Bovine Coronavirus Infection Research Articles

Brain invasion of bovine coronavirus: molecular analysis of bovine coronavirus infection in calves with severe pneumonia and neurological signs.

Although the role of bovine coronavirus (BCoV) in calf diarrhea and respiratory disorders is well documented, its contribution to neurological diseases is unclear. This study conducted virological investigations of calves showing diarrhea and respiratory and neurological signs. An outbreak of diarrhea, respiratory, and neurological disorders occurred among the 12 calves in July 2022 in Istanbul, Türkiye. Two of these calves exhibited neurological signs and died a few days after the appearance of symptoms. One of these calves was necropsied and analyzed using molecular and histopathological tests. BCoV RNA was detected in the brain, lung, spleen, liver, and intestine of the calf that had neurological signs by real-time reverse transcription polymerase chain reaction. Immunostaining was also observed in the intestine and brain. A 622 bp S1 gene product was noted on gel electrophoresis only in the brain. Phylogenetic analysis indicated that the BCoV detected in this study had a high proximity to the BCoV strain GIb with 99.19% nucleotide sequence homology to the strains detected in Poland, Israel, Türkiye, and France. No distinct genetic lineages were observed when the brain isolate was compared with the respiratory and enteric strains reported to GenBank. In addition, the highest identity (98,72%) was obtained with the HECV 4408 and L07748 strains of human coronaviruses. The strain detected in a calf brain belongs to the GIb-European lineage and shares high sequence homology with BCoV strains detected in Europe and Israel. In addition, the similarity between the human coronaviruses (4408 and L07748) raises questions about the zoonotic potential of the strains detected in this study.

Humoral Immune Response in Immunized Sheep with Bovine Coronavirus Glycoproteins Delivered via an Adenoviral Vector.

Bovine coronavirus (BCoV) is distributed globally and mainly causes different clinical manifestations: enteric diarrhea in calves, winter dysentery in adults, and respiratory symptoms in cattle of all ages. Low mortality and high morbidity are the hallmarks of BCoV infection, usually associated with substantial economic losses for the livestock industry. Vaccination, combined with the implementation of biosecurity measures, is the key strategy for the prevention of infections. This pilot study evaluates the immunogenicity of a recombinant vaccine containing two BCoV antigens (S and M) in sheep, compared to vaccines containing only the M or S protein. Three groups of sheep were inoculated intramuscularly at day 0 and day 21 with recombinant adenoviruses expressing BCoV S protein (AdV-BCoV-S), BCoV M protein (AdV-BCoV-M), or both proteins (AdV-BCoV-S + M). Serum antibodies were evaluated using immunofluorescence (IF) and serum neutralization (SN) tests. Moderate seroconversion was observed by day 21, but serum antibodies detected via SN increased from 1:27.5 (day 21) to 1:90 (day 28) in sheep inoculated with the recombinant AdV expressing both the S- and M-BCoV proteins. Based on the SN results, a repeated-measures ANOVA test indicated a more significant difference in immune response between the three groups (F = 20.47; p < 0.001). The experimental investigation produced satisfactory results, highlighting that the S + M recombinant vaccine was immunogenic, stimulating a valid immune response. Despite some inherent limitations, including a small sample size and the absence of challenge tests, the study demonstrated the efficacy of the immune response induced via the recombinant vaccine containing both S and M proteins compared to that induced via the individual proteins S or M.

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-1β 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.

Decoding bovine coronavirus immune targets: an epitope informatics approach

Bovine coronavirus (BCoV) poses a significant threat to the global cattle industry, causing both respiratory and gastrointestinal infections in cattle populations. This necessitates the development of efficacious vaccines. While several inactivated and live BCoV vaccines exist, they are predominantly limited to calves. The immunization of adult cattle is imperative for BCoV infection control, as it curtails viral transmission to calves and ameliorates the impact of enteric and respiratory ailments across all age groups within the herd. This study presents an in silico methodology for devising a multiepitope vaccine targeting BCoV. The spike glycoprotein (S) and nucleocapsid (N) proteins, which are integral elements of the BCoV structure, play pivotal roles in the viral infection cycle and immune response. We constructed a remarkably effective multiepitope vaccine candidate specifically designed to combat the BCoV population. Using immunoinformatics technology, B-cell and T-cell epitopes were predicted and linked together using linkers and adjuvants to efficiently trigger both cellular and humoral immune responses in cattle. The in silico construct was characterized, and assessment of its physicochemical properties revealed the formation of a stable vaccine construct. After 3D modeling of the vaccine construct, molecular docking revealed a stable interaction with the bovine receptor bTLR4. Moreover, the viability of the vaccine’s high expression and simple purification was demonstrated by codon optimization and in silico cloning expression into the pET28a (+) vector. By applying immunoinformatics approaches, researchers aim to better understand the immune response to bovine coronavirus, discover potential targets for intervention, and facilitate the development of diagnostic tools and vaccines to mitigate the impact of this virus on cattle health and the livestock industry. We anticipate that the design will be useful as a preventive treatment for BCoV sickness in cattle, opening the door for further laboratory studies.

Study on the susceptibility of bovine coronavirus to BALB/c mice

There are no other bovine coronavirus (BCoV) infection models except calves, which makes efficacy evaluation of vaccines and pathogenic mechanism research of BCoV inconvenient owing to their high value and inconvenient operation. This study aimed to establish a mouse model of BCoV infection. BCoV was used to infect 4-week-old male BALB/c mice and the optimal infection conditions were screened, including the following infection routes: gavage, intraperitoneal injection, and tail vein injection at doses of 1 × 108 TCID50, 2 × 108 TCID50 and 4 × 108 TCID50. Using the optimal infection conditions, BALB/c mice were infected with BCoV, and their body weight, blood routine, inflammatory factors, autopsy, virus distribution, and viral load were measured at 1, 3, 5, and 7 days after infection. The results showed that the optimal conditions for infecting BALB/c mice with BCoV HLJ-325 strain were continuous oral gavage for 3 days with a dose of 4 × 108 TCID50. On the 7th day after infection, there was significant extensive consolidation of the lungs and thinning of the colon wall. Significant inflammation was observed in various organs, especially in the colon and alveoli, where a large number of inflammatory cells infiltrate. Both BCoV Ag and nucleic acid are positive in visceral organs. The viral load in the colon and lungs was significantly higher than that in the other organs (p < 0.001). BCoV-infected mice showed a decreasing trend in body weight starting from day 5, and there was a significant difference compared to the control group on days 6 and 7 (p < 0.001). The total number of white blood cells and lymphocytes began to decrease and was significantly lower than that in the control group 24 h after infection (p < 0.001), and gradually returned to the control level. The cytokine TNF-α, IL-1β, and IL-6 showed an increasing trend, significantly higher than the control group on day 5 and 7 (p < 0.001).These results indicate that the BCoV HLJ-325 strain can infect BALB/c mice and cause inflammatory reactions and tissue lesions. The most significant effect was observed on the seventh day after infection with a dose of 4 × 108 TCID50 and three consecutive gavages. This study established, for the first time, a BALB/c mouse model of BCoV infection, providing a technical means for evaluating the immune efficacy of BCoV vaccines and studying their pathogenic mechanisms.

The Potential Roles of Host Cell miRNAs in Fine-Tuning Bovine Coronavirus (BCoV) Molecular Pathogenesis, Tissue Tropism, and Immune Regulation.

Bovine coronavirus (BCoV) infection causes significant economic loss to the dairy and beef industries worldwide. BCoV exhibits dual tropism, infecting the respiratory and enteric tracts of cattle. The enteric BCoV isolates could also induce respiratory manifestations under certain circumstances. However, the mechanism of this dual tropism of BCoV infection has not yet been studied well. MicroRNAs (miRNAs) are small non-coding RNAs that regulate gene expression and play a dual role in virus infection, mediating virus or modulating host immune regulatory genes through complex virus-host cell interactions. However, their role in BCoV infection remains unclear. This study aims to identify bovine miRNAs crucial for regulating virus-host interaction, influencing tissue tropism, and explore their potential as biomarkers and therapeutic agents against BCoV. We downloaded 18 full-length BCoV genomes (10 enteric and eight respiratory) from GenBank. We applied several bioinformatic tools to study the host miRNAs targeting various regions in the viral genome. We used the criteria of differential targeting between the enteric/respiratory isolates to identify some critical miRNAs as biological markers for BCoV infection. Using various online bioinformatic tools, we also searched for host miRNA target genes involved in BCoV infection, immune evasion, and regulation. Our results show that four bovine miRNAs (miR-2375, miR-193a-3p, miR-12059, and miR-494) potentially target the BCoV spike protein at multiple sites. These miRNAs also regulate the host immune suppressor pathways, which negatively impacts BCoV replication. Furthermore, we found that bta-(miR-2338, miR-6535, miR-2392, and miR-12054) also target the BCoV genome at certain regions but are involved in regulating host immune signal transduction pathways, i.e., type I interferon (IFN) and retinoic acid-inducible gene I (RIG-I) pathways. Moreover, both miR-2338 and miR-2392 also target host transcriptional factors RORA, YY1, and HLF, which are potential diagnostic markers for BCoV infection. Therefore, miR-2338, miR-6535, miR-2392, and miR-12054 have the potential to fine-tune BCoV tropism and immune evasion and enhance viral pathogenesis. Our results indicate that host miRNAs play essential roles in the BCoV tissue tropism, pathogenesis, and immune regulation. Four bovine miRNAs (miR-2375, bta-miR-193a-3p, bta-miR-12059, and bta-miR-494) target BCoV-S glycoprotein and are potentially involved in several immune suppression pathways during the viral infection. These miRNA candidates could serve as good genetic markers for BCoV infection. However, further studies are urgently needed to validate these identified miRNAs and their target genes in the context of BCoV infection and dual tropism and as genetic markers.

Haemato-biochemical Changes and Various Risk Factors Associated with Bovine Coronavirus Infection in Cattle Calves

Haemato-biochemical Changes and Various Risk Factors Associated with Bovine Coronavirus Infection in Cattle Calves

Detection of bovine coronavirus (BCoV) infection in cattle with clinical respiratory signs by PCR and investigation of the serological status of these animals

Aim: It was aimed to reveal the antigenic and serological prevalence of bovine coronavirus (BCoV) in cattle with clinically respiratory infection and to evaluate the relationship between virus shedding and serological status of animals in this study. Materials and Methods: In the study, nasal swab samples were collected from a total of 92 cattle of different gender between 0 and 2 years of age with clinical respiratory symptoms. These samples were subjected to RT-PCR using specific primer pairs. Furthermore, serum samples were also collected from the same animals and evaluated for BCoV-specific antibodies through an indirect ELISA method. Results: Ten (10.87%) swap samples were defined as positive for BCoV using RT-PCR. The seropositivity rate was 93.48% (86/92) in cattle by ELISA and 90% (9/10) in virus-shedding animals with nasal swab samples. Conclusion: The fact that 93.47% seropositivity was detected in unvaccinated animals against coronavirus in this study indicates that the animals were exposed to BCoV through colostrum or at a certain period of their lives. However the fact that 9 out of 10 animals with BCoV detected in swab samples also have BCoV-specific antibodies casts doubt on the efficacy of these antibodies in protecting against respiratory BCoV infections. We concluded that more detailed studies are needed to reveal the role of antibodies in respiratory coronavirus infections.

Alterations of microbiota and metabolites in the feces of calves with diarrhea associated with rotavirus and coronavirus infections.

The changes in the composition of intestinal microbiota and metabolites have been linked to digestive disorders in calves, especially neonatal calf diarrhea. Bovine rotavirus (BRV) and bovine coronavirus (BCoV) are known to be the primary culprits behind neonatal calf diarrhea. In this study, we analyzed changes in the fecal microbiota and metabolites of calves with neonatal diarrhea associated with BRV and BCoV infection using high-throughput 16S rRNA sequencing and metabolomics technology. The microbial diversity in the feces of calves infected with BRV and BCoV with diarrhea decreased significantly, and the composition changed significantly. The significant increase of Fusobacterium and the reductions of some bacteria genera, including Faecalibacterium, Bifidobacterium, Ruminococcus, Subdoligranulum, Parabacteroides, Collinsella, and Olsenella, etc., were closely related to diarrhea associated with BRV and BCoV infection. Metabolites in the feces of BRV and BCoV-infected calves with diarrhea were significantly changed. Phosphatidylcholine [PC; 16:1(9 Z)/16:1(9 Z)], lysophosphatidylethanolamine (LysoPE; 0:0/22:0), lysophosphatidylcholine (LysoPC; P-16:0) and LysoPE (0:0/18:0) were significantly higher in the feces of BRV-infected calves with diarrhea. In contrast, some others, such as desthiobiotin, were significantly lower. BRV infection affects glycerophospholipid metabolism and biotin metabolism in calves. Two differential metabolites were significantly increased, and 67 differential metabolites were significantly reduced in the feces of BCoV-infected calves with diarrhea. Seven significantly reduced metabolites, including deoxythymidylic acid (DTMP), dihydrobiopterin, dihydroneopterin triphosphate, cortexolone, cortisol, pantetheine, and pregnenolone sulfate, were enriched in the folate biosynthesis, pantothenate and CoA biosynthesis, pyrimidine metabolism, and steroid hormone biosynthesis pathway. The decrease in these metabolites was closely associated with increased harmful bacteria and reduced commensal bacteria. The content of short-chain fatty acids (SCFAs) such as acetic acid and propionic acid in the feces of BRV and BCoV-infected calves with diarrhea was lower than that of healthy calves, which was associated with the depletion of SCFAs-producing bacteria such as Parabacteroides, Fournierella, and Collinsella. The present study showed that BRV and BCoV infections changed the composition of the calf fecal microbiota and were associated with changes in fecal metabolites. This study lays the foundation for further revealing the roles of intestinal microbiota in neonatal calf diarrhea associated with BRV and BCoV infection.

Bovine Enteroids as an In Vitro Model for Infection with Bovine Coronavirus.

Bovine coronavirus (BCoV) is one of the major viral pathogens of cattle, responsible for economic losses and causing a substantial impact on animal welfare. Several in vitro 2D models have been used to investigate BCoV infection and its pathogenesis. However, 3D enteroids are likely to be a better model with which to investigate host-pathogen interactions. This study established bovine enteroids as an in vitro replication system for BCoV, and we compared the expression of selected genes during the BCoV infection of the enteroids with the expression previously described in HCT-8 cells. The enteroids were successfully established from bovine ileum and permissive to BCoV, as shown by a seven-fold increase in viral RNA after 72 h. Immunostaining of differentiation markers showed a mixed population of differentiated cells. Gene expression ratios at 72 h showed that pro-inflammatory responses such as IL-8 and IL-1A remained unchanged in response to BCoV infection. Expression of other immune genes, including CXCL-3, MMP13, and TNF-α, was significantly downregulated. This study shows that the bovine enteroids had a differentiated cell population and were permissive to BCoV. Further studies are necessary for a comparative analysis to determine whether enteroids are suitable in vitro models to study host responses during BCoV infection.

Seroprevalence and Risk Factors for Bovine Coronavirus Infection among Dairy Cattle and Water Buffalo in Campania Region, Southern Italy

Cattle and water buffalo are the main livestock species that are raised in the Campania region, southern Italy, and they contribute significantly to the regional rural economy. Currently there are limited data on the prevalence of relevant impact infections, such as bovine coronavirus (BCov), an RNA virus that causes acute enteric and respiratory disease. Although these diseases are described primarily in cattle, there have been reports of spillovers to other ruminants, including water buffalo. Here, we determined the seroprevalence of BCoV in cattle and water buffalo in the Campania region of southern Italy. An overall seroprevalence of 30.8% was determined after testing 720 sampled animals with a commercial enzyme-linked immunosorbent assay. A risk factor analysis revealed that the seropositivity rates in cattle (49.2%) were higher than in water buffalo (5.3%). In addition, higher seroprevalence rates were observed in older and purchased animals. In cattle, housing type and location were not associated with higher seroprevalence. The presence of BCoV antibodies in water buffalo was associated with the practice of co-inhabiting with cattle, demonstrating that this practice is incorrect and promotes the transmission of pathogens between different species. Our study found a considerable seroprevalence, which is consistent with previous research from other countries. Our results provide information on the widespread distribution of this pathogen as well as the risk factors that are involved in its transmission. This information could be useful in the control and surveillance of this infection.

Development of a multienzyme isothermal rapid amplification and lateral flow dipstick combination assay for bovine coronavirus detection.

Bovine coronavirus (BCoV) is a major cause of infectious disease in cattle, causing huge economic losses to the beef and dairy industries worldwide. BCoV can infect humans and multiple other species of animals. A rapid, reliable, and simple test is needed to detect BCoV infection in suspected farms. In this study, we developed a novel multienzyme isothermal rapid amplification (MIRA) and lateral flow dipstick (LFD) combination assay, targeting a highly conserved region of the viral nucleocapsid (N) gene for BCoV detection. The MIRA-LFD assay was highly specific and sensitive, comparable to a published reverse transcription quantitative PCR (RT-qPCR) assay for BCoV detection. Compared with the published RT-qPCR assay, the κ value of the MIRA-LFD assay in the detection of 192 cattle clinical samples was 0.982. The MIRA-LFD assay did not require sophisticated instruments and the results could be observed with eyes. Our results showed that the MIRA-LFD assay was a useful diagnostic tool for rapid on-site detection of BCoV.

Evolutionary adaptation of bovine coronavirus (BCoV): Screening of natural recombinations across the complete genomes.

Bovine coronavirus (BCoV) is a member of pathogenic Betacoronaviruses that has been circulating for several decades in multiple host species. Given the similarity between BCoV and human coronaviruses, the current study aimed to review the complete genomes of 107 BCoV strains available on the GenBank database, collected between 1983 and 2017 from different countries. The maximum-likelihood based phylogenetic analysis revealed three main BCoV genogroups: GI, GII, and GIII. GI is further divided into nine subgenogroups: GI-a to GI-i. The GI-a to GI-d are restricted to Japan, and GI-e to GI-i to the USA. The evolutionary relationships were also inferred using phylogenetic network analysis, revealing two major distinct networks dominated by viruses identified in the USA and Japan, respectively. The USA strains-dominated Network Cluster includes two sub-branches: France/Germany and Japan/China in addition to the United States, while Japan strains-dominated Network Cluster is limited to Japan. Twelve recombination events were determined, including 11 intragenogroup (GI) and one intergenogroup (GII vs. GI-g). The breakpoints of the recombination events were mainly located in ORF1ab and the spike glycoprotein ORF. Interestingly, 10 of 12 recombination events occurred between Japan strains, one between the USA strains, and one from intercontinental recombination (Japan vs. USA). These findings suggest that geographical characteristics, and population density with closer contact, might significantly impact the BCoV infection and co-infection and boost the emergence of more complex virus lineages.

VIRAL PNEUMOENTERITIS OF CATTLE IN FARMS OF THE REPUBLIC OF BELARUS

The article describes the etiological structure of pneumoenteritis in cattle (cattle) in livestock farms of the Re-public of Belarus for the period 2020–2021. based on the analysis of the results of our own research and describes the role of associations of viruses involved in their occurrence.&#x0D; Among the etiological causes of pneumoenteritis in cattle, there are such infections as BVDV, IBR, PIV-3, BRSV, bovine rotavirus and bovine coronavirus infections that cause significant economic damage to livestock. Preven-tion of diseases is the most important task of modern animal husbandry. In many cases, preventive measures can pre-vent the development of diseases or make their prognosis and consequences more favorable. Prevention and treatment requires rapid and accurate laboratory diagnosis. In addition to establishing the etiology of the disease, laboratory diagnostics is important in establishing the epizootic situation and in organizing anti-epizootic measures.

Egyptian cobra (Naja haje haje) venom phospholipase A2: a promising antiviral agent with potent virucidal activity against simian rotavirus and bovine coronavirus

Viral infections are linked to a variety of human diseases. Despite the achievements made in drug and vaccine development, several viruses still lack preventive vaccines and efficient antiviral compounds. Thus, developing novel antiviral agents is of great concern, particularly the natural products that are promising candidates for such discoveries. In this study, we have purified an approximately 15 kDa basic phospholipase A2 (PLA2) enzyme from the Egyptian cobra Naja haje haje venom. The purified N. haje PLA2 showed a specific activity of 22 units/mg protein against 6 units/mg protein for the whole crude venom with 3.67-fold purification. The antiviral activity of purified N. haje PLA2 has been investigated in vitro against bovine coronavirus (BCoV) and simian rotavirus (RV SA-11). Our results showed that the CC50 of PLA2 were 33.6 and 29 µg/ml against MDBK and MA104 cell lines, respectively. Antiviral analysis of N. haje PLA2 showed an inhibition of BCoV and RV SA-11 infections with a therapeutic index equal to 33.6 and 16, respectively. Moreover, N. haje PLA2 decreased the BCoV and RV SA-11 titers by 4.25 log10 TCID50 and 2.5 log10 TCID50, respectively. Thus, this research suggests the potential antiviral activity of purified N. haje PLA2 against BCoV and RV SA-11 infections in vitro.

Evaluation of diagnostic methods for the Detection of Bovine Coronavirus and Rotavirus in feces of diarrheic calves

The purpose of the present study was to evaluate the efficacy of Enzyme-Linked Immunosorbent Assay (ELISA), immunochromatographic (ICG), and reverse transcription-polymerase chain reaction (RT-PCR) methods for the detection of rotavirus (RV) and bovine coronavirus (BCV). Feces samples were collected from 90 diarrhoeic calves (male and female) up to one month of age and the immune response against RV and BCV infection was assessed by using ELISA (Ag and Ab), ICG, and RT-PCR. To determine the performance and accuracy of each diagnostic method in comparison to the diagnostic gold standard (RT-PCR) method, different statistical tests including receiver operating characteristic curve (ROC), concordance correlation were used. Results shown the prevalence of RV and BCV and RV+BCV according to RT-PCR were equal to 8.89 (95% CI: 6.64-10.07), 14.44 (95% CI: 11.23-6.90), and 2.22 (95% CI: 0.89-3.72), respectively. The best agreement and the highest sensitivity and specificity were obtained between the RT-PCR and AgELISA (100% and 94.3%), and then AbELISA (100% and 94.3%) was the second-best test and also the ICG test (95% and 94.3%) was less accurate method in comparison to ELISA methods for identifying RV and BCV, but a good correlation and concordance between ICG diagnostic techniques and RT-PCR were observed. To put it in a nutshell, our results suggest that the ICG assay may improve the ability to diagnose calves RV and BCV infections accurately and quickly. Promoting rapid IGG kit with higher accuracy in early diagnosis of the cause of diarrhoea plays an important role in its therapeutic regimes, management protocols, and control procedures, but ELISA is preferred due to more precise results.

Prevalence and genetic characterization of bovine coronavirus identified from diarrheic pre-weaned native Korean calves from 2019 to 2021.

Bovine coronavirus (BCoV) is associated with severe diarrhea in calves, winter dysentery in adult cattle, and respiratory diseases in cattle. However, there is currently limited information regarding its molecular characterization in the Republic of Korea (KOR). Therefore, this study investigated the prevalence of BCoV in diarrheic pre-weaned calves (aged ≤60days) and compared BCoV genome sequences identified globally. A total of 846 fecal samples were collected from calves with diarrhea across 100 beef farms in the KOR. The samples were divided into three groups based on age as follows: 1-10days (n=490), 11-30days (n=277), and 31-60days (n=79). BCoV infection was detected in 50 calves by real-time RT-PCR analysis. The results showed that the prevalence of BCoV was associated with calf age (P=0.028) and was significantly higher in calves aged 31-60days (odds ratio: 2.69, 95% confidence interval: 1.24-5.85; P=0.012) than in those aged 1-10days. Our findings show that BCoV is an important etiological agent of diarrhea in calves aged 31-60days. Fifteen full genome sequences (2019-2021 variants) of the spike, hemagglutinin/esterase, and nucleocapsid were obtained from the 50 BCoV-positive samples. Phylogenetic analysis of each gene revealed that BCoVs circulating worldwide might have no boundary between enteric and respiratory tropisms, demonstrating the presence of three BCoVs groups: the classical, Asia/USA, and European. Initially, Korean BCoVs were originated from the USA, but diverged since the 1980s and rapidly evolved independently, unlike in other Asian countries. In this study, Korean BCoVs are more recent BCoVs and present relatively high nucleotide substitution rates in all genes compared with other BCoVs. Our results showed that the 2019-2021 variants undergo continuous genetic evolution and that there are genetic differences among globally distributed BCoVs.

Advances in Bovine Coronavirus Epidemiology.

Bovine coronavirus (BCoV) is a causative agent of enteric and respiratory disease in cattle. BCoV has also been reported to cause a variety of animal diseases and is closely related to human coronaviruses, which has attracted extensive attention from both cattle farmers and researchers. However, there are few comprehensive epidemiological reviews, and key information regarding the effect of S-gene differences on tissue tendency and potential cross-species transmission remain unclear. In this review, we summarize BCoV epidemiology, including the transmission, infection-associated factors, co-infection, pathogenicity, genetic evolution, and potential cross-species transmission. Furthermore, the potential two-receptor binding motif system for BCoV entry and the association between BCoV and SARS-CoV-2 are also discussed in this review. Our aim is to provide valuable information for the prevention and treatment of BCoV infection throughout the world.

Bovine Coronavirus Infects the Respiratory Tract of Cattle Challenged Intranasally.

Bovine Coronavirus (BCoV) is a member of a family of viruses associated with both enteric and respiratory diseases in a wide range of hosts. BCoV has been well-established as a causative agent of diarrhea in cattle, however, its role as a respiratory pathogen is controversial. In this study, fifteen calves were challenged intranasally with virulent BCoV in order to observe the clinical manifestation of the BCoV infection for up to 8 days after initial challenge, looking specifically for indication of symptoms, pathology, and presence of viral infection in the respiratory tract, as compared to six unchallenged control calves. Throughout the study, clinical signs of disease were recorded and nasal swabs were collected daily. Additionally, bronchoalveolar lavage (BAL) was performed at 4 days Post-challenge, and blood and tissue samples were collected from calves at 4, 6, or 8 days Post-challenge to be tested for the presence of BCoV and disease pathology. The data collected support that this BCoV challenge resulted in respiratory infections as evidenced by the isolation of BCoV in BAL fluids and positive qPCR, immunohistochemistry (IHC), and histopathologic lesions in the upper and lower respiratory tissues. This study can thus be added to a growing body of data supporting that BCoV is a respiratory pathogen and contributor to respiratory disease in cattle.

Targeting of Protein Kinase CK2 Elicits Antiviral Activity on Bovine Coronavirus Infection

Coronaviruses constitute a global threat to the human population; therefore, effective pan-coronavirus antiviral drugs are required to tackle future re-emerging virus outbreaks. Protein kinase CK2 has been suggested as a promising therapeutic target in COVID-19 owing to the in vitro antiviral activity observed after both pharmacologic and genetic inhibition of the enzyme. Here, we explored the putative antiviral effect of the anti-CK2 peptide CIGB-325 on bovine coronavirus (BCoV) infection using different in vitro viral infected cell-based assays. The impact of the peptide on viral mRNA and protein levels was determined by qRT-PCR and Western blot, respectively. Finally, pull-down experiments followed by Western blot and/or mass spectrometry analysis were performed to identify CIGB-325-interacting proteins. We found that CIGB-325 inhibited both the cytopathic effect and the number of plaque-forming units. Accordingly, intracellular viral protein levels were clearly reduced after treatment of BCoV-infected cells, with CIGB-325 determined by immunocytochemistry. Pull-down assay data revealed the physical interaction of CIGB-325 with viral nucleocapsid (N) protein and a group of bona fide CK2 cellular substrates. Our findings evidence in vitro antiviral activity of CIGB-325 against bovine coronavirus as well as some molecular clues that might support such effect. Altogether, data provided here strengthen the rationale of inhibiting CK2 to treat betacoronavirus infections.