Senecavirus A (SVA) continues to cause vesicular lesions in swine in Canada and many regions worldwide. Since the vesicular lesions caused by SVA are similar to those caused by foot and mouth disease virus, swine vesicular disease virus and vesicular stomatitis virus, a foreign animal disease investigation must be initiated to rule out these diseases. SVA isolates from pigs displaying vesicular lesions in Canada from 2015 to 2023 were sequenced, and phylogeographic analysis was performed using the complete genome sequences. The results infer that SVA has spread between the United States and Canada several times. In addition, the results suggest that SVA spreads from different regions. SVA spread was inferred from Canada into Thailand, India and Mexico and inferred from the United States to Brazil, Columbia, Chile and China with ten separate introductions. Furthermore, recombination was observed in SVA genomes from Canada, the United States and China.

Foot-and-mouth disease (FMD) is one of the most infectious viral transboundary diseases of livestock, which causes devastating global economic losses. Different enzyme-linked immunosorbent assays (ELISAs) are used for sero-surveillance of the foot-and-mouth disease virus (FMDV). However, more sensitive, accurate, and convenient ELISAs are still required to detect antibodies against FMDV serotypes. The primary goal of this study was to establish serotype-specific monoclonal antibody (mAb)-based blocking ELISAs (mAb-bELISAs) that would provide better performance characteristics or be equivalent in performance characteristics compared with a conventional polyclonal antibody (pAb)-based competitive ELISA (pAb-cELISA). Four mAb-bELISAs were developed using FMDV serotype-specific mAbs for the detection of anti-FMDV/O/A/Asia1/SAT2 antibodies. Using a 50% cut-off, all four mAb-bELISAs exhibited species-independent 99.74%, 98.01%, 96.59%, and 98.55% diagnostic specificity (DSp) and 98.93%, 98.25%, 100%, and 87.50% diagnostic sensitivity (DSe) for FMDV serotypes O, A, Asia1, and SAT2, respectively. In addition, a 100% DSe of serotypes O- and SAT2-specific mAb-bELISAs was observed for porcine sera when the cut-off was 30%. All mAb-bELISAs developed in this study displayed high repeatability/reproducibility without cross-reactivity. Finally, the diagnostic performance of mAb-bELISAs was found to be better than or equivalent to compared with pAb-cELISAs, suggesting that mAb-bELISAs can be used to replace existing pAb-ELISAs for the detection of antibodies against these four FMDV serotypes.

Foot-and-mouth disease (FMD) is a devastating infectious viral disease of cloven-hoofed animals. Differentiating FMD from other vesicular diseases is difficult based on only clinical symptoms, requiring an appropriate laboratory diagnostic test. The double-antibody sandwich (DAS)-ELISA is a reliable diagnostic technique for antigen detection and serotyping of FMDV. However, classical DAS-ELISAs use polyclonal antibodies (pAbs), which are inconsistent in yields and limited in large-scale applications compared to hybridoma cell-secreted laboratory-made monoclonal antibodies (mAbs). Therefore, this study aimed to develop simplified and sensitive FMD serotype-specific DAS-ELISAs using HRP-conjugated mAbs and a TMB substrate. Six FMDV serotype-specific mAb-DAS-ELISAs were developed. All assays were optimized using BEI-inactivated FMD antigens. Real-time reverse-transcriptase PCR (RRT-PCR) was also used to verify the detection efficiency of all assays. Known negative and positive 10% tissue suspensions of different animal origins were examined to calculate the diagnostic specificity (DSp) and sensitivity (DSe). Serotype-specific mAb-DAS-ELISAs demonstrated 100%, 97%, 97%, 99%, 99%, and 94% DSp and 100%, 95%, 90%, 95%, 100%, and 100% DSe for serotypes O, A, Asia-1, SAT-1, SAT-2, and SAT-3, respectively. The detection efficiency of mAb-DAS-ELISAs was better than that of classical DAS-ELISAs. Also, all assays demonstrated minimal cross-reactivity and optimal reproducibility. Therefore, the mAb-DAS-ELISAs developed in this study could be useful for detecting and serotyping FMDV and ultimately replacing the classical DAS-ELISA.

During the past 25 years, vesicular stomatitis virus (VSV) has produced multiple outbreaks in the US, resulting in the emergence of different viral lineages. Currently, very little is known about the pathogenesis of many of these lineages, thus limiting our understanding of the potential biological factors favoring each lineage in these outbreaks. In this study, we aimed to determine the potential phenotypic differences between two VSV Indiana (VSIV) serotype epidemic strains using a pig model. These strains are representative of the epidemic lineages that affected the US between 1997 and 1998 (IN98COE) and between 2019 and 2020 (IN0919WYB2), the latter responsible for one of the most extensive outbreaks in the US. Our initial genome analysis revealed the existence of 121 distinct mutations between both strains, including the presence of a 14-nucleotide insertion in the intergenic region between the G and L genes observed in IN0919WYB2. The levels of viral RNA in clinical samples between pigs infected with IN98COE or IN0919WYB2 were compared. Overall, higher and prolonged expression of viral RNA in pigs infected with IN98COE was observed. However, clinically, IN0919WYB2 was slightly more virulent than IN98COE, as well as more efficient at producing infection through contact transmission. Additionally, infectious virus was recovered from more samples when the pigs were infected with IN0919WYB2, as revealed by virus isolation in cell culture, indicating the increased ability of this virus to replicate in pigs. Sequence analyses conducted from isolates recovered from both experimental groups showed that IN0919WYB2 produced more variability during the infection, denoting the potential of this strain to evolve rapidly after a single infection–contact transmission event in pigs. Collectively, the results showed that epidemic strains of VSIV may represent disparate phenotypes in terms of virulence/transmissibility for livestock, a situation that may impact the intensity of an epidemic outbreak. This study also highlights the relevance of pathogenesis studies in pigs to characterize phenotypic differences in VSV strains affecting livestock in the field.

Diagnostic testing of foot-and-mouth disease virus (FMDV) currently utilizes reverse transcription quantitative PCR (RT-qPCR) to detect the presence of viral RNA and double antibody sandwich ELISAs (DAS-ELISAs) to determine viral serotype. Serotype identification is critical to support informed vaccine selection to combat outbreaks. While DAS-ELISAs are capable of serotype identification, the test suffers from low sensitivity and requires a viral isolate for successful detection. In this study, we developed FMDV-ONTAPS: an Oxford Nanopore Technologies Amplicon P1 Sequencing protocol involving reverse transcription-PCR to amplify P1 of the FMDV genome, and Nanopore sequencing of the amplicons to provide genetic data for serotype and subtype/topotype identification. FMDV isolates representing all seven serotypes were successfully sequenced with this method. Additionally, the protocol successfully provided serotype identification from a variety of specimen matrices obtained from experimentally infected animals that included milk, serum, oral and nasal swabs, tissue suspensions, vesicular fluid, and oral fluid. The limit of detection for FMDV cell culture isolates was comparable for both sequencing and RT-qPCR detection. RT-qPCR Cq values for clinical samples evaluated ranged from 8 to 28.21. Sequencing was successful for all samples except for a single tissue suspension sample (Cq of 28.21). Identification of FMDV serotype in clinical samples is critical for effective outbreak response, and Nanopore sequencing offers a timelier and more sensitive alternative to DAS-ELISAs.