Bovine Parainfluenza Virus Type Research Articles

The Prevalence, Coexistence, and Correlations between Seven Pathogens Detected by a PCR Method from South-Western Poland Dairy Cattle Suffering from Bovine Respiratory Disease.

Bovine respiratory disease (BRD) is a very important disease that contributes to economic losses in dairy and beef cattle breeding worldwide. The molecular testing of material from 296 calves showing BRD symptoms from 74 dairy herds located in south-western Poland was performed in 2019–2021. Molecular tests were performed using a commercial kit “VetMAXTM Ruminant Respiratory Screening Kit” (Thermo Fisher Scientific) for the simultaneous detection of genetic material of seven pathogens responsible for BRD. At least one pathogen was detected in 95.95% of herds. The overall prevalence was: Pasteurella multocida 87.84%, Mannheimia haemolytica 44.59%, bovine coronavirus (BcoV) 32.43%, Mycoplasma bovis 29.73%, Histophilus somni 28.38%, bovine parainfluenza virus type 3 (BPIV-3) 13.51%, and bovine respiratory syncytial virus (BRSV) 10.81%. Twenty-nine configurations of pathogen occurrences were found. Bacterial infections were the most frequently recorded as 56.7% of all results. Coinfections mainly consisted of two pathogens. Not a single purely viral coinfection was detected. The most frequent result was a single P. multocida infection accounting for 18.31% of all results. The statistically significant correlation (p = 0.001) with the highest strength of effect (ϕ 0.38) was between M. bovis and H. somni.

Serodiagnosis and Risk Factors Associated with Infectious Agents of Reproductive Diseases in Bovines of Chiquinquirá, District of Boyacá (Colombia)

The productivity of cattle farms is affected by infectious and noninfectious factors that generate economic losses and cause reproductive failure represented by low conception rates, embryonic mortality, abortions, and fetal mummification. The infectious agents that most impact the reproductive health of the bovine species from conception to birth are bovine herpesvirus type 1 (BoHV-1) causing infectious bovine rhinotracheitis (IBR), bovine viral diarrhea virus (BVDV), bovine parainfluenza virus type 3 (PI3), Neospora caninum and Leptospira spp. The objective of this study was to diagnose the presence of BoHV-1, bovine viral diarrhea (BVD), PI3, Neospora caninum, and Leptospira spp. by serology and identify the risk factors associated with infectious agents of reproductive interest in bovines of Boyacá (Colombia). A descriptive cross-sectional study was developed, with simple random sampling, where a sample size of 601 female cattle of Holstein, Jersey, and Normande breeds of different age groups was determined. Blood samples were taken and processed using the indirect ELISA technique (SYNBIOTICS®, SERELISA® BVD p80 Ab Mono Blocking, Ingezim R.12.NC.K, PRIMACHECK VPI-3®) and the MAT test for the diagnosis of bovine leptospirosis. The data were processed with the statistical program Epi Info™. The highest apparent seroprevalence was established for infectious bovine rhinotracheitis (61.1%), followed by BVD (37.6%), PI3 (40.9%), neosporosis (51.1%), and leptospirosis (14.8%). Variables such as age >4 years and Holstein breed for IBR and >4 years for BVD were established risk factors. Considering our results, we suggest implementing prevention and control plans that include vaccination as a prophylactic measure and biosecurity tools that reduce the probability of contagion and transmission of pathogens.

Quantitative proteomic analysis shows involvement of the p38 MAPK pathway in bovine parainfluenza virus type 3 replication

BackgroundBovine parainfluenza virus type 3 (BPIV3) infection often causes respiratory tissue damage and immunosuppression and further results in bovine respiratory disease complex (BRDC), one of the major diseases in dairy cattle, caused huge economical losses every year. However, the pathogenetic and immunoregulatory mechanisms involved in the process of BPIV3 infection remain unknown. However, the pathogenetic and immunoregulatory mechanisms involved in the process of BPIV3 infection remain unknown. Proteomics is a powerful tool for high-throughput identification of proteins, which has been widely used to understand how viruses interact with host cells.MethodsIn the present study, we report a proteomic analysis to investigate the whole cellular protein alterations of MDBK cells infected with BPIV3. To investigate the infection process of BPIV3 and the immune response mechanism of MDBK cells, isobaric tags for relative and absolute quantitation analysis (iTRAQ) and Q-Exactive mass spectrometry-based proteomics were performed. The differentially expressed proteins (DEPs) involved in the BPIV3 invasion process in MDBK cells were identified, annotated, and quantitated.ResultsA total of 116 proteins, which included 74 upregulated proteins and 42 downregulated proteins, were identified as DEPs between the BPIV3-infected and the mock-infected groups. These DEPs included corresponding proteins related to inflammatory response, immune response, and lipid metabolism. These results might provide some insights for understanding the pathogenesis of BPIV3. Fluorescent quantitative PCR and western blotting analysis showed results consistent with those of iTRAQ identification. Interestingly, the upregulated protein MKK3 was associated with the p38 MAPK signaling pathway.ConclusionsThe results of proteomics analysis indicated BPIV3 infection could activate the p38 MAPK pathway to promote virus replication.

Epidermal growth factor receptor (EGFR) promotes uptake of bovine parainfluenza virus type 3 into MDBK cells

Bovine parainfluenza virus 3 (BPIV3) is an important respiratory pathogen of both young and adult cattle. The pathways of cell entry are highly related to viral transmission and pathogenicity. In previous studies, we demonstrated that macropinocytosis and clathrin-dependent endocytosis play critical roles in the entry of BPIV3 into MDBK cells. Macropinocytosis is special endocytic process which need to activate signaling pathways that remodle the actin cytoskeleton. Parainfluenza viruses (PIVs) initiate infection by binding to sialic acid receptors on cell surfaces. Nevertheless, sialic acids are not able to transmit signals across the plasma membrane, indicating the necessity for additional signaling receptors. Here, we have demonstrated that specific inhibitors and siRNAs targeting EGFR inhibit the entry of BPIV3 into MDBK cells. BPIV3 productive infection in MDBK cells led to activation of EGFR. Inactivation of EGFR suppressed BPIV3-induced rearrangement of the F-actin cytoskeleton. In addition, PI3K-Akt and ERK1/2 were activated in an EGFR-dependent manner during BPIV3 infection. Specific inhibitors targeting these canonical downstream effectors of EGFR could significantly reduce viral entry efficacy. Moreover, we also demonstrated that the important regulators of macropinocytosis, Rac1 and Pak1, are downstream mediators of EGFR during BPIV3 internalization. These results indicated that EGFR is a host-entry cofactor used by BPIV3 to enter MDBK cells.

Temporal Dynamics of the Ruminant Type I IFN-Induced Antiviral State against Homologous Parainfluenza Virus 3 Challenge In Vitro.

Viruses have evolved diverse strategies to evade the antiviral response of interferons (IFNs). Exogenous IFNs were applied to eliminate the counteracting effect and possess antiviral properties. Caprine parainfluenza virus 3 (CPIV3) and bovine parainfluenza virus type 3 (BPIV3) are important pathogens associated with respiratory diseases in goat and cattle, respectively. To explore the feasibility of type I IFNs for control of CPIV3 and BPIV3 infection, the activated effects of IFN-stimulated genes (ISGs) and the immunomodulation responses of goat IFN-α were detected by transcriptomic analysis. Then, the antiviral efficacy of goat IFN-α and IFN-τ against CPIV3 and BPIV3 infection in MDBK cells was evaluated using different treatment routes at different infection times. The results showed that CPIV3 infection inhibited the production of type I IFNs, whereas exogenous goat IFN-α induced various ISGs, the IFN-τ encoding gene, and a negligible inflammatory response. Consequently, goat IFN-α prophylaxis but not treatment was found to effectively modulate CPIV3 and BPIV3 infection; the protective effect lasted for 1 week, and the antiviral activity was maintained at a concentration of 0.1 μg/mL. Furthermore, the antiviral activity of goat IFN-τ in response to CPIV3 and BPIV3 infection is comparable to that of goat IFN-α. These results corroborate that goat IFN-α and IFN-τ exhibit prophylactic activities in response to ruminant respiratory viral infection in vitro, and should be further investigated for a potential use in vivo.

Self-assembled BPIV3 nanoparticles can induce comprehensive immune responses and protection against BPIV3 challenge by inducing dendritic cell maturation in mice

Bovine parainfluenza virus type 3 (BPIV3) is one of the most important viral respiratory pathogens of cattle. No specific therapies are available for BPIV3 infection; vaccination is one of the most effective ways to prevent BPIV3 infection. We therefore prepared the self-assembled BPIV3 nanoparticles by genetically fusing the ectodomain of BPIV3 haemagglutinin-neuraminidase (HN) (HNex) to the NH2 terminus of ferritin (HNex-RFNp) using a baculovirus expression system. It was found that HNex-RFNp-induced bone marrow-derived dendritic cell (BMDC) maturation through the upregulated expression of surface molecules (MHC II, CD80, CD86, and CD40), increased the secretion of inflammatory cytokines (IL-6, IL-12, TNF-α, and IFN-γ), and reduced antigen phagocytosis and T cell activation capacity. HNex-RFNp positively regulated IκBα and NF-κB (p65) phosphorylation and facilitated NF-κB (p65) translocation into the nuclei of mature BMDCs. Incubating RFNp-treated BMDCs with TLR4 and NF-κB (p65) inhibitors, suppressed surface molecule expression as well as pro-inflammatory cytokine production and IκBα and NF-κB (p65) activities. The BPIV3 HNex protein induced BMDC maturation to some extent but was significantly weaker than HNex-RFNp. We found that HNex-RFNp induced a higher titre of specific antibodie, haemagglutinin inhibition (HI) antibody, and virus neutralisation (VN) antibody, and a comprehensive cellular immune response. We examined protection against BPIV3 challenge in a mouse model. Pathological changes were not observed in the lungs of HNex-RFNp-vaccinated mice. Levels of BPIV3 RNA and virus titres in the lungs and trachea were significantly lower in the HNex-RFNp, than HNex, inactivated BPIV3, and PBS groups. In summary, HNex-RFNp elicited better immunogenicity than HNex or inactivated BPIV3 and could be developed as an effective vaccine to protect against BPIV3 infection.

A two-year serological study of bovine viral diarrhea virus, bovine alphaherpesvirus 1 and bovine parainfluenza virus type 3 in Qazvin dairy cattle farms, Northwestern of Iran

Infections with bovine viral diarrhea virus (BVDV), bovine alphaherpesvirus 1 (BoHV-1), and bovine parainfluenza virus type 3 (BPIV-3) cause diseases in cattle with serious economic consequences worldwide. The objective of the present study was to determine of herd-level and animal-level BVDV, BoHV-1, and BPIV-3 seroprevalence, and evaluate some of the associated risk factors on farms in Qazvin province, Northwestern Iran. A total of 1036 cattle in 16 herds were randomly selected, and their serum samples were tested to detect antibodies to these viruses in a cross-sectional study over 2 years. The results showed the seroprevalence of BVDV, BoHV-1, and BPIV-3 was 100%, 56.3%, and 100% at herd-level and 55.1%, 5.1%, and 95.2% at animal-level, respectively. Statistical analysis revealed that the farm was a strong risk factor for all the studied viruses, while the year was determined as a risk factor for only BVDV (P<0.001). The seroprevalence of BVDV and BPIV-3 was significantly (P<0.01) affected by season. The proportion of seropositive cows increased with age for BVDV and BoHV-1 (P<0.001). Concurrent infection was the highest in mixed infections with BVDV and BPIV-3 (53.2%), and there was a positive correlation between BVDV and BoHV-1 seropositivity (R2= 0.106, P<0.001). The present study shows that infections of BVDV and BPIV-3 are common in cattle in Northwestern Iran and which implies the need to implement control programs to reduce the risk of the spread of these viruses.

Limitations of bacterial culture, viral PCR, and tulathromycin susceptibility from upper respiratory tract samples in predicting clinical outcome of tulathromycin control or treatment of bovine respiratory disease in high-risk feeder heifers.

A cross-sectional prospective cohort study including 1026 heifers administered tulathromycin due to high risk of clinical signs of bovine respiratory disease (BRD), measured poor association between BRD clinical outcomes and results of bacterial culture and tulathromycin susceptibility from BRD isolates of deep nasopharyngeal swabs (DNS) and adequate association with viral polymerase chain reaction (PCR) results from nasal swabs. Isolation rates from DNS collected on day-0 and at 1st BRD-treatment respectively were: Mannheimia haemolytica (10.9% & 34.1%); Pasteurella multocida (10.4% & 7.4%); Mycoplasma bovis (1.0% & 36.6%); and Histophilus somni (0.7% & 6.3%). Prevalence of BRD viral nucleic acid on nasal swabs collected exclusively at 1st BRD-treatment were: bovine parainfluenza virus type-3 (bPIV-3) 34.1%; bovine viral diarrhea virus (BVDV) 26.3%; bovine herpes virus type-1 (BHV-1) 10.8%; and bovine respiratory syncytial virus (BRSV) 54.1%. Increased relative risk, at 95% confidence intervals, of 1st BRD-treatment failure was associated with positive viral PCR results: BVDV 1.39 (1.17–1.66), bPIV-3 1.26 (1.06–1.51), BHV-1 1.52 (1.25–1.83), and BRSV 1.35 (1.11–1.63) from nasal swabs collected at 1st BRD-treatment and culture of M. haemolytica 1.23 (1.00–1.51) from DNS collected at day-0. However, in this population of high-risk feeder heifers, the predictive values of susceptible and resistant isolates had inadequate association with BRD clinical outcome. These results indicate, that using tulathromycin susceptibility testing of isolates of M. haemolytica or P. multocida from DNS collected on arrival or at 1st BRD-treatment to evaluate tulathromycin clinical efficacy, is unreliable.

Development of a One-Step Multiplex Real-Time PCR Assay for the Detection of Viral Pathogens Associated With the Bovine Respiratory Disease Complex.

Bovine respiratory disease complex (BRDC) occurs widely in cattle farms. The main viral pathogens include bovine viral diarrhea virus (BVDV), Bovine herpesvirus 1 (BoHV-1), bovine parainfluenza virus type 3 (BPIV3), and bovine respiratory syncytial virus (BRSV), and the newly emerged influenza D virus (IDV). In this study, we have developed a one-step multiplex real-time Polymerase Chain Reaction (PCR) capable of simultaneously detecting these five viral pathogens causing BRDC. The established assay could specifically detect targeted viruses without cross-reaction with others. The detection limit was ~10 copies/reaction for single real-time PCR and 100 copies/ reaction for multiplex real-time PCR assay. A total of 213 nasal samples from cattle with signs of respiratory tract disease were then collected for performance evaluation of the established platform, proving that the method has good specificity and sensitivity. The surveillance data suggested that BVDV and BoHV-1 infections are the dominant cause of BRDC in the herd, whereas the detection rate of IDV, BIPV3, and BRSV is relatively lower. In summary, the established assay provides technical support for rapid clinical detection of BRDC associated viral pathogens to guide the formulation of BRDC prevention and control measures.

A lateral flow dipstick combined with reverse transcription recombinase polymerase amplification for rapid and visual detection of the BVDV and BPIV3

Bovine respiratory disease complex (BRDC) is a serious disease affecting feedlot cattle in China and likely other places worldwide. Bovine viral diarrhea virus (BVDV) and bovine parainfluenza virus type 3 (BPIV3) are principally responsible for causing BRDC, and are a major strain to the industrial economy. Eradication of these viruses/disease requires swift viral identification and treatment. Hence, this study established a fast and easy procedure of BVDV and BPIV3 identification that employs reverse transcription recombinase polymerase amplification (RT-RPA) and lateral flow dipstick (LFD), and uses primers and lateral flow (LF) probe targeting the 5′-UTR gene of BVDV and phosphoprotein P gene of BPIV3, respectively. Our assay was able to successfully amplify BVDV and BPIV3 RNA within 25 min at 35 °C using RT-RPA, with products visible on the LFD within 5 min at room temperature (RT). The lowest detection limits were 50 RNA molecules for BVDV and 34 RNA molecules for BPIV3 per reaction. We also demonstrated that the established dual RT-RPA LFD assay was precise and targeted, harboring excellent potential to become an onsite molecular diagnostic tool in the detection of BVDV and BPIV3. This method can detect BVDV (Pestivirus A, B) and BPIV3, and exhibit no cross-reaction with other viruses like the classical swine fever virus (CSFV) and infectious bovine rhinotracheitis virus (IBRV). The assay performance was further assessed with clinical samples, and demonstrated good performance in comparison to real-time RT-PCR (RT-qPCR). Moreover, the RT-RPA LFD assay was comparitively rapid and required minimal training.

Entry of bovine parainfluenza virus type 3 into MDBK cells occurs via clathrin-mediated endocytosis and macropinocytosis in a acid-dependent manner

Bovine parainfluenza virus 3 (BPIV3) is an important respiratory pathogen of both young and adult cattle. No specific therapies are available for BPIV3. Understanding the viral internalization pathway of BPIV3 will provide new strategies for the development of antiviral treatments. Here, the entry mechanism of BPIV3 into MDBK cells was analyzed using chemical inhibitors and RNA silencing. Our data demonstrated that treatment with an inhibitor targeting the clathrin-mediated pathway or clathrin heavy chain (CHC) knockdown suppressed the entry of BPIV3 into MDBK cells. In contrast, sequestration of cellular cholesterol by nystatin or silencing of caveolin-1 had no effect on viral entry. Moreover, inhibition of critical modulators of macropinocytosis significantly reduced BPIV3 uptake. In addition, fluid-phase uptake was significantly increased in cells infected with BPIV3, which is indicative of virus-induced facilitation of macropinocytosis. These results suggest that BPIV3 enters MDBK cells via macropinocytosis and clathrin- but not caveolar-dependent endocytosis. Furthermore, inhibition of endosomal acidification and activation of cathepsin blocked BPIV3 entry, demonstrating that BPIV3 entered MDBK cells in a acid-dependent manner and required cathepsin L. Finally, we demonstrated that macropinocytosis but not clathrin-mediated endocytosis is dependent on actin dynamics during BPIV3 infection.

Amino- and carboxyl-terminal ends of the bovine parainfluenza virus type 3 matrix protein are important for virion and virus-like particle release

Paramyxovirus matrix (M) proteins are key drivers of virus particle assembly and budding at the plasma membrane. To identify regions important for the M protein function, we generated a series of deletion mutants of the bovine parainfluenza virus type 3 (BPIV3) M protein. We found that M proteins lacking 10 amino acids in the amino-terminal end (ΔN10) or 4 amino acids in the carboxyl-terminal end (ΔC4) did not support M-deficient BPIV3 virion release and M protein-induced virus-like particle (VLP) release. Both ΔN10 and ΔC4 retained M protein-M protein and M protein-nucleocapsid (N) protein interactions. However, neither was transported to the plasma membrane. Our results indicate that both amino- and carboxyl-terminal ends of the BPIV3 M protein are essential for M protein transport to the plasma membrane, where it facilitates virion and VLP release.

Non-epidermidis coagulase-negative Staphylococcus isolated from farm animals can inhibit the hemagglutinating activity of Newcastle disease virus and bovine parainfluenza virus type 3

The Embp protein of Staphylococcus epidermidis inhibits the hemagglutination of the H1N1 influenza virus and protects birds from a viral respiratory infection. Several species of Coagulase-negative Staphylococcus (CoNS) are present in the respiratory cavity, particularly in nostrils. We hypothesize that non-epidermidis CoNS found in animals can have the same function as observed in S. epidermidis. Thirty Non-epidermidis CoNS isolates were obtained from poultry, sheep, goat, pig, and dairy cow nostrils. Haemagglutination inhibition (HI) activity was assayed in bacteria-free supernatants from non-epidermidis CoNS against Newcastle disease virus (NDV) and bovine parainfluenza virus type 3 (BPIV). In 13 of the 30 strains (43.3 %), bacteria-free supernatants showed HI activity for NDV and BPIV-3. Staphylococcus xylosus supernatants from poultry (one isolate), sheep (two isolates), goat (one isolate), and dairy cow (three isolates) had the highest frequency of HI activity on NDV and BPIV-3, followed by Staphylococcus sp. supernatants from goat (one isolate), dairy cow (two isolates), and finally Staphylococcus equorum, Staphylococcus chromogens and Staphylococcus gallinarum supernatants with single isolation from poultry, pig and poultry, respectively. Nine isolates had the homologous gene to the embp gene of S. epidermidis, and it was associated with HI activity in the studied viruses. By Pulsed‐field gel electrophoresis, S. xylosus isolates showed to be different clones and related to the origin of isolation and HI activity. These results demonstrate that non-epidermidis CoNS supernatants from different animals and origins have the ability of HI on NDV and BPIV-3, indicating that not only S. epidermidis has the same function.

Evolution of the Seroprevalence of Pestivirus and Respiratory Viral Infections in Spanish Feedlot Lambs.

Simple SummaryThe presence of viruses, such as bovine parainfluenza type 3, bovine herpesvirus type 1, bovine respiratory syncytial virus and pestiviruses, has been widely demonstrated in sheep in the past, but it is unknown how these infections may affect the lambs during their fattening period under commercial conditions. In the present study, the exposure to the aforementioned viruses was studied in 120 feedlot lambs throughout the whole fattening period. Seroprevalences were measured by enzyme-linked immunosorbent assays (ELISA), relating their presence to health and production parameters in the studied lambs. During the studied period, the lambs seroconverted against 3 out of the 4 viruses analysed and only 10.8% of the lambs were seronegative for all the tested viruses throughout the entire study. Finally, no associations were found between the seroconversion to one or more viruses and the presence of respiratory clinical signs or lung lesions at the slaughterhouse. In addition, no disturbances were observed in the performance of the lambs, except in the case of pestivirus infections, for which the presence of antibodies in the animals was associated with reduced final weight at the end of the fattening period.The presence of respiratory viruses and pestiviruses in sheep has been widely demonstrated, and their ability to cause injury and predispose to respiratory processes have been proven experimentally. A longitudinal observational study was performed to determine the seroprevalence of bovine parainfluenza virus type 3 (BPIV-3), bovine respiratory syncytial virus (BRSV), bovine herpesvirus type 1 (BHV-1) and pestiviruses in 120 lambs at the beginning and the end of the fattening period. During this time, the animals were clinically monitored, their growth was recorded, and post-mortem examinations were performed in order to identify the presence of pneumonic lesions in the animals. Seroconversion to all viruses tested except BHV-1 was detected at the end of the period. Initially, BPIV-3 antibodies were the most frequently found, while the most common seroconversion through the analysed period occurred to BRSV. Only 10.8% of the lambs showed no detectable levels of antibodies against any of the tested viruses at the end of the survey. In addition, no statistical differences were found in the presentation of respiratory clinical signs, pneumonic lesions nor in the production performance between lambs that seroconverted and those which did not, except in the case of pestiviruses. The seroconversion to pestiviruses was associated with a reduction in the final weight of the lambs.

Investigation of viral pathogens in cattle with bovine respiratory disease complex in Inner Mongolia, China

As a multifactor disease, the bovine respiratory disease complex (BRDC) causes high morbidity and mortality that is devastating to the cattle industry. To assess viral infections in beef cattle suffering from respiratory diseases in Inner Mongolia, 302 nasal swabs and serum samples were randomly collected from cattle with mild respiratory symptoms between March 2018 and May 2019. Our results showed that the rate of RT-PCR results positive for nucleic acids of viral pathogens in 6 cities was between 54 and 80%.The rates of bovine viral diarrhea virus (BVDV), bovine herpesvirus 1 (BHV-1), bovine parainfluenza virus type 3(BPIV3), and bovine respiratory syncytial virus(BRSV)infections were 44.70% (135/302), 24.83% (75/302), 5.63% (17/302), and 6.95% (21/302),respectively. There are also 8.94% (27/302) of samples were positive for BVDV and BHV-1, and 3.97% (12/302) of samples were positive for BPIV3 and BRSV. In addition, the RT-PCR products were sequenced, and phylogenetic analysis based on these sequences was performed. The results indicated that: a) all of the BVDV isolates were BVDV-1 and were classified as BVDV-1a (66.67%) and BVDV-1b (33.33%); b) all of the BHV-1 isolates were classified as subtype 1.1; 44.44% of the isolates were closely related to modified live viral vaccine strains, and 55.56% of the isolates were closer to epidemic strains; c) all of the BPIV3 isolates belonged to BPIV3c; d) all of the BRSV isolates were classified into subgroup III. It is suggested that an important cause of respiratory diseases for beef cattle is viral infection, and phylogenetic analysis can help us choose the proper strain to develop a vaccine.

A spraying applicator device is not required for efficacy of a live intranasal respiratory vaccine in young calves, which improves user convenience

Almost all bovine intranasal respiratory vaccines require a spraying device, but there are no published reports which prove the necessity of this. The objective of this investigation was to compare the efficacy of the Bovilis® INtranasal RSP™ Live vaccine when applied with or without an (intra)nasal spraying device. This vaccine contains live, attenuated strains of bovine respiratory syncytial virus (BRSV) and bovine parainfluenza virus type 3 (BPI3V) and is licensed to protect young cattle against respiratory infections with wild-type field isolates of these viruses.Two efficacy studies, for BRSV and BPI3V respectively, were performed in which the vaccine was administered using a spraying device or directly from the tip of a syringe as a liquid stream.Both BRSV-vaccinated groups showed a reduction of nasal shedding, BRSV titres in lung washings and clinical symptoms. The BPI3V vaccinated groups showed a significant reduction of nasal shedding and a non-significant reduction of clinical symptoms. Overall, in both studies, the groups in which vaccine was administered without the spraying device performed better than the groups with the spraying device, although this difference was not statistically significant.In conclusion, a spraying device to administer Bovilis® INtranasal RSP™ Live was not required and both application methods induced a protective immune response. This makes application more convenient and flexible for future users and animals.

Phylogenetic and antigenic analysis of bovine parainfluenza virus type 3 isolated in Japan between 2002 and 2019

Bovine parainfluenza virus type 3 (BPIV3) is one of the most important viral respiratory pathogens of cattle. In addition to the classical BPIV3 genotype A (BPIV3a), new genetic groups, genotype B (BPIV3b) and C (BPIV3c), have been identified and isolated in certain parts of the world. The present study aimed to investigate the genetic and antigenic characteristics of BPIV3 circulating in Japan. Seventy-three BPIV3 field strains were isolated from nasal samples of cattle between 2002 and 2019. Phylogenetic analysis of the phosphoprotein and hemagglutinin-neuraminidase genes showed that the isolates clustered into two genotypes, BPIV3a (49 %) and BPIV3c (51 %). The BPIV3a strains had more wide genetic variation than the rest of the genotypes. Additionally, new variants were obtained and designated them tentatively as subgroup 4 of the BPIV3a. The first Japanese BPIV3c was isolated in 2012, but here the BPIV3c NM2 strain was isolated from a sample collected four years earlier than the previous report. The antigenicity of ten BPIV3 strains including all three genotypes was assessed with a viral cross-neutralization test. Anti-sera against BPIV3a and BPIV3b cross-reacted well with both homologous and heterologous viruses. On the other hand, anti-sera against BPIV3c had reduced cross-reactivity to the heterologous viruses. Overall, our findings showed that genetically and antigenically divergent BPIV3 is prevalent in cattle in Japan. These results could provide a reference for molecular epidemiological characterization of BPIV3 and vaccine development.

Detection of bovine viral diarrhea virus genotype 1 in aerosol by a real time RT-PCR assay

BackgroundAs a pestivirus of the Flaviviridae family, bovine viral diarrhea virus (BVDV), has imposed a large burden on animal husbandry worldwide, and such virus can be transmitted mainly through direct contact with other infected animals and probably via aerosols. In the present study, we aimed to develop a real-time RT-PCR method for detection of BVDV-1 in aerosol samples.MethodsA pair of primers specific for highly conserved regions of the BVDV-1 5′-UTR was designed. The standard curve and sensitivity of the developed assay were assessed based on 10-fold serial dilutions of RNA molecular standard. The specificity of the assay was evaluated with other pestiviruses and infectious bovine viruses. The clinical performance was examined by testing 169 aerosol samples.ResultsThe results showed that a good linear relationship existed between the standard curve and the concentration of template. The lowest detection limit was 5.2 RNA molecules per reaction. This assay was specific for detection of BVDV-1, and no amplification was found for other pestiviruses such as classical swine fever virus (CSFV), border disease virus (BDV), and common infectious bovine viruses, including BVDV-2, infectious bovine rhinotracheitis virus (IBRV), bovine parainfluenza virus type 3 (BPIV-3), bovine respiratory syncytial virus (BRSV), bovine ephemeral fever virus (BEFV) and bovine coronavirus (BcoV). The assay was highly reproducible with low variation coefficient values (CVs) for intra-assay and inter-assay. A total of 169 aerosol samples collected from six dairy herds were tested using this method. The results showed that the positive detection rate of BVDV-1 was 17.2% (29/169), which was significantly higher compared with the conventional RT-PCR. Additionally, the positive samples (n = 29) detected by real-time RT-PCR were verified by BVDV RPA-LFD, and a concordance rate of 100% was obtained between them.ConclusionsTaken together, we developed a real-time RT-PCR assay for quantitative analysis of BVDV-1 in aerosol samples, and our finding provided valuable insights into the risk on aerosol transmission of BVDV-1.

Serodetection Bovine Herpesvirus Types 1, 4 and Bovine Parainfluenza Virus Type 3 Infections in Milk of Cows with Clinical Mastitis Based in Dairy Cattle Management in Turkey

Aims: The present study aimed to Searching Bovine Herpesvirus Types 1, 4 and Bovine Parainfluenza Virus Type 3 Infections in Milk of Cows with Clinical Mastitis Based in Dairy Cattle Managements.
 Study Design: In 35 managements around Burdur region, milk of 123 cows with clinical mastitis was searched for Bovine Herpesvirus Types 1 (BHV-1), 4 (BHV-4) and Bovine Parainfluenza Virus Type 3 (BPIV-3) infections.
 Results: In the study, the highest seropositivity was detected against BPIV-3. The highest seropositivity on infection distribution according to age was found against every four viruses in animals within the three-year-old group. The highest seropositivity in this group and other age groups was detected against BPIV-3. Seropositivity against these viruses was found in the highest right anterior one and the lowest left posterior one out of determined udder lobes. The highest seropositivity was found in semi-outdoor managements with concrete and dirty grounds where cleaning/disinfection of teats before and after milking was performed, mastitis treatment and viral vaccination was not applied, the ground of stable was cleaned monthly, only feces was collected from the grounds, water was used for cleaning of milking machines and hands and iodised disinfectant was used. During udder inspection evaluation of animals with clinical mastitis, the highest seropositivity was detected in the ones having normal udder, teat and skin appearance. Out of udder and teat skin lesions, the highest seropositivity was found in crushed ones. In the evaluation of udder palpation in animals with clinical mastitis, the highest seropositivity was detected in the ones with teat tissue thickening and with elastic teat sinuses and lobes. In the milk of these animals, the highest seropositivity was found in the ones showing coagulation.
 Conclusion: It was stated that viruses took a vital part in clinical mastitis cases, the structure, practice, cleaning and disinfection of managements was really important, udder and teat skin lesions, tissue thickening, elastic teat sinuses and lobes and coagulation of milk was also crucial. Besides, the appearance of udder, teat and skin was not relevant.

Determination of serum amyloid A, haptoglobin and hepcidin levels in calves with endemic viral pneumonia

The aim of this study was to investigate serum amyloid A, haptoglobin and hepcidin levels in respiratory infections caused by various viral agents in calves. Pneumonia is an infectious or non-infectious disease affecting the lungs of all animals, and during the inflammation, blood levels of acute phase proteins increase rapidly. Hepcidin is a hormone that has many functions and is known as type II acute phase reactant. The blood samples were collected from a total of 20 calves to form the study group. The control group consisted of 10 healthy animals. Sera were analyzed with ELISA for antigen detection. According to the results, Bovine herpesvirus type-1 (BHV-1) was positive in 3 calves, Bovine viral diarrhea virus (BVDV) was positive in 3 calves, Bovine respiratory syncytial virus (BRSV) was positive in 1 calf, and Bovine parainfluenza virus type 3 (BPI3) was positive in 1 calf. Adenovirus infection was detected in 5 animals together with other viruses. Mixed infections were observed in 12 calves. Haptoglobin, serum amyloid A and hepcidin values were measured in sera. According to the results, statistically significant differences were observed between the two groups for the haptoglobin (P<0.01), serum amyloid A (P<0.05) and hepcidin (P<0.05). It was concluded that these parameters can be used to support the diagnosis of respiratory infections of the calves.