Surface display of the COE antigen of porcine epidemic diarrhoea virus on Bacillus subtilis spores.

Porcine epidemic diarrhoea virus (PEDV) infects pigs of all ages by invading small intestine, causing acute diarrhoea, vomiting, and dehydration with high morbidity and mortality among newborn piglets. However, current PEDV vaccines are not effective to protect the pigs from field epidemic strains because of poor mucosal immune response and strain variation. Therefore, it is indispensable to develop a novel oral vaccine based on epidemic strains. Bacillus subtilis spores are attractive delivery vehicles for oral vaccination on account of the safety, high stability, and low cost. In this study, a chimeric gene CotC-Linker-COE (CLE), comprising of the B. subtilis spore coat gene cotC fused to the core neutralizing epitope CO-26 K equivalent (COE) of the epidemic strain PEDV-AJ1102 spike protein gene, was constructed. Then recombinant B. subtilis displaying the CLE on the spore surface was developed by homologous recombination. Mice were immunized by oral route with B. subtilis 168-CLE, B. subtilis 168, or phosphate-buffered saline (PBS) as control. Results showed that the IgG antibodies and cytokine (IL-4, IFN-γ) levels in the B. subtilis 168-CLE group were significantly higher than the control groups. This study demonstrates that B. subtilis 168-CLE can generate specific systemic immune and mucosal immune responses and is a potential vaccine candidate against PEDV infection.

Monitoring emerging pathogens using negative nucleic acid test results from endemic pathogens in pig populations: Application to porcine enteric coronaviruses.

This study evaluated the use of endemic enteric coronaviruses polymerase chain reaction (PCR)-negative testing results as an alternative approach to detect the emergence of animal health threats with similar clinical diseases presentation. This retrospective study, conducted in the United States, used PCR-negative testing results from porcine samples tested at six veterinary diagnostic laboratories. As a proof of concept, the database was first searched for transmissible gastroenteritis virus (TGEV) negative submissions between January 1st, 2010, through April 29th, 2013, when the first porcine epidemic diarrhea virus (PEDV) case was diagnosed. Secondly, TGEV- and PEDV-negative submissions were used to detect the porcine delta coronavirus (PDCoV) emergence in 2014. Lastly, encountered best detection algorithms were implemented to prospectively monitor the 2023 enteric coronavirus-negative submissions. Time series (weekly TGEV-negative counts) and Seasonal Autoregressive-Integrated Moving-Average (SARIMA) were used to control for outliers, trends, and seasonality. The SARIMA's fitted and residuals were then subjected to anomaly detection algorithms (EARS, EWMA, CUSUM, Farrington) to identify alarms, defined as weeks of higher TGEV-negativity than what was predicted by models preceding the PEDV emergence. The best-performing detection algorithms had the lowest false alarms (number of alarms detected during the baseline) and highest time to detect (number of weeks between the first alarm and PEDV emergence). The best-performing detection algorithms were CUSUM, EWMA, and Farrington flexible using SARIMA fitted values, having a lower false alarm rate and identified alarms 4 to 17 weeks before PEDV and PDCoV emergences. No alarms were identified in the 2023 enteric negative testing results. The negative-based monitoring system functioned in the case of PEDV propagating epidemic and in the presence of a concurrent propagating epidemic with the PDCoV emergence. It demonstrated its applicability as an additional tool for diagnostic data monitoring of emergent pathogens having similar clinical disease as the monitored endemic pathogens.

Genotypic characterization of novel S-DEL variants of porcine epidemic diarrhea virus identified in South Korea.

The highly pathogenic genotype 2b (HP-G2b) of porcine epidemic diarrhea virus (PEDV), which caused a pandemic in 2013-2014, evolved in South Korea and became endemic, affecting the domestic pig industry. This study describes the genotypic traits of novel HP-G2b PEDV strains identified on affected farms experiencing low disease severity with < 10% neonatal mortality. Nucleotide sequencing revealed common deletion patterns, termed S-DEL2, resulting in a two-amino-acid deletion at positions 60 and 61, 61 and 62, or 63 and 64 in the N-terminal domain of the spike (S) protein of all isolates. The S barcode profiles of S-DEL2 variants differed from each other and shared 96.0-99.4% and 98.5-99.6% nt sequence identity with other South Korean HP-G2b PEDV strains in the S gene and in the complete genome sequence, respectively. Genetic and phylogenetic analysis showed that the S-DEL2 strains belonged to diverse domestic clades: CK, CK.1, CK.2, or NC. The emergence of novel S-DEL2 strains suggests that continuous evolution of PEDV occurs under endemic circumstances, resulting in genetic diversity and distinct clinical presentations. This study advances our knowledge regarding the genetic and pathogenic heterogeneity of PEDV and emphasizes the importance of active monitoring and surveillance to identify novel variants and determine their genotypic and phenotypic characteristics.

Porcine deltacoronavirus nonstructural protein 2 inhibits type I and III IFN production by targeting STING for degradation.

Porcine deltacoronavirus (PDCoV) is an enteropathogenic coronavirus that has been reported to use various strategies to counter the host antiviral innate immune response. The cGAS-STING signalling pathway plays an important role in antiviral innate immunity. However, it remains unclear whether PDCoV achieves immune evasion by regulating the cGAS-STING pathway. Here, we demonstrated that the nonstructural protein 2 (nsp2) encoded by PDCoV inhibits cGAS-STING-mediated type I and III interferon (IFN) responses via the regulation of porcine STING (pSTING) stability. Mechanistically, ectopically expressed PDCoV nsp2 was found to interact with the N-terminal region of pSTING. Consequently, pSTING was degraded through K48-linked ubiquitination and the proteasomal pathway, leading to the disruption of cGAS-STING signalling. Furthermore, K150 and K236 of pSTING were identified as crucial residues for nsp2-mediated ubiquitination and degradation. In summary, our findings provide a basis for elucidating the immune evasion mechanism of PDCoV and will contribute to the development of targets for anti-coronavirus drugs.

Tris stabilized AuNPs based lateral flow immunochromatography for the simultaneous detection of porcine epidemic diarrhea virus and rotavirus on-site.

Porcine epidemic diarrhea virus (PEDV) and rotavirus has posed a significant threat to the pig industry annually across different nations, resulting in huge economic losses. The frequent co-infection of these two viruses in clinical settings complicates the process of differential diagnoses. Rapid and accurate detection of PEDV and rotavirus is in great demand for timely diarrhea disease prevention and control. In this study, tris stabilized AuNPs were prepared and a sensitive lateral flow immunoassay (LFIA) sensor was developed for the simultaneous and rapid detection of PEDV and rotavirus on site. After the system optimization, the established LFIA can simultaneously identify PEDV and rotavirus with limits of detection (LOD) of 1.25 × 103 TCID50 mL-1 and 3.13 × 102 pg mL-1, respectively. When applying for clinical samples, the LFIA show a concordance of 95 % and 100 % to reverse transcript polymerase chain reaction (RT-PCR) for PEDV and rotavirus respectively. Therefore, this LFIA can qualitatively detect PEDV and rotavirus in 18 min with high sensitivity and accuracy without any sophisticated equipment and operation, making it a promising candidate for the early diagnosis of PEDV or/and rotavirus diarrhea on site.

Isolation and phylogenetic analysis of avian infectious bronchitis virus from an imported chicken meat product in Malaysia.

Avian infectious bronchitis (IB), a Gammacoronavirus, is a highly contagious upper respiratory disease, affecting chickens of all ages with a significant economic threat to the poultry industry. In February 2020, a specimen of imported chicken meat product was received and requested for coronavirus testing. The result was positive for the avian coronavirus, the IB virus (IBV) by molecular detection in the pre-screening test. Thus, this study aimed to isolate and characterize the IBV from the specimen. Virus isolation via egg inoculation was attempted and IBV was successfully isolated. The S1 subunit of the spike (S) gene of the IBV was amplified, sequenced, and the Basic Local Alignment Search Tool (BLAST) analysis showed that the IBV has 99% and 98% nucleotide similarity with the Malaysian and China IBVs, respectively. The phylogenetic analysis indicated that the virus belongs to the GI-19 lineage (also known as the QX strain) and is grouped with other IBVs from Malaysia and China. The GI-19 lineage is one of the primary IB strains that circulate in Malaysia. The recovery of the virus may be due to the persistence characteristic of the virus on meat; and the cold chain practices in the imported food product prolong the survival of this coronavirus. Though IBV is not identified as a hazard in chicken meat or meat products, raw food should be cooked thoroughly before being consumed. With the increase in international trade in poultry and poultry products, disease screening at the entry point and import risk analysis is crucial to ensure food safety and prevent the introduction of new viruses into Malaysia.

Protection Against Infectious Bronchitis Virus Vaccine Recombinants and Chicken-Selected Vaccine Subpopulations.

Outbreaks of infectious bronchitis (IB) continue to occur from novel variants of IB virus (IBV) emerging from selection of vaccine subpopulations and/or naturally occurring recombination events. S1 sequencing of Arkansas (Ark) -type viruses obtained from clinical cases in Alabama broilers and backyard chickens shows both Ark Delmarva Poultry Industry (ArkDPI) vaccine subpopulations as well as Ark vaccine viruses showing recombination with other IB vaccine viruses. IB Ark-type isolates AL5, most similar to an ArkDPI vaccine subpopulation selected in chickens, AL4, showing a cluster of three nonsynonymous changes from ArkDPI subpopulations selected in chickens, and AL9, showing recombination with Massachusetts (Mass) -type IBV, were examined for pathogenicity and ability to break through immunity elicited by vaccination with a commercial ArkDPI vaccine. Analysis of predicted S1 protein structures indicated the changes were in regions previously shown to comprise neutralizing epitopes. Thus, they were expected to contribute to immune escape and possibly virulence. Based on clinical signs, viral load, and histopathology, all three isolates caused disease in naïve chickens, although AL9 and AL5 viral loads in trachea were statistically significantly higher (30- and 40-fold) than AL4. S1 gene sequencing confirmed the stability of the relevant changes in the inoculated viruses in the chickens, although virus in some individual chickens exhibited additional S1 changes. A single amino acid deletion in the S1 NTD was identified in some individual chickens. The location of this deletion in the predicted structure of S1 suggested the possibility that it was a compensatory change for the reduced ability of AL4 to replicate in the trachea of naïve chickens. Chickens vaccinated with a commercial ArkDPI vaccine at day of hatch and challenged at 21 days of age showed that vaccination provided incomplete protection against challenge with these viruses. Moreover, based on viral RNA copy numbers in trachea, differences were detected in the ability of the vaccine to protect against these IBV isolates, with the vaccine protecting the most poorly against AL4. These results provide additional evidence supporting that IBV attenuated vaccines, especially ArkDPI vaccines, contribute to perpetuating the problem of IB in commercial chickens.

Protección contra los virus de la bronquitis infecciosa vacunales recombinantes y las subpoblaciones de vacunas seleccionadas en pollos. Los brotes de la bronquitis infecciosa aviar continúan presentándose a partir de nuevas variantes de dicho virus, que surgen de la selección de subpoblaciones de vacunas y/o eventos de recombinación que ocurren naturalmente. La secuenciación del gene S1 de virus tipo Arkansas (Ark) obtenidos de casos clínicos en pollos de engorde y de traspatio de Alabama muestra que tanto las subpoblaciones de la cepa vacunal Arkansas Delmarva Poultry Industry (ArkDPI) así como los virus de la vacuna Arkansas muestran recombinación con otros virus vacunales de la bronquitis infecciosa. Los aislamientos del virus de la bronquitis infecciosa Arkansas tipo "AL5", más similares a una subpoblación de vacuna ArkDPI seleccionada en pollos, "AL4", que muestra un grupo de tres cambios no sinónimos de subpoblaciones de ArkDPI seleccionadas en pollos y el tipo "AL9", que muestra recombinación con el serotipo Massachusetts, se examinaron para determinar su patogenicidad y capacidad para traspasar la inmunidad generada por la vacunación con una vacuna comercial ArkDPI. El análisis de las estructuras predichas de la proteína S1 indicó que los cambios se produjeron en regiones que previamente se había demostrado comprendían epítopos neutralizantes. Por lo tanto, se esperaba que contribuyeran al escape inmunológico y posiblemente a la virulencia. Con base en los signos clínicos, la carga viral y la histopatología, los tres aislados causaron enfermedad en pollos sin exposición previa, aunque las cargas virales de AL9 y AL5 en la tráquea fueron estadísticamente significativamente mayores (30 y 40 veces) en comparación con AL4. La secuenciación del gene S1 confirmó la estabilidad de los cambios relevantes en los virus inoculados en los pollos, aunque el virus en algunos pollos individuales exhibió cambios adicionales en el gene S1. Se identificó una deleción de un solo aminoácido en el dominio terminal N del gene S1 (NTD S1) en algunos pollos individuales. La ubicación de esta eliminación en la estructura predicha del gene S1 sugirió la posibilidad de que se tratara de un cambio compensatorio por la capacidad reducida de AL4 para replicarse en la tráquea de pollos sin exposición previa. Los pollos vacunados con una vacuna comercial ArkDPI el día de la eclosión y desafiados a los 21 días de edad mostraron que la vacunación proporcionó una protección incompleta contra el desafío con estos virus. Además, basándose en el número de copias del ARN viral en la tráquea, se detectaron diferencias en la capacidad de la vacuna para proteger contra estos aislados del virus de la bronquitis infecciosa, siendo la vacuna con la protección más deficiente contra AL4. Estos resultados proporcionan evidencia adicional que respalda que las vacunas atenuadas contra el virus de la bronquitis infecciosa, especialmente las vacunas ArkDPI, contribuyen a perpetuar esta enfermedad en los pollos comerciales.

Comparison of postweaning bovine respiratory disease treatment rates between non-vaccinated control beef calves and calves variably primed and boosted using commercially available bovine coronavirus vaccines.

Objective: Bovine respiratory disease (BRD) and overall postweaning treatment rates were compared among 3 groups of calves either differentially primed and boosted with commercially available bovine coronavirus (BCoV) vaccine or not vaccinated against BCoV. Animals: Commercial heifer and steer beef calves born in April and May 2022. Procedure: In June 2022, calves were randomly enrolled into 3 treatment groups. Those in 2 groups [V1 (n = 160) and V2 (n = 160)] were administered a mucosal priming dose of 1 of 2 commercial BCoV vaccines; those in the 3rd group [CTL (n = 151)] were unvaccinated against BCoV. The V1 and V2 groups were boosted by intramuscular injection pre-weaning with the same vaccine used for priming. Weaning occurred 3 wk after the last preweaning processing day. Ranch staff used a BRD case definition provided by their herd veterinarian to identify, treat, and record treatments for 45 d post-weaning. Results: Postweaning BRD treatment rates for V1, V2, and CTL were 7%, 9%, and 14%, respectively. The CTL calves had 2.2× greater odds of receiving treatment for BRD than V1 calves. There were no differences in odds of treatment between CTL and V2 calves or V1 and V2 calves. Conclusion: In a herd with previously diagnosed BCoV BRD cases, prime-boost vaccination of calves is associated with a difference in odds of BRD treatment post-weaning compared to not vaccinating calves against BCoV. Clinical relevance: Prime-boost vaccination with commercial BCoV vaccine may be an important management tool for herds with known BCoV BRD outbreaks.

Comparaison des taux de traitement des maladies respiratoires bovines après le sevrage entre des veaux de boucherie témoins non vaccinés et des veaux vaccinés amorce-rappel de manière variable à l'aide de vaccins contre le coronavirus bovin commercialement disponibles. Objectif: La maladie respiratoire bovine (BRD) et les taux globaux de traitement post-sevrage ont été comparés parmi 3 groupes de veaux soit vaccinés de manière différentielle et avec un rappel avec le vaccin contre le coronavirus bovin (BCoV) disponible commercialement, soit non vaccinés contre le BCoV. Animaux: Génisses et veaux de boucherie commerciaux nés en avril et mai 2022. Procédure: En juin 2022, les veaux ont été randomisés lors du recrutement dans 3 groupes de traitement. Ceux des 2 groupes [V1 (n = 160) et V2 (n = 160)] ont reçu une dose d'amorce par voie muqueuse de l'un des deux vaccins commerciaux BCoV; ceux du 3ème groupe [CTL (n = 151)] n'étaient pas vaccinés contre le BCoV. Les groupes V1 et V2 ont eu un rappel par injection intramusculaire avant le sevrage avec le même vaccin que celui utilisé pour l'amorçage. Le sevrage a eu lieu 3 semaines après le dernier jour de conditionnement pré-sevrage. Le personnel du ranch a utilisé une définition de cas de BRD fournie par le vétérinaire de leur troupeau pour identifier, traiter et enregistrer les traitements pendant 45 jours après le sevrage. Résultats: Les taux de traitement BRD post-sevrage pour V1, V2 et CTL étaient respectivement de 7 %, 9 % et 14 %. Les veaux CTL avaient 2,2 fois plus de chances de recevoir un traitement contre la BRD que les veaux V1. Il n'y avait aucune différence dans les probabilités de traitement entre les veaux CTL et V2 ou entre les veaux V1 et V2. Conclusion: Dans un troupeau avec des cas de BRD causés par le BCoV déjà diagnostiqués, la vaccination amorce-rappel des veaux est associée à une différence de probabilité de traitement par le BRD après le sevrage par rapport à la nonvaccination des veaux contre le BCoV. Pertinence clinique: La vaccination amorce-rappel avec le vaccin commercial BCoV peut être un outil de gestion important pour les troupeaux présentant des foyers connus de BCoV BRD.(Traduit par Dr Serge Messier).

Prevalence of coronaviruses in European bison (Bison bonasus) in Poland.

Coronaviruses have been confirmed to infect a variety of species, but only one case of associated winter dysentery of European bison has been described. The study aimed to analyze the prevalence, and define the impact on the species conservation, the source of coronavirus infection, and the role of the European bison in the transmission of the pathogen in Poland. Molecular and serological screening was performed on 409 European bison from 6 free-ranging and 14 captive herds over the period of 6 years (2017-2023). Presence of coronavirus was confirmed in one nasal swab by pancoronavirus RT-PCR and in 3 nasal swab samples by bovine coronavirus (BCoV) specific real time RT-PCR. The detected virus showed high (> 98%) homology in both RdRp and Spike genes to BCoV strains characterised recently in Polish cattle and strains isolated from wild cervids in Italy. Antibodies specific to BCoV were found in 6.4% of tested samples, all originating from free-ranging animals. Seroprevalence was higher in adult animals over 5 years of age (p = 0.0015) and in females (p = 0.09). Our results suggest that European bison play only a limited role as reservoirs of bovine-like coronaviruses. Although the most probable source of infections in the European bison population in Poland is cattle, other wild ruminants could also be involved. In addition, the zoonotic potential of bovine coronaviruses is quite low.

High-affinity monoclonal antibodies against the porcine epidemic diarrhea virus S1 protein.

The porcine epidemic diarrhea virus (PEDV) infection inflicted substantial economic losses upon the global pig-breeding industry. This pathogen can infect all pigs and poses a particularly high fatality risk for suckling piglets. The S1 subunit of spike protein is a crucial target protein for inducing the particularly neutralizing antibodies that can intercept the virus-host interaction and neutralize virus infectivity. In the present study, the HEK293F eukaryotic expression system was successfully utilized to express and produce recombinant S1 protein. Through quantitative analysis, five monoclonal antibodies (mAbs) specifically targeting the recombinant S1 protein of PEDV were developed and subsequently evaluated using enzyme-linked immunosorbent assay (ELISA), indirect immunofluorescence assay (IFA), and flow cytometry assay (FCA). The results indicate that all five mAbs belong to the IgG1 isotype, and their half-maximal effective concentration (EC50) values measured at 84.77, 7.42, 0.89, 14.64, and 7.86 pM. All these five mAbs can be utilized in ELISA, FCA, and IFA for the detection of PEDV infection. MAb 5-F9 exhibits the highest sensitivity to detect as low as 0.3125 ng/mL of recombinant PEDV-S1 protein in ELISA, while only 0.096 ng/mL of mAb 5-F9 is required to detect PEDV in FCA. The results from antigen epitope analysis indicated that mAb 8-G2 is the sole antibody capable of recognizing linear epitopes. In conclusion, this study has yielded a highly immunogenic S1 protein and five high-affinity mAbs specifically targeting the S1 protein. These findings have significant implications for early detection of PEDV infection and provide a solid foundation for further investigation into studying virus-host interactions.

Evaluation of bovine coronavirus in Korean native calves challenged through different inoculation routes.

Bovine coronavirus (BCoV) is a pneumoenteric virus that can infect the digestive and respiratory tracts of cattle, resulting in economic losses. Despite its significance, information regarding BCoV pathogenesis is limited. Hence, we investigated clinical signs, patterns of viral shedding, changes in antibody abundance, and cytokine/chemokine production in calves inoculated with BCoV via intranasal and oral. Six clinically healthy Korean native calves (< 30 days old), initially negative for BCoV, were divided into intranasal and oral groups and monitored for 15 days post-infection (dpi). BCoV-infected calves exhibited clinical signs such as nasal discharge and diarrhea, starting at 3 dpi and recovering by 12 dpi, with nasal discharge being the most common symptoms. Viral RNA was detected in nasal and fecal samples from all infected calves. Nasal shedding occurred before fecal shedding regardless of the inoculation route; however, fecal shedding persisted longer. Although the number of partitions was very few, viral RNA was identified in the blood of two calves in the oral group at 7 dpi and 9 dpi using digital RT-PCR analysis. The effectiveness of maternal antibodies in preventing viral replication and shedding appeared limited. Our results showed interleukin (IL)-8 as the most common and highly induced chemokine. During BCoV infection, the levels of IL-8, monocyte chemoattractant protein-1, and macrophage inflammatory protein-1ß were significantly affected, suggesting that these emerge as potential and reliable biomarkers for predicting BCoV infection. This study underscores the importance of BCoV as a major pathogen causing diarrhea and respiratory disease.

Heterologous maternal antibodies derived from infectious bronchitis vaccines prevent the development of lesions associated with false layer syndrome.

Infectious bronchitis virus (IBV) strains of the Delmarva (DMV)/1639 genotype have been causing false layer syndrome (FLS) in the Eastern Canadian layer operations since the end of 2015. FLS is characterized by the development of cystic oviducts in layer pullets infected at an early age. Currently, there are no homologous vaccines for the control of this IBV genotype. Our previous research showed that a heterologous vaccination regimen incorporating Massachusetts (Mass) and Connecticut (Conn) IBV types protects layers against DMV/1639 genotype IBV. The aim of this study was to investigate the role of maternal antibodies conferred by breeders received the same vaccination regimen in the protection against the development of DMV/1639-induced FLS in pullets. Maternal antibody-positive (MA+) and maternal antibody-negative (MA-) female progeny chicks were challenged at 1 day of age and kept under observation for 16 weeks. Oviductal cystic formations were observed in 3 of 14 birds (21.4 %) in the MA- pullets, while the lesions were notably absent in the MA+ pullets. Milder histopathological lesions were observed in the examined tissues of the MA+ pullets. However, the maternal derived immunity failed to demonstrate protection against the damage to the tracheal ciliary activity, viral shedding, and viral tissue distribution. Overall, this study underscores the limitations of maternal derived immunity in preventing certain aspects of viral pathogenesis, emphasizing the need for comprehensive strategies to address different aspects of IBV infection.

Prevalence, Genotype Diversity, and Distinct Pathogenicity of 205 Gammacoronavirus Infectious Bronchitis Virus Isolates in China during 2019-2023.

Gammacoronavirus infectious bronchitis virus (IBV) causes a highly contagious disease in chickens and seriously endangers the poultry industry. The emergence and co-circulation of diverse IBV serotypes and genotypes with distinct pathogenicity worldwide pose a serious challenge to the development of effective intervention measures. In this study, we report the epidemic trends of IBV in China from 2019 to 2023 and a comparative analysis on the antigenic characteristics and pathogenicity of isolates among major prevalent lineages. Phylogenetic and recombination analyses based on the nucleotide sequences of the spike (S) 1 gene clustered a total of 205 isolates into twelve distinct lineages, with GI-19 as a predominant lineage (61.77 ± 4.56%) exhibiting an overall increasing trend over the past five years, and demonstrated that a majority of the variants were derived from gene recombination events. Further characterization of the growth and pathogenic properties of six representative isolates from different lineages classified four out of the six isolates as nephropathogenic types with mortality rates in one-day-old SPF chickens varying from 20-60%, one as a respiratory type with weak virulence, and one as a naturally occurring avirulent strain. Taken together, our findings illuminate the epidemic trends, prevalence, recombination, and pathogenicity of current IBV strains in China, providing key information for further strengthening the surveillance and pathogenicity studies of IBV.

Transcriptome Analysis in Air-Liquid Interface Porcine Respiratory Epithelial Cell Cultures Reveals That the Betacoronavirus Porcine Encephalomyelitis Hemagglutinating Virus Induces a Robust Interferon Response to Infection.

Porcine hemagglutinating encephalomyelitis virus (PHEV) replicates in the upper respiratory tract and tonsils of pigs. Using an air-liquid interface porcine respiratory epithelial cells (ALI-PRECs) culture system, we demonstrated that PHEV disrupts respiratory epithelia homeostasis by impairing ciliary function and inducing antiviral, pro-inflammatory cytokine, and chemokine responses. This study explores the mechanisms driving early innate immune responses during PHEV infection through host transcriptome analysis. Total RNA was collected from ALI-PRECs at 24, 36, and 48 h post inoculation (hpi). RNA-seq analysis was performed using an Illumina Hiseq 600 to generate 100 bp paired-end reads. Differential gene expression was analyzed using DeSeq2. PHEV replicated actively in ALI-PRECs, causing cytopathic changes and progressive mucociliary disruption. Transcriptome analysis revealed downregulation of cilia-associated genes such as CILK1, DNAH11, LRRC-23, -49, and -51, and acidic sialomucin CD164L2. PHEV also activated antiviral signaling pathways, significantly increasing the expression of interferon-stimulated genes (RSAD2, MX1, IFIT, and ISG15) and chemokine genes (CCL5 and CXCL10), highlighting inflammatory regulation. This study contributes to elucidating the molecular mechanisms of the innate immune response to PHEV infection of the airway epithelium, emphasizing the critical roles of the mucociliary, interferon, and chemokine responses.

Isolation and Characterization of Contemporary Bovine Coronavirus Strains.

Bovine coronavirus (BCoV) poses a threat to cattle health worldwide, contributing to both respiratory and enteric diseases. However, few contemporary strains have been isolated. In this study, 71 samples (10 nasal and 61 fecal) were collected from one farm in Ohio in 2021 and three farms in Georgia in 2023. They were screened by BCoV-specific real-time reverse transcription-PCR, and 15 BCoV-positive samples were identified. Among them, five BCoV strains from fecal samples were isolated using human rectal tumor-18 (HRT-18) cells. The genomic sequences of five strains were obtained. The phylogenetic analysis illustrated that these new strains clustered with US BCoVs that have been detected since the 1990s. Sequence analyses of the spike proteins of four pairs of BCoVs, with each pair originally collected from the respiratory and enteric sites of one animal, revealed the potential amino acid residue patterns, such as D1180 for all four enteric BCoVs and G1180 for three of four respiratory BCoVs. This project provides new BCoV isolates and sequences and underscores the genetic diversity of BcoVs, the unknown mechanisms of disease types, and the necessity of sustained surveillance and research for BCoVs.

Immunoinformatics approach for designing a multiepitope subunit vaccine against porcine epidemic diarrhea virus genotype IIA spike protein.

Background: Porcine epidemic diarrhea (PED), caused by the porcine epidemic diarrhea virus (PEDV), is associated with high mortality and morbidity rates, especially in neonatal pigs. This has resulted in significant economic losses for the pig industry. PEDV genotype II-based vaccines were found to confer better immunity against both heterologous and homologous challenges; specifically, spike (S) proteins, which are known to play a significant role during infection, are ideal for vaccine development. Aim: This study aims to design a multi-epitope subunit vaccine targeting the S protein of the PEDV GIIa strain using an immunoinformatics approach. Methods: Various bioinformatics tools were used to predict HTL, CTL, and B-cell epitopes. The epitopes were connected using appropriate linkers and conjugated with the CTB adjuvant and M-ligand. The final multiepitope vaccine construct (fMEVc) was then docked to toll-like receptor 4 (TLR4). The stability of the fMEVc-TLR4 complex was then simulated using GROMACS. C-immsim was then used to predict the in vitro immune response of the fMEVc. Results: Six epitopes were predicted to induce antibody production, ten epitopes were predicted to induce CTL responses, and four epitopes were predicted to induce HTL responses. The assembled epitopes conjugated with the CTB adjuvant and M-ligand, fMEVc, is antigenic, non-allergenic, stable, and soluble. The construct showed a favorable binding affinity for TLR4, and the protein complex was shown to be stable through molecular dynamics simulations. A robust immune response was induced after immunization, as demonstrated through immune stimulation. Conclusion: In conclusion, the multi-epitope subunit vaccine construct for PEDV designed in this study exhibits promising antigenicity, stability, and immunogenicity, eliciting robust immune responses and suggesting its potential as a candidate for further vaccine development.

Calmodulin-like 5 promotes PEDV replication by regulating late-endosome synthesis and innate immune response.

The infection caused by porcine epidemic diarrhea virus (PEDV) is associated with high mortality in piglets worldwide. Host factors involved in the efficient replication of PEDV, however, remain largely unknown. Our recent proteomic study in the virus-host interaction network revealed a significant increase in the accumulation of CALML5 (EF-hand protein calmodulin-like 5) following PEDV infection. A further study unveiled a biphasic increase of CALML5 in 2 and 12 â€‹h after viral infection. Similar trends were observed in the intestines of piglets in the early and late stages of the PEDV challenge. Moreover, CALML5 depletion reduced PEDV mRNA and protein levels, leading to a one-order-of-magnitude decrease in virus titer. At the early stage of PEDV infection, CALML5 affected the endosomal trafficking pathway by regulating the expression of endosomal sorting complex related cellular proteins. CALML5 depletion also suppressed IFN-ß and IL-6 production in the PEDV-infected cells, thereby indicating its involvement in negatively regulating the innate immune response. Our study reveals the biological function of CALML5 in the virology field and offers new insights into the PEDV-host cell interaction.

Infectious bronchitis virus: an overview of the "chicken coronavirus".

Infectious bronchitis virus (IBV) is a highly contagious avian Gammacoronavirus that affects mainly chickens (Gallus gallus) but can circulate in other avian species. IBV constitutes a significant threat to the poultry industry, causing reduced egg yield, growth and mortality levels that can vary in impact. The virus can be transmitted horizontally by inhalation or direct/indirect contact with infected birds or contaminated fomites, vehicles, farm personnel and litter (Figure 1). The error-prone viral polymerase and recombination mechanisms mean diverse viral population results, with multiple genotypes, serotypes, pathotypes and protectotypes. This significantly complicates control and mitigation strategies based on vigilance in biosecurity and the deployment of vaccination.

Epidemiological investigation of enteric canine coronaviruses in domestic dogs: A systematic review and meta-analysis.

Canine enteric coronavirus (CeCoV) is a globally distributed enteric pathogen that causes significant harm to canines. The objective of this systematic review was to examine the global dissemination of CeCoV and assess the potential for infected canines to be exposed to various CeCoV genotypes and subtypes. With an aggregated prevalence of 18.8%, the study predicted regional variations, indicating that CeCoV is an exceptionally prevalent disease. The increased likelihood that infected canines will be asymptomatic is a significant cause for concern, as undetected cases of CeCoV infection could persist and spread the disease. This underscores the significance of ongoing surveillance of CeCoV in order to avert its transmission. Nevertheless, further investigation is necessary in order to ascertain the moderators that significantly impact the prevalence and distribution of distinct subtypes and genotypes of CeCoV. Hence, it is imperative to undertake randomized clinical trials in order to acquire a more accurate understanding of the variables that influence the prevalence of CeCoV. By conducting ongoing surveillance, regional variations in the prevalence of CeCoV in canines can be accounted for, thereby enhancing our comprehension of the illness and ultimately impeding its transmission.

Characterisation of the tracheal transcriptional response of chickens to chronic infection with Mycoplasma synoviae.

Mycoplasma synoviae causes infectious synovitis and respiratory tract infections in chickens and is responsible for significant economic losses in the poultry industry. Effective attachment and colonisation of the trachea is critical for the persistence of the organism and progression of the disease it causes. The respiratory tract infection is usually sub-clinical, but concurrent infection with infectious bronchitis virus (IBV) is known to enhance the pathogenicity of M. synoviae. This study aimed to explore differentially expressed genes in the tracheal mucosa, and their functional categories, during chronic infection with M. synoviae, using a M. synoviae-IBV infection model. The transcriptional profiles of the trachea were assessed 2 weeks after infection using RNA sequencing. In chickens infected with M. synoviae or IBV, only 1 or 8 genes were differentially expressed compared to uninfected chickens, respectively. In contrast, the M. synoviae-IBV infected chickens had 621 upregulated and 206 downregulated genes compared to uninfected chickens. Upregulated genes and their functional categories were suggestive of uncontrolled lymphoid cell proliferation and an ongoing pro-inflammatory response. Genes associated with anti-inflammatory effects, pathogen removal, apoptosis, regulation of the immune response, airway homoeostasis, cell adhesion and tissue regeneration were downregulated. Overall, transcriptional changes in the trachea, 2 weeks after infection with M. synoviae and IBV, indicate immune dysregulation, robust inflammation and a lack of cytotoxic damage during chronic infection. This model provides insights into the pathogenesis of chronic infection with M. synoviae.