction and immunogenicity of Senecavirus A Virus-Like Particle Vaccine with adjuvant

Senecavirus A (SVA) is constantly associated with vesicular disease in pigs, and the clinical symptoms of pig infection with SVA are indistinguishable from other porcine vesicular diseases. Vaccine is one of the best methods to eliminate and control the spread of SVA. Virus-like particles (VLPs) can play important roles in prevention for infectious diseases. Here, the SVA VLPs was assembled by the baculovirus expression vector system, and the immunogenicity of the SVA VLPs mixed with different adjuvants were evaluated in mice and pigs. Two recombinant baculoviruses (rPFBD-VP1-VP3 and rPFBD-VP2-VP4) were constructed, which co-infected with Sf9 suspension cells to assemble SVA VLPs successfully. SVA VLPs mixed with ISA201 adjuvant and ISA201+Poly(I:C) adjuvant produced higher levels of neutralizing antibody, specific antibody (total IgG, IgG1, IgG2a and IgG2b) and cytokines in the T cells. And there was no significant difference between SVA VLPs+201 group and SVA VLPs+Poly(I:C)+201 group. Pigs immunized with high dose of SVA VLPs mixed with ISA201 adjuvant could produce higher titers of neutralizing antibody and SVA-specific antibody. Furthermore, the protection rates of SVA VLPs-H and SVA VLPs-L were 100% and 80%, and the viral load of SVA VLPs-H group is the lowest in all SVA VLPs groups. It is the first time to develop the SVA VLPs using the baculovirus expression vector system, which may lay the foundation for the research and development of SVA vaccine.

Experimental Seneca Valley virus infection in sows and their offspring

Neonatal mortality has been increasingly reported on swine breeding farms experiencing swine idiopathic vesicular disease (SIVD) outbreaks, which can be accompanied by lethargy, diarrhea, and neurologic signs in neonates. Seneca Valley Virus (SVV), or Senecavirus A, has been detected in clinical samples taken from pigs with SIVD. Experimental SVV inoculation has caused vesicular disease in pigs, particularly during the stages from weaning to finishing. However, it remains crucial to investigate whether SVV directly contributes to the increase in neonatal mortality rates. The following study was conducted to chronicle the pathogenesis of SVV infection in sows and their offspring. Ten sows were intranasally inoculated with 4.75 × 107 plaque-forming units of the virus per sow either late in gestation (n = 5) or within fourteen days of farrowing (n = 5). Each sow replicated SVV following intranasal inoculation, but only one out of ten sows developed a vesicular lesion on the snout. Evidence of transplacental infection was observed in two litters, and an additional two litters became infected following parturition out of five litters from sows inoculated in late gestation. No clinical signs were observed in the infected neonates. Likewise, no clinical signs were observed in the other five litters inoculated after farrowing, although each piglet did replicate the challenge virus. In this study, the experimental challenge of SVV did not result in neonatal mortality in contrast to observations in the field; however, it has shed light on the pathogenesis of the virus, the transmission of SVV between sows and their offspring, and host immune response that can help shape control measures in the field.

Seneca Valley virus 3C protease cleaves OPTN (optineurin) to Impair selective autophagy and type I interferon signaling

ABSTRACT Seneca Valley virus (SVV) causes vesicular disease in pigs, posing a threat to global pork production. OPTN (optineurin) is a macroautophagy/autophagy receptor that restricts microbial propagation by targeting specific viral or bacterial proteins for degradation. OPTN is degraded and cleaved at glutamine 513 following SVV infection via the activity of viral 3C protease (3C[pro]), resulting in N-terminal and a C-terminal OPTN fragments. Moreover, OPTN interacts with VP1 and targets VP1 for degradation to inhibit viral replication. The N-terminal cleaved OPTN sustained its interaction with VP1, whereas the degradation capacity targeting VP1 decreased. The inhibitory effect of N-terminal OPTN against SVV infection was significantly reduced, C-terminal OPTN failed to inhibit viral replication, and degradation of VP1 was blocked. The knockdown of OPTN resulted in reduced TBK1 activation and phosphorylation of IRF3, whereas overexpression of OPTN led to increased TBK1-IRF3 signaling. Additionally, the N-terminal OPTN diminished the activation of the type I IFN (interferon) pathway. These results show that SVV 3C[pro] targets OPTN because its cleavage impairs its function in selective autophagy and type I IFN production, revealing a novel model in which the virus develops diverse strategies for evading host autophagic machinery and type I IFN response for survival. Abbreviations: Co-IP: co-immunoprecipitation; GFP-green fluorescent protein; hpi: hours post-infection; HRP: horseradish peroxidase; IFN: interferon; IFNB/IFN-β: interferon beta; IRF3: interferon regulatory factor 3; LIR: LC3-interacting region; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MOI: multiplicity of infection; OPTN: optineurin; PBS: phosphate-buffered saline; SVV: Seneca Valley virus; SQSTM1: sequestosome 1; TAX1BP1: Tax1 binding protein 1; TBK1: TANK binding kinase 1; TCID50: 50% tissue culture infectious doses; UBAN: ubiquitin binding in TNIP/ABIN (TNFAIP3/A20 and inhibitor of NFKB/NF-kB) and IKBKG/NEMO; UBD: ubiquitin-binding domain; ZnF: zinc finger.

Seneca Valley Virus Degrades STING via PERK and ATF6-Mediated Reticulophagy.

Seneca Valley Virus (SVV), a member of the Picornaviridae family, is an emerging porcine virus that can cause vesicular disease in pigs. However, the immune evasion mechanism of SVV remains unclear, as does its interaction with other pathways. STING (Stimulator of interferon genes) is typically recognized as a critical factor in innate immune responses to DNA virus infection, but its role during SVV infection remains poorly understood. In the present study, we observed that STING was degraded in SVV-infected PK-15 cells, and SVV replication in the cells was affected when STING was knockdown or overexpressed. The STING degradation observed was blocked when the SVV-induced autophagy was inhibited by using autophagy inhibitors (Chloroquine, Bafilomycin A1) or knockdown of autophagy related gene 5 (ATG5), suggesting that SVV-induced autophagy is responsible for STING degradation. Furthermore, the STING degradation was inhibited when reticulophagy regulator 1 (FAM134B), a reticulophagy related receptor, was knocked down, indicating that SVV infection induces STING degradation via reticulophagy. Further study showed that in eukaryotic translation initiation factor 2 alpha kinase 3 (PERK)/activating transcription factor 6 (ATF6) deficient cells, SVV infection failed to induce reticulophagy-medaited STING degradation, indicating that SVV infection caused STING degradation via PERK/ATF6-mediated reticulophagy. Notably, blocking reticulophagy effectively hindered SVV replication. Overall, our study suggested that SVV infection resulted in STING degradation via PERK and ATF6-mediated reticulophagy, which may be an immune escape strategy of SVV. This finding improves the understanding of the intricate interplay between viruses and their hosts and provides a novel strategy for the development of novel antiviral drugs.

Seneca Valley virus 3Cpro antagonizes type I interferon response by targeting STAT1-STAT2-IRF9 and KPNA1 signals.

Type I interferon (IFN) signaling plays a principal role in host innate immune responses against invading viruses. Viruses have evolved diverse mechanisms that target the Janus kinase-signal transducer and activator of transcription (STAT) signaling pathway to modulate IFN response negatively. Seneca Valley virus (SVV), an emerging porcine picornavirus, has received great interest recently because it poses a great threat to the global pork industry. However, the molecular mechanism by which SVV evades host innate immunity remains incompletely clear. Our results revealed that SVV proteinase (3Cpro) antagonizes IFN signaling by degrading STAT1, STAT2, and IRF9, and cleaving STAT2 to escape host immunity. SVV 3Cpro also degrades karyopherin 1 to block IFN-stimulated gene factor 3 nuclear translocation. Our results reveal a novel molecular mechanism by which SVV 3Cpro antagonizes the type I IFN response pathway by targeting STAT1-STAT2-IRF9 and karyopherin α1 signals, which has important implications for our understanding of SVV-evaded host innate immune responses.

Senecavirus A seroprevalence and risk factors in United States pig farms.

Senecavirus A (SVA) is a non-enveloped, single-stranded, positive-sense RNA virus belonging to the Picornaviridae family. Senecavirus A is constantly associated with outbreaks of vesicular disease in pigs and has been reported in several countries since its first large-scale outbreak in 2014. Senecavirus A's clinical disease and lesions are indistinguishable from other vesicular foreign animal diseases (FAD). Therefore, an FAD investigation needs to be conducted for every SVA case. For this reason, SVA has been attributed as the cause of an alarming increase in the number of yearly FAD investigations performed by the United States Department of Agriculture (USDA). The objectives of this study were to estimate the seroprevalence of SVA antibodies in breeding and growing pig farms in the United States and to determine the farm-level risk factors associated with seropositivity. A total of 5,794 blood samples were collected from 98 and 95 breeding and growing pig farms in 17 states. A farm characteristics questionnaire was sent to all farms, to which 80% responded. The responses were used to conduct logistic regression analyses to assess the risk factors associated with SVA seropositivity. The estimated farm-level seroprevalences were 17.3% and 7.4% in breeding and growing pig farms, respectively. Breeding farms had 2.64 times higher odds of SVA seropositivity than growing pig farms. One key risk factor identified in breeding farms was the practice of rendering dead animal carcasses. However, the adoption of a higher number of farm biosecurity measures was associated with a protective effect against SVA seropositivity in breeding farms.

Senecavirus A Entry Into Host Cells Is Dependent on the Cholesterol-Mediated Endocytic Pathway.

Senecavirus A (SVA), an important member of the Picornaviridae family, causes vesicular disease in pigs. Here, we generated an EGFP-expressing recombinant SVA re-SVA-EGFP, which exhibited similar growth kinetics to its parental virus. The reporter SVA was used to study the role of pig ANTXR1 (pANTXR1) in SVA infection in a porcine alveolar macrophage cell line (PAM-Tang cells). Knockdown of the pANTXR1 significantly reduced SVA infection and replication in PAM-Tang cells, while re-expression of the pANTXR1 promoted the cell susceptibility to SVA infection. The results indicated that pANTXR1 is a crucial receptor mediating SVA infection. Subsequently, the viral endocytosis pathways for SVA entry into pig cells were investigated and the results showed that cholesterol played an essential role in receptor-mediated SVA entry. Together, these results demonstrated that SVA entered into host cells through the pANTXR1-mediated cholesterol pathway. Our findings provide potential targets to develop antiviral drugs for the prevention of SVA infection in the pig population.

Disruption of anthrax toxin receptor 1 in pigs leads to a rare disease phenotype and protection from senecavirus A infection

Senecavirus A (SVA) is a cause of vesicular disease in pigs, and infection rates are rising within the swine industry. Recently, anthrax toxin receptor 1 (ANTXR1) was revealed as the receptor for SVA in human cells. Herein, the role of ANTXR1 as a receptor for SVA in pigs was investigated by CRISPR/Cas9 genome editing. Strikingly, ANTXR1 knockout (KO) pigs exhibited features consistent with the rare disease, GAPO syndrome, in humans. Fibroblasts from wild type (WT) pigs supported replication of SVA; whereas, fibroblasts from KO pigs were resistant to infection. During an SVA challenge, clinical symptoms, including vesicular lesions, and circulating viremia were present in infected WT pigs but were absent in KO pigs. Additional ANTXR1-edited piglets were generated that were homozygous for an in-frame (IF) mutation. While IF pigs presented a GAPO phenotype similar to the KO pigs, fibroblasts showed mild infection, and circulating SVA nucleic acid was decreased in IF compared to WT pigs. Thus, this new ANTXR1 mutation resulted in decreased permissiveness of SVA in pigs. Overall, genetic disruption of ANTXR1 in pigs provides a unique model for GAPO syndrome and prevents circulating SVA infection and clinical symptoms, confirming that ANTXR1 acts as a receptor for the virus.

Recombinase polymerase amplification assay for rapid detection of Seneca Valley Virus

Seneca Valley virus (SVV) is related to vesicular disease in pigs, and its clinical symptoms are indistinguishable from other notifiable clinical symptoms of vesicular disease such as foot-and-mouth disease. The rapid and accurate detection of SVV is essential to confirm the pathogenic factors and initiate the implementation of control measures. The development of a rapid, simple, convenient, and low-cost molecular (nucleic acid amplification) test that can be used at the sample collection point has been identified as a key component for controlling SVV. This study describes the development and demonstration of recombinase polymerase amplification (RPA) test targeting the conserved regions of SVV for detection of SVV. The Primers and probes designed by us have shown good sensitivity and specificity in RPA test, which is helpful for RPA to be an effective tool for rapid diagnosis of SVV.

Construction of eGFP-Tagged Senecavirus A for Facilitating Virus Neutralization Test and Antiviral Assay.

Senecavirus A (SVA), also known as Seneca Valley virus, is an emerging virus that causes vesicular disease in pigs. This virus belongs to the genus Senecavirus in the family Picornaviridae. The SVA CH-LX-01-2016 was isolated from Guangdong Province of China in 2016. In this study, a recombinant SVA CH-LX-01-2016 was constructed using reverse genetics, and proven to be able to express efficiently an enhanced green fluorescent protein (eGFP) in vitro. This eGFP-tagged recombinant SVA (rSVA-eGFP) exhibited a high capacity for viral replication. Its fluorescence-tracked characteristics greatly facilitated both virus neutralization test (VNT) and antiviral assay. The rSVA-eGFP-based VNT was used to detect eight porcine serum samples, out of which four were determined to be neutralization titer-positive. Subsequently, two antiviral drugs, ribavirin and apigenin, were assayed for evaluating both effects against the rSVA-eGFP in vitro. The result showed that only the ribavirin exhibited an anti-SVA activity.

Analysis of the Performance of the Animal Health Surveillance System in the Outbreak of Swine Vesicular Disease in the State of Santa Catarina - Brazil

Background: The occurrence of vesicular disease associated with Senecavirus A in a pig-producing region of Santa Catarina increased in 2015, reflected by the number of syndromic notifications to the official animal health service. In view of the recurrence of this event in 2018, the objectives of this study were to analyze the official data related to cases of suspected vesicular disease in pigs and to evaluate whether the experience in conducting the investigations of 2015 was incorporated into the years subsequent to 2015. We addressed this goal by analysis of the performance parameters of the state animal health surveillance system.Materials, Methods & Results: Descriptive analyses of data from official investigations of suspected vesicular disease in swine in different regions were carried out, and statistical models were used to: i) test the effect of the year on the age of the investigated injuries; ii) assess whether there was an association between the year and the type of outcome of the official investigation (discarded case or a probable case of vesicular disease, which resulted in the collection of samples for laboratory diagnosis and interdiction of the affected properties); iii) evaluate whether there was an association between the year and the detection of Senecavirus A RNA among the molecular analyses carried out after a case was classified as probable vesicular disease. From 05/22/2015 to 03/28/2019, there were 2093 notifications of suspected vesicular disease in pigs to the official service of Santa Catarina, with 1538 (73.5%) occurring in 2015 and 555 (26.5%) in subsequent years. After 2015, when compared to the base year, the chances of detecting late vesicular lesions (>3 days) were similar (increased 1.11 times, but there was no statistically significant association), in view of a panorama in which 55.29% of cases had the lesions classified as late throughout the analyzed period. The variation in the odds was relatively homogeneous among the regional units (Intraclass Correlation Coefficient [ICC] = 10.9%), but in the São Miguel do Oeste unit, it was significantly lower than the average. It was also lower than the average, in regional units with a high ratio of properties with pigs attended by the official veterinarian. The chances of cases being considered probable increased 32.3 times, but the descriptive analysis of the average interdiction period decreased. The estimated ICC was 34.9%, and in Campos Novos and Caçador, the chances of cases being probable were significantly higher than the average, while in the Canoinhas unit, the chances were significantly lower. The prevalence of cases with molecular detection of Senecavirus A was 78% lower after 2015. Discussion: Despite the increased sensitivity of the surveillance process for suspected vesicular events in the years after 2015, the specificity of the molecular diagnosis of Senecavirus A decreased. This is likely, because there was difficulty in sampling lesions with vesicular fluid and/or when the lesion epithelium had recently been disrupted, conditions most conducive to the identification of viral genetic material. In this context, the western region of the state, the main pig producer, continued to be the most accurate in confirming cases of Senecavirus A. Even with the improvements of laboratory diagnosis and the reduction of the interdiction period, it was important to standardize the differentiation of a late infectious lesion from a traumatic injury, as well as any association with other sample analyses, in order to minimize the disorders linked to the mischaracterization of a manifestation of Senecavirus A, without compromising the syndromic focus of its vesicular character.

Senecavirus A: An emerging pathogen causing vesicular disease in pigs

Senecavirus A (SVA) is a single representative species of the Senecavirus genus within the family Picornaviridae. This review presents the current knowledge regarding SVA epidemiology, transmission, pathogenesis, clinical signs, differential diagnosis and control measures. SVA is not debilitating, but significant because of its resemblance to acute, highly contagious and economically devastating viral diseases, such as FMD. The incubation period of SVA is 4-5 days, the viremia period is short, lasting 3 to 10 days post infection (dpi). SVA shedding lasts up to 28 days. SVA can be shed by oral and nasal secretions and by faeces. The virus excretion peak occurs between 1 and 5 dpi, especially in oral secretions, which contain higher virus loads relative to nasal secretions and faeces. SVA lesions are found most frequently on the snout, lips and tongue, as well as on hooves, specifically, on coronary bands, dewclaws, hoof pads and in interdigital space. The vesicles quickly rupture to form ulcers that may be covered by serofibrinous exudates. The ulcers begin to repair in 7 days, and the regeneration of epithelium is usually complete within 2 weeks. Since clinical lesions induced by SVA are indistinguishable from those observed in other vesicular diseases of swine, accurate and reliable laboratory differential diagnosis is critical to the precise identification of the infectious agent. SVA has potential cytolytic activity and high selectivity for tumour cell lines with neuroendocrine properties versus adult normal cells. Because of its potential oncolytic activity, the virus can be useful in human cancer therapy. The example of SVA shows that the risk of emerging infectious diseases in swine populations is high and that emerging diseases of swine have significant potential impact on the productivity and economics of the pork industry. The SVA infection is currently limited to the United States, Canada, Brazil, China and Thailand. However, descriptions of the SVA infection in Asia suggest that the virus is not restricted to a specific geographic region and may be distributed on a global scale in the future.

A ten years (2007-2016) retrospective serological survey for Seneca Valley virus infection in major pig producing states of Brazil.

Seneca Valley virus (SVV) is the etiological agent of vesicular disease in pigs, clinically indistinguishable of classical viral vesicular infections, including foot-and-mouth disease. The first outbreaks of SVV infection in Brazil were reported in 2014. However, it was not known whether the virus was circulating in Brazilian pig herds before this year. This study is a retrospective serological investigation of porcine health status to SVV in Brazil. Serum samples (n=594) were grouped in before (2007-2013, n=347) and after (2014-2016, n=247) SVV outbreaks in Brazil. Twenty-three pig herds were analyzed, of which 19 and 4 were sampled before and after the beginning of SVV outbreaks, respectively. Two herds sampled after 2014 presented animals with SVV-associated clinical manifestations, while the other two housed asymptomatic pigs. Anti-SVV antibodies were evaluated by virus neutralization test. The results demonstrated that pig herds of different Brazilian geographical regions and distinct pig categories were negative to anti-SVV antibodies in sera obtained before 2014. Antibodies to SVV were detected only in serum samples obtained after 2014, particularly in herds with the presence of pigs with SVV-clinical signs. These results present robust serological evidence that the SVV was not present in the major Brazilian pig producing regions prior to 2014.

Generation, characterization, and application in serodiagnosis of recombinant swine vesicular disease virus-like particles.

Swine vesicular disease (SVD) is a highly contagious viral disease that causes vesicular disease in pigs. The importance of the disease is due to its indistinguishable clinical signs from those of foot-and-mouth disease, which prevents international trade of swine and related products. SVD-specific antibody detection via an enzyme-linked immunosorbent assay (ELISA) is the most versatile and commonly used method for SVD surveillance and export certification. Inactivated SVD virus is the commonly used antigen in SVD-related ELISA. A recombinant SVD virus-like particle (VLP) was generated by using a Bac-to-Bac baculovirus expression system. Results of SVD-VLP analyses from electron microscopy, western blotting, immunofluorescent assay, and mass spectrometry showed that the recombinant SVD-VLP morphologically resemble authentic SVD viruses. The SVD-VLP was evaluated as a replacement for inactivated whole SVD virus in competitive and isotype-specific ELISAs for the detection of antibodies against SVD virus. The recombinant SVD-VLP assay produced results similar to those from inactivated whole virus antigen ELISA. The VLP-based ELISA results were comparable to those from the virus neutralization test for antibody detection in pigs experimentally inoculated with SVD virus. Use of the recombinant SVD-VLP is a safe and valuable alternative to using SVD virus antigen in diagnostic assays.

Detection of pseudocowpox virus in water buffalo (Bubalus bubalis) with vesicular disease in the state of São Paulo, Brazil, in 2016

ABSTRACTBackground: Parapoxviruses are zoonotic viruses that infect cattle, goats and sheep; there have also been reports of infections in camels, domestic cats and seals.Objective: The objective of this report was to describe a case of vesicular disease caused by pseudocowpox virus (PCPV) in water buffalo (Bubalus bubalis) in Brazil.Animals: Sixty buffalo less than 6 months old exhibited ulcers and widespread peeling of the tongue epithelium. There were no cases of vesicular disease in pigs or horses on the same property.Methods: Samples were analysed by PCR and sequencing. Phylogenetic analysis in MEGA 7.01 was reconstructed using major envelope protein (B2L) by the Tamura three-parameter nucleotide substitution model and the maximum likelihood and neighbor joining models, both with 1000 bootstrap replicates. The genetic distance between the groups was analysed in MEGA using the maximum composite likelihood model. The rate variation among sites was modeled using gamma distribution.Results: The presence of PCPV in the buffalo herd could be demonstrated in epithelium and serum. The minimum genetic distance between the isolated PCPV strain (262-2016) and orf virus and bovine papular stomatitis virus was 6.7% and 18.4%, respectively. The maximum genetic distance calculated was 4.6% when compared with a PCPV detected in a camel.Conclusions/Clinical Importance: The peculiar position of the isolated strain in the phylogenetic trees does not necessarily indicate a different kind of PCPV that infects buffalo. More samples from cattle and buffalo in Brazil must be sequenced and compared to verify if PCPV from buffalo are genetically different from samples derived from cattle.