Comparison of the immune response following subcutaneous versus intranasal modified-live virus booster vaccination against bovine respiratory disease in pre-weaning beef calves that had received primary vaccination by the intranasal route.

This study was performed to elucidate whether the route of booster vaccination affects the immune response against respiratory vaccine viruses in pre-weaning beef calves that receive primary intranasal (IN) vaccination during the first month of life. The objective was to compare the serum neutralizing antibody (SNA) titers to BHV1, BRSV, and BPI3V, cytokine mRNA expression and mucosal BHV1- and BRSV-specific IgA in nasal secretions following administration of IN or subcutaneous (SC) modified-live virus (MLV) booster vaccines 60 days after primary IN vaccination in young beef calves. Twenty-one beef calves were administered 2 mL of an IN MLV vaccine containing BHV1, BRSV, and BPI3V (Inforce3®) between one and five weeks of age. Sixty days after primary vaccination, calves were randomly assigned to one of two groups: IN-MLV (n = 11): Calves received 2 mL of the same IN MLV vaccine used for primary vaccination and 2 mL of a SC MLV vaccine containing BVDV1 & 2 (Bovi- Shield GOLD® BVD). SC-MLV (n = 10): Calves were administered 2 mL of a MLV vaccine containing, BHV1, BRSV, BPI3V, and BVDV1 & 2 (Bovi-Shield GOLD® 5). Blood and nasal secretion samples were collected on days -61 (primary vaccination), -28, -14, 0 (booster vaccination), 14, 21, 28, 42 and 60 for determination of SNA titers, cytokine gene expression analysis and nasal virus-specific IgA concentrations. Statistical analysis was performed using a repeated measures analysis through PROC GLIMMIX of SAS®. Booster vaccination by neither IN nor SC routes induced a significant increase in SNA titers against BHV1, BRSV, and BPI3V. Subcutaneous booster vaccination induced significantly greater BRSV-specific SNA titers (on day 42) and IgA concentration in nasal secretions (on days 21 and 42) compared to calves receiving IN booster vaccination. Both IN and SC booster vaccination were able to stimulate the production of BHV1-specific IgA in nasal secretions. In summary, booster vaccination of young beef calves using either SC or IN route two months after IN MLV primary vaccination resulted in comparable SNA titers, cytokine gene expression profile and virus-specific IgA concentration in nasal secretions. Only a few differences in the systemic and mucosal immune response against BHV1 and BRSV were observed. Subcutaneous booster vaccination induced significantly greater BRSV-specific SNA and secretory IgA titers compared to IN booster vaccination.

A Recombinant BCG Vaccine Is Safe and Immunogenic in Neonatal Calves and Reduces the Clinical Disease Caused by the Respiratory Syncytial Virus.

The human respiratory syncytial virus (hRSV) constitutes a major health burden, causing millions of hospitalizations in children under five years old worldwide due to acute lower respiratory tract infections. Despite decades of research, licensed vaccines to prevent hRSV are not available. Development of vaccines against hRSV targeting young infants requires ruling out potential vaccine-enhanced disease presentations. To achieve this goal, vaccine testing in proper animal models is essential. A recombinant BCG vaccine that expresses the Nucleoprotein of hRSV (rBCG-N-hRSV) protects mice against hRSV infection, eliciting humoral and cellular immune protection. Further, this vaccine was shown to be safe and immunogenic in human adult volunteers. Here, we evaluated the safety, immunogenicity, and protective efficacy of the rBCG-N-hRSV vaccine in a neonatal bovine RSV calf infection model. Newborn, colostrum-replete Holstein calves were either vaccinated with rBCG-N-hRSV, WT-BCG, or left unvaccinated, and then inoculated via aerosol challenge with bRSV strain 375. Vaccination with rBCG-N-hRSV was safe and well-tolerated, with no systemic adverse effects. There was no evidence of vaccine-enhanced disease following bRSV challenge of rBCG-N-hRSV vaccinated animals, suggesting that the vaccine is safe for use in neonates. Vaccination increased virus-specific IgA and virus-neutralization activity in nasal fluid and increased the proliferation of virus- and BCG-specific CD4+ and CD8+ T cells in PBMCs and lymph nodes at 7dpi. Furthermore, rBCG-N-hRSV vaccinated calves developed reduced clinical disease as compared to unvaccinated control calves, although neither pathology nor viral burden were significantly reduced in the lungs. These results suggest that the rBCG-N-hRSV vaccine is safe in neonatal calves and induces protective humoral and cellular immunity against this respiratory virus. These data from a newborn animal model provide further support to the notion that this vaccine approach could be considered as a candidate for infant immunization against RSV.

Impact of meloxicam on respiratory virus titers and health outcomes when administered concurrently with a modified live respiratory vaccine in abruptly weaned beef steers.

Abruptly weaned crossbred steer calves (N = 271) were used in a randomized, blinded 2-arm clinical trial to assess the impact of a long-acting non-steroidal anti-inflammatory drug on bovine herpesvirus type 1, bovine respiratory syncytial virus, parainfluenza virus type 3, and coronavirus titers and health outcomes when administered concurrently with a modified live respiratory vaccine upon arrival at a feedlot. Treatment groups included a control (saline; n = 135) and an experimental group (injectable meloxicam; n = 136). Viral antibody titers and body weight were measured on arrival, day 7, and day 21, along with a final weight on day 45. Body weight and antibody titers for all viruses increased over time (P < 0.001); however, there were no differences by treatment group or a significant group × time interaction when evaluated using repeated measures analysis of variance. Interestingly, the use of meloxicam was associated with increased treatment risk (P < 0.05). In conclusion, the administration of meloxicam may adversely affect health; however, a decreased vaccine response is likely not a contributing factor.

Des bouvillons croisés sevrés rapidement (N = 271) ont été utilisés dans un essai clinique randomisé en aveugle à deux bras pour évaluer l'impact d'un anti-inflammatoire non stéroïdien à action prolongée sur les titres du virus de la rhinotrachéite infectieuse bovine, du virus respiratoire syncytial bovin, du virus parainfluenza 3 et du coronavirus, et les résultats pour la santé lorsqu'administré en même temps qu'un vaccin vivant modifié respiratoire à l'arrivée dans un parc d'engraissement. Les groupes de traitement comprenaient un témoin (solution saline; n = 135) et un groupe expérimental (méloxicam injectable; n = 136). Les titres d'anticorps viraux et le poids corporel ont été mesurés à l'arrivée, au jour 7 et au jour 21, ainsi qu'un poids final au jour 45. Le poids corporel et les titres d'anticorps pour tous les virus ont augmenté avec le temps (P < 0,001); cependant, il n'y avait aucune différence selon le groupe de traitement ou une interaction groupe × temps significative lors de l'évaluation à l'aide de mesures répétées d'analyse de la variance. Fait intéressant, l'utilisation du méloxicam était associée à un risque de traitement accru (P < 0,05). En conclusion, l'administration de méloxicam peut nuire à la santé; cependant, une réponse vaccinale réduite n'est probablement pas un facteur contributif.(Traduit par Docteur Serge Messier).

NASYM, a live intranasal vaccine, protects young calves from bovine respiratory syncytial virus in the presence of maternal antibodies.

BACKGROUND: Bovine respiratory syncytial virus (BRSV) is a major problem for cattle worldwide during their first year of life. The aim of the present study was to evaluate efficacy and longevity of immunity of a live vaccine (NASYM, HIPRA) in the presence of maternally derived antibodies (MDA). METHOD: Calves (36) were distributed in four groups, based on MDA status and treatment. They received NASYM or a placebo at an early age (less than two weeks) by intranasal route. Eight weeks later, animals were challenged with the Asquith strain of BRSV. Efficacy was assessed by monitoring clinical signs and mortality, PaO2 , virus shedding and lung lesions. The immunological response was evaluated by measuring IgG in serum and IgA in nasal secretions. RESULTS: A reduction of mortality, lung lesions, shedding and a higher PaO2 was achieved in NASYM vaccinated groups, independently of MDA status. An anamnestic IgG response was observed after challenge in vaccinated animals, both in MDA+ and MDA- groups. An IgA response was also observed in vaccinated animals after vaccination and challenge. CONCLUSION: NASYM protected newborn calves with MDAs during the first 10 weeks of life, against a very virulent challenge that caused extensive pulmonary lesions and deaths in control animals, with just a single intranasal dose.

Dynamics of neutralizing antibodies against Bovine respiratory syncytial virus in a dairy herd from Santa Fe Province, Argentina

Abstract Bovine respiratory syncytial virus (BRSV) is one of the most relevant agents responsi-ble for respiratory disease in cattle from both dairy and beef farms. BRSV is spread by horizontalcontact causing a constant presence of seropositive animals that favors viral circulation throughout the year. Moreover, reinfections with BRSV are frequent between animals regardless of theirage as BRSV does not confer long-lasting protective immunity. Several studies have demonstrated the circulation of BRSV in cattle from different regions of the world; however, little isknown about the dynamics of BRSV infection in cows before and after they begin lactation. Theaim of this work was to study the dynamics of BRSV neutralizing antibodies from birth up to36 months of age in a closed dairy herd of Argentina specifically around the lactation period. Passive maternal antibodies against BRSV started to decrease monthly and became almost undetectable at 8 months of age. We detected two potential infection points at months 11 and 27after birth, in which 30% and 45% of the animals showed seroconversion, respectively. Specifically, an increase in the proportion of seropositive cows after the start of lactation suggests thatthey became reinfected around the time they began lactating. We demonstrate the importanceof understanding BRSV dynamics in a closed dairy herd to review the vaccination schedule ofthe animals to achieve protection against BRSV infection.

Resumen El virus respiratorio sincitial bovino (Bovine respiratory syncytial virus, [BRSV]) es uno de los principales agentes responsables de la enfermedad respiratoria en bovinos, tanto de tambos como de cría. El virus se transmite horizontalmente y causa la presencia constante de animales seropositivos, lo cual favorece la circulación viral a lo largo del ano. A su vez, las reinfecciones por BRSV son frecuentes entre animales independientemente de su edad, dado que el virus no confiere inmunidad protectora a largo plazo. Numerosos estudios han demostrado la circulación de BRSV en bovinos de diferentes regiones del mundo, sin embargo, poco se conoce acerca de la dinámica de infección en vacas antes y después del inicio de la fase de lactancia. El objetivo de este trabajo fue estudiar la dinámica de anticuerpos neutralizantes anti- BRSV en vacas lecheras desde el nacimiento hasta los 36 meses de vida en un tambo cerrado de Argentina, específicamente, en el período de lactancia. Los anticuerpos pasivos específicos para BRSV comenzaron a declinar mensualmente hasta ser casi indetectables a los 6 meses. Detectamos dos potenciales puntos de infección a los meses 11 y 27 luego del nacimiento, momentos en los que el 30 y el 45% de los animales mostraron seroconversión, respectivamente. El incremento en la proporción de vacas seropositivas luego del comienzo de la lactancia sugiere que estas se reinfectaron en el inicio de dicha etapa. Demostramos la importancia de entender la dinámica de circulación del BRSV en un tambo cerrado, a fin de revisar el esquema de vacunación de los animales para que estén protegidos frente a la posible infección por este virus.

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.

Effectiveness of two intranasal vaccines for the control of bovine respiratory disease in newborn beef calves: A randomized non-inferiority multicentre field trial.

Bovine respiratory syncytial virus (BRSV) and bovine parainfluenza-3 virus (bPI3V) are major causes of bovine respiratory disease (BRD) in newborn calves worldwide. Vaccination is widely used to prevent BRD, and intranasal vaccines for BRSV and bPI3V were developed to overcome interference from BRSV and bPI3V-specific maternally derived antibodies. Many experimental challenge trials have demonstrated that intranasal vaccines for BRSV and bPI3V are efficacious, but effectiveness under field conditions has been demonstrated less often, especially for newborn beef calves. The objective of this field trial was to compare the effectiveness of a newly available commercial BRSV-bPI3V intranasal vaccine with that of a benchmarked one in newborn beef calves reared in a cow-calf system. A total of 935 calves from 39 farms were randomized into two vaccine groups (Bovalto Respi Intranasal [Vaccine A], n=468; Rispoval RS+PI3 Intranasal [Vaccine B], n=467), and monitored during the in-house risk period up to three months after vaccination. Non-inferiority analysis was performed by calculating the difference in BRD prevalence between the two vaccine groups. No significant differences were observed between vaccines regarding clinical outcomes of morbidity, mortality, duration between vaccination and BRD occurrence, or treatments required. Because the upper limit of the 2-sided 95% confidence interval of the difference in BRD prevalence between the two treatment groups (0.8%) was less than the margin of non-inferiority (δ=5%), a non-inferiority of Vaccine A was concluded. In conclusion, Vaccine A is at least as effective as Vaccine B for the prevention of BRD in newborn beef cattle in a cow-calf system under field conditions.

Supplementing a Saccharomyces cerevisiae fermentation product modulates innate immune function and ameliorates bovine respiratory syncytial virus infection in neonatal calves.

The objectives of this study were to determine the effects of oral supplementation with Saccharomyces cerevisiae fermentation products (SCFP; SmartCare and NutriTek; Diamond V, Cedar Rapids, IA) on immune function and bovine respiratory syncytial virus (BRSV) infection in preweaned dairy calves. Twenty-four Holstein × Angus, 1- to 2-d-old calves (38.46 ± 0.91 kg initial body weight [BW]) were assigned two treatment groups: control or SCFP treated, milk replacer with 1 g/d SCFP (SmartCare) and calf starter top-dressed with 5 g/d SCFP (NutriTek). The study consisted of one 31-d period. On days 19 to 21 of the supplementation period, calves were challenged via aerosol inoculation with BRSV strain 375. Calves were monitored twice daily for clinical signs, including rectal temperature, cough, nasal and ocular discharge, respiration effort, and lung auscultation. Calves were euthanized on day 10 postinfection (days 29 to 31 of the supplementation period) to evaluate gross lung pathology and pathogen load. Supplementation with SCFP did not affect BW (P = 0.762) or average daily gain (P = 0.750), percentages of circulating white blood cells (P < 0.05), phagocytic (P = 0.427 for neutrophils and P = 0.460 for monocytes) or respiratory burst (P = 0.119 for neutrophils and P = 0.414 for monocytes) activity by circulating leukocytes either before or following BRSV infection, or serum cortisol concentrations (P = 0.321) after BRSV infection. Calves receiving SCFP had reduced clinical disease scores compared with control calves (P = 0.030), reduced airway neutrophil recruitment (P < 0.002), reduced lung pathology (P = 0.031), and a reduced incidence of secondary bacterial infection. Calves receiving SCFP shed reduced virus compared with control calves (P = 0.049) and tended toward lower viral loads in the lungs (P = 0.051). Immune cells from the peripheral blood of SCFP-treated calves produced increased (P < 0.05) quantities of interleukin (IL)-6 and tumor necrosis factor-alpha in response to toll-like receptor stimulation, while cells from the bronchoalveolar lavage (BAL) of SCFP-treated calves secreted less (P < 0.05) proinflammatory cytokines in response to the same stimuli. Treatment with SCFP had no effect on virus-specific T cell responses in the blood but resulted in reduced (P = 0.045) virus-specific IL-17 secretion by T cells in the BAL. Supplementing with SCFP modulates both systemic and mucosal immune responses and may improve the outcome of an acute respiratory viral infection in preweaned dairy calves.

Technical Note: Using enzyme-linked immunosorbent assays to evaluate humoral responses to vaccination against respiratory viruses in beef cattle.

This experiment evaluated humoral responses in beef calves vaccinated against parainfluenza-3 virus (PI3), bovine respiratory syncytial virus (BRSV), and bovine herpesvirus-1 (BHV-1) using serum neutralization (SN) tests or enzyme-linked immunosorbent assays (ELISA). Blood samples were collected from 50 overtly healthy Angus-influenced steers (183 ± 3 kg of body weight, 212 ± 2 d of age) on days 0, 21, 35, and 49 of the experiment. Steers were vaccinated against respiratory viruses on days 0 and 21. Blood was processed for serum collection and frozen in duplicates. One of the duplicates was analyzed for antibodies against BRSV, PI3, and BHV-1 using commercially available ELISA (IDEXX Switzerland AG, Liebefeld-Bern, Switzerland), and results reported as sample:positive control (S/P, %) ratio. The other duplicate was analyzed for antibodies against the same vaccine antigens via SN. This method reports results as titers, the greatest dilution that provides complete protection of the cells, which were transformed with base 2 log for statistical analyses. Samples were classified as positive for the presence of antibodies by SN if log-transformed titer ≥ 2 for all viruses, and by ELISA if S/P ratio ≥ 50% for BHV-1 or ≥ 20% for PI3 and BRSV. Day effects were detected (P < 0.01) for SN and ELISA across all vaccine antigens, as antibody levels increased after vaccine administration. Linear fits were detected (P < 0.01) across all vaccine antigens when regressing the SN and ELISA results; as SN titer increased, the ELISA S/P ratio linearly increased (P < 0.01). Kendall (τ) and Spearman's rank (ρ) correlations were also detected (P < 0.01) between SN and ELISA results across all vaccine antigens. The SN and ELISA were very strongly correlated (ρ ≥ 0.83) for BHV-1 and PI3 and strongly correlated (ρ = 0.66) for BRSV. Cohen's kappa coefficient for diagnosis agreement between methods was strong for BHV-1 and PI3 (κ ≥ 0.88), but weak (κ = 0.47) for BRSV. The sensitivity of the ELISA in yielding true positive results approached 100% across all antigens. The specificity of the ELISA in yielding negative results was satisfactory for BHV-1 and PI3 assays (84.0% and 88.5%, respectively) but not for BRSV (34.4%). Despite limitations in detecting true BRSV negatives, results from this experiment indicate that the commercial ELISAs tested herein can be used as surrogate for SN tests in quantifying humoral responses to vaccination against BHV-1, PI3, and BRSV in beef cattle.

Dynamics of neutralizing antibodies against Bovine respiratory syncytial virus in a dairy herd from Santa Fe Province, Argentina.

Bovine respiratory syncytial virus (BRSV) is one of the most relevant agents responsible for respiratory disease in cattle from both dairy and beef farms. BRSV is spread by horizontal contact causing a constant presence of seropositive animals that favors viral circulation throughout the year. Moreover, reinfections with BRSV are frequent between animals regardless of their age as BRSV does not confer long-lasting protective immunity. Several studies have demonstrated the circulation of BRSV in cattle from different regions of the world; however, little is known about the dynamics of BRSV infection in cows before and after they begin lactation. The aim of this work was to study the dynamics of BRSV neutralizing antibodies from birth up to 36 months of age in a closed dairy herd of Argentina specifically around the lactation period. Passive maternal antibodies against BRSV started to decrease monthly and became almost undetectable at 8 months of age. We detected two potential infection points at months 11 and 27 after birth, in which 30% and 45% of the animals showed seroconversion, respectively. Specifically, an increase in the proportion of seropositive cows after the start of lactation suggests that they became reinfected around the time they began lactating. We demonstrate the importance of understanding BRSV dynamics in a closed dairy herd to review the vaccination schedule of the animals to achieve protection against BRSV infection.

El virus respiratorio sincitial bovino (Bovine respiratory syncytial virus, [BRSV]) es uno de los principales agentes responsables de la enfermedad respiratoria en bovinos, tanto de tambos como de cría. El virus se transmite horizontalmente y causa la presencia constante de animales seropositivos, lo cual favorece la circulación viral a lo largo del año. A su vez, las reinfecciones por BRSV son frecuentes entre animales independientemente de su edad, dado que el virus no confiere inmunidad protectora a largo plazo. Numerosos estudios han demostrado la circulación de BRSV en bovinos de diferentes regiones del mundo, sin embargo, poco se conoce acerca de la dinámica de infección en vacas antes y después del inicio de la fase de lactancia. El objetivo de este trabajo fue estudiar la dinámica de anticuerpos neutralizantes anti- BRSV en vacas lecheras desde el nacimiento hasta los 36 meses de vida en un tambo cerrado de Argentina, específicamente, en el período de lactancia. Los anticuerpos pasivos específicos para BRSV comenzaron a declinar mensualmente hasta ser casi indetectables a los 6 meses. Detectamos dos potenciales puntos de infección a los meses 11 y 27 luego del nacimiento, momentos en los que el 30 y el 45% de los animales mostraron seroconversión, respectivamente. El incremento en la proporción de vacas seropositivas luego del comienzo de la lactancia sugiere que estas se reinfectaron en el inicio de dicha etapa. Demostramos la importancia de entender la dinámica de circulación del BRSV en un tambo cerrado, a fin de revisar el esquema de vacunación de los animales para que estén protegidos frente a la posible infección por este virus.

Mycoplasma bovis and viral agents associated with the development of bovine respiratory disease in adult dairy cows.

The etiology and pathologic findings of bovine respiratory disease (BRD) in adult dairy cows (n = 35) from a commercial dairy herd in Southern Brazil were investigated. Pulmonary samples were examined for histopathologic patterns and specific features within these patterns, while immunohistochemical (IHC) assays were designed to detect the intralesional antigens of viral infectious disease agents and Mycoplasma bovis. Pneumonia was diagnosed in 91.4% (32/35) of these cases; neither pneumonia nor any of the infectious disease pathogens evaluated occurred in three cows. The presence of multiple respiratory pathogens in 75% (24/32) of these cases indicated the complex origin of pneumonia in cattle. Interstitial pneumonia, necrosuppurative bronchopneumonia and suppurative bronchopneumonia were the principal patterns of pulmonary disease identified by histopathology. The most frequent pathogens identified by IHC were bovine viral diarrhea virus (BVDV; n = 18), M. bovis (n = 16) and bovine alphaherpesvirus type 1 (BoHV-1; n = 14), followed by bovine respiratory syncytial virus (BRSV; n = 11) and bovine parainfluenza virus type 3 (BPIV-3; n = 5). Obliterative bronchiolitis and peribronchial lymphocytic cuffings were the characteristic histopathologic features associated with M. bovis. Necrohemorrhagic bronchitis with bronchial angiogenesis was associated with BoHV-1. Necrotizing bronchitis and bronchiolitis were associated with BVDV, BoHV-1 and BRSV. Ballooning degeneration of the bronchial and bronchiolar epithelia was associated with BRSV and BoHV-1. This is the first report from Brazil that correlated the histopathologic findings of BRD with the associated infectious disease agents by immunohistochemistry. M. bovis was frequently detected in the tissues of cows with fatal pulmonary disease during this study and may be a possible primary disease pathogen associated with the development of BRD in dairy cows. Additionally, the histopathologic features identified within patterns of pulmonary disease during this investigation may be an efficient diagnostic tool to associate histopathologic findings with specific agents of BRD in dairy cows.

Protection against bovine respiratory syncytial virus in calves vaccinated with adjuvanted modified live vaccine administered in the face of maternal antibody.

Bovine respiratory syncytial virus (BRSV) is major viral contributor to bovine respiratory disease (BRD). BRD is a major cause of morbidity and mortality in all classes of cattle but particularly young beef and dairy calves. Passive antibodies not only help protect the calf against infection, but may interfere with the immune responses following vaccination. The purpose of this study was to evaluate the efficacy of an adjuvanted modified live virus (MLV) vaccine in the presence of well-defined maternal passive immunity. Calves were vaccinated at approximately 1 month of age and challenged ~90 days later when BRSV systemic antibodies were ≤1:4. Body temperature was lower at 6 and 7 days post challenge and other clinical signs were also lower in the vaccinates. Nasal viral shed was 3-4 times lower in the vaccinated animals as measured by virus isolation and polymerase chain reaction (PCR) and peaked 5 days post challenge compared to the controls (who peaked at days 6 and 7). On day 8 following challenge, animals were necropsied, and lung lobes were scored and tested for virus by PCR and indirect fluorescent assay (IFA). There was a 25-fold reduction in PCR virus detection in vaccinates and two of the vaccinated calves' lungs were PCR negative. Only 29.4% of vaccinated calves were BRSV positive on IFA testing at necropsy, while 87.5% of control calves were BRSV positive. Vaccinated calves developed a mucosal BRSV IgA response with over 50% of the vaccinated calves having IgA prior to challenge and all vaccinated calves were positive following challenge. Additionally, vaccination stimulated the production of Interferon gamma (IFN-γ) in mononuclear cells to prime the immune system. This study established that an adjuvanted MLV vaccine could provide protection against BRSV as measured by clinical, virological, and pathological parameters while also activating both mucosal and systemic immunity.

Vitamin A deficiency impairs the immune response to intranasal vaccination and RSV infection in neonatal calves.

Respiratory syncytial virus (RSV) infection is a leading cause of severe acute lower respiratory tract infection in infants and children worldwide. Vitamin A deficiency (VAD) is one of the most prevalent nutrition-related health problems in the world and is a significant risk factor in the development of severe respiratory infections in infants and young children. Bovine RSV (BRSV) is a primary cause of lower respiratory tract disease in young cattle. The calf model of BRSV infection is useful to understand the immune response to human RSV infection. We have previously developed an amphiphilic polyanhydride nanoparticle (NP)-based vaccine (i.e., nanovaccine) encapsulating the fusion and attachment proteins from BRSV (BRSV-NP). Calves receiving a single, intranasal dose of the BRSV-NP vaccine are partially protected from BRSV challenge. Here, we evaluated the impact of VAD on the immune response to the BRSV-NP vaccine and subsequent challenge with BRSV. Our results show that VAD calves are unable to respond to the mucosal BRSV-NP vaccine, are afforded no protection from BRSV challenge and have significant abnormalities in the inflammatory response in the infected lung. We further show that acute BRSV infection negatively impacts serum and liver retinol, rendering even well-nourished individuals susceptible to VAD. Our results support the use of the calf model for elucidating the impact of nutritional status on mucosal immunity and respiratory viral infection in infants and underline the importance of VA in regulating immunity in the respiratory mucosa.

[Risk factors for calf mortality influence the occurrence of antibodies against the pathogens of enzootic bronchopneumonia]. / Risikofaktoren für die Kälbersterblichkeit beeinflussen den Nachweis von Antikörpern gegen die Erreger der enzootischen Bronchopneumonie.

OBJECTIVE: Bovine respiratory diseases are a common cause of calf loss. This study aimed to analyse associations between an occurrence of enzootic bronchopneumonia (EBP), calf mortality and calving management. MATERIAL AND METHODS: A total of 153 dairy farms participated in the study on a voluntary basis from November 2006 to July 2007. Calf management was inspected on-site during a farm visit and farm managers were required to complete a questionnaire on personal assessment of calving procedures, neonate management and environmental factors. Results were collated and matched with the calf mortality rate of 2006 determined from the HI-Tier database for each farm. Randomly selected serum samples of a mean number of 7 calves at the age 6 months per herd were investigated for antibodies against bovine respiratory syncytial virus (BRSV-AB) and parainfluenzavirus type 3 (PIV3-AB). According to the proportion of calves with BRSV-AB or PIV3-AB (≤ 20 % or > 20 %) farms were divided into 2 groups. RESULTS: Customary timing of the first colostrum feeding as well as the perceived level of importance of EBP to the farm manager, as described in the questionnaire, showed a positive correlation to calf mortality. BRSV-AB occurred more frequently on farms where managers stated that the first colostrum feeding occurred later than 4 hours after birth, that birth monitoring was rarely practiced and that the estimated level of dust in the calf barn was considered high. PIV3-AB was more frequently found at farms practicing tethered calving. CONCLUSION AND CLINICAL RELEVANCE: The results of this study indicate that peri- and postnatal calf management procedures may affect calf mortality and the frequency of occurrence of BRSV-AB or PIV3-AB. The influences of birth monitoring and the time of first colostrum feeding as well as dust exposure should be taken into account in future studies on the frequency of EBP and be included in the veterinary cause analysis of herd EBP-related problems.

Profiling of local disease-sparing responses to bovine respiratory syncytial virus in intranasally vaccinated and challenged calves.

Bovine and human respiratory syncytial viruses (BRSV, HRSV) are primary causes of pneumonia in calves and children respectively, with vaccination offering protection via antibody and cellular immune responses. However, with no vaccines currently licensed for human use, evaluation of local responses to BRSV vaccination may provide insights to aid the design of effective safe HRSV vaccines. Calves received intranasal single component BRSV vaccine or "3-Way" vaccine (BRSV, Bovine Herpes Virus-1 (BHV-1), Bovine Parainfluenza Virus Type-3 (BPIV-3)), and were BRSV-challenged 42 days post-vaccination. All vaccinates exhibited reduced pulmonary lesioning with elevated anti-BRSV serum IgG, and higher nasal anti-BRSV IgA in 3-Way vaccinates. Thirty-nine proteins associated with homeostatic and immune processes were altered in vaccinates, with enhanced 3-Way vaccinate group proteins associated with Th1/Th2 balance and immunoglobulin class switching. Proteins altered in the pharyngeal tonsil of animals euthanized early related to anti-inflammatory responses and lymphoid tissue remodeling. These findings indicate that multivalent vaccines distinctly modulate local immune responses, with clear correlation between the pharyngeal tonsil proteome profile and resulting immune protection and disease-sparing. This suggests that the efficacy of low-antigenic subunit vaccine components for problematic pathogens such as HRSV could be enhanced by use in combination with existing safe live vaccines. SIGNIFICANCE: This study demonstrates that vaccine valency can alter post-challenge proteome responses within the pharyngeal tonsil, a sentinel site of primary immune responses, with the magnitude of response dependent on antigen formulation. Observed differential responses can be attributed to antigenic material and viral nucleic acid from multivalent formulations providing additional T-cell epitopes and PAMPS. These findings indicate that incorporation of subunits proteins within multivalent formulations containing live virus has the potential to induce/skew a favorable immune response, utilising the natural adjuvanting effects of safe proven live vaccines.

The role of the light chain in the structure and binding activity of two cattle antibodies that neutralize bovine respiratory syncytial virus.

Cattle antibodies have unusually long CDR3 loops in their heavy chains (HCs), and limited light chain (LC) diversity, raising the question of whether these mask the effect of LC variation on antigen recognition. We have investigated the role of the LC in the structure and activity of two neutralizing cattle antibodies (B4 and B13) that bind the F protein of bovine respiratory syncytial virus (bRSV). Recombinant Fab fragments of B4 and B13 bound bRSV infected cells and showed similar affinities for purified bRSV F protein. Exchanging the LCs between the Fab fragments produced hybrid Fabs: B13* (B13 HC/B4 LC) and B4* (B4 HC/B13 LC). The affinity of B13* to the F protein was found to be two-fold lower than B13 whilst the binding affinity of B4* was reduced at least a hundred-fold compared to B4 such that it no longer bound to bRSV infected cells. Comparison of the structures of B4 and B13 with their LC exchanged counterparts B4* and B13* showed that paratope of the HC variable domain (VH) of B4 was disrupted on pairing with the B13 LC, consistent with the loss of binding activity. By contrast, B13 H3 adopts a similar conformation when paired with either B13 or B4 LCs. These observations confirm the expected key role of the extended H3 loop in antigen-binding by cattle antibodies but also show that the quaternary LC/HC subunit interaction can be crucial for its presentation and thus the LC variable domain (VL) is also important for antigen recognition.

Prophylactic digoxin treatment reduces IL-17 production in vivo in the neonatal calf and moderates RSV-associated disease.

Respiratory syncytial virus (RSV) is a leading cause of morbidity and mortality in human infants. Bovine RSV infection of neonatal calves is pathologically and immunologically similar to RSV infection in infants, and is therefore a useful preclinical model for testing novel therapeutics. Treatment of severe RSV bronchiolitis relies on supportive care and may include use of bronchodilators and inhaled or systemic corticosteroids. Interleukin-17A (IL-17) is an inflammatory cytokine that plays an important role in neutrophil recruitment and activation. IL-17 is increased in children and rodents with severe RSV infection; and in calves with severe BRSV infection. It is currently unclear if IL-17 and Th17 immunity is beneficial or detrimental to the host during RSV infection. Digoxin was recently identified to selectively inhibit IL-17 production by antagonizing its transcription factor, retinoid-related orphan receptor γ t (RORγt). Digoxin inhibits RORγt binding to IL-17 and Th17 associated genes, and suppresses IL-17 production in vitro in human and murine leukocytes and in vivo in rodent models of autoimmune disease. We demonstrate here that in vitro and in vivo digoxin treatment also inhibits IL-17 production by bovine leukocytes. To determine the role of IL-17 in primary RSV infection, calves were treated prophylactically with digoxin and infected with BRSV. Digoxin treated calves demonstrated reduced signs of clinical illness after BRSV infection, and reduced lung pathology compared to untreated control calves. Digoxin treatment did not adversely affect virus shedding or lung viral burden, but had a significant impact on pulmonary inflammatory cytokine expression on day 10 post infection. Together, our results suggest that exacerbated expression of IL-17 has a negative impact on RSV disease, and that development of specific therapies targeting Th17 immunity may be a promising strategy to improve disease outcome during severe RSV infection.

Genome-wide association study for response to vaccination in Angus calves1.

BACKGROUND: Bovine respiratory disease complex (BRDC) is one of the most important sources of loss within the beef cattle industry in the USA. Steps have been taken to reduce the incidence of BRDC through vaccination. Despite the effectiveness of vaccines, large proportions of cattle still experience morbidity and mortality. Identification of genomic regions that are associated with variation in response to vaccination would allow for the selection of individuals genetically predisposed to respond to vaccination based on specific markers, while heritability and accuracy estimates would help facilitate genomic selection. This in turn may lead to selection for beef cattle herds that may have lower incidence rate of BRDC after vaccination. This study utilizes an Angus herd of more than 2000 head of cattle to identify these regions of association. RESULTS: Genome wide association studies were performed for viral neutralization antibody level and response to vaccination traits against four different viruses associated with BRDC: bovine viral diarrhea virus 1 and 2 (BVDV1 and BVDV2), bovine respiratory syncytial virus (BRSV), and bovine herpesvirus (BHV1). A total of six 1-Mb windows were associated with greater than 1% of the genetic variance for the analyzed vaccination response traits. Heritabilities ranged from 0.08 to 0.21 and prediction accuracy ranged from 0.01 to 0.33 across 7 different vaccination traits. CONCLUSIONS: Although six 1-Mb windows were identified as associated with 1% or greater genetic variance for viral neutralization antibody level and response to vaccination traits, few genes around these windows could readily be considered candidates. This indicates the need for further functional genomic annotation, as these regions appear to be gene deserts. Traits ranged from lowly to moderately heritable, which indicated the potential for selection of individuals that are genetically pre-disposed to respond to vaccination. The relatively low amount of genetic variance accounted for by any 1-Mb window indicated that viral neutralization antibody level and response to vaccination traits are polygenic in nature. Selection for these traits is possible, but likely to be slow due to the low heritabilities and absence of markers with high genetic variation associated with them.

Comparative efficacy of modified-live and inactivated vaccines in boosting responses to bovine respiratory syncytial virus following neonatal mucosal priming of beef calves.

Bovine respiratory syncytial virus (BRSV) is the leading cause of viral pneumonia in calves, making young passively immune calves candidates for vaccination, and raising issues concerning boosting of neonatally primed responses. To address this, 18, 2-month-old Angus-cross passively immune beef heifer calves that had been primed at birth with a combination viral intranasal vaccine were administered either a parenteral combination vaccine containing modified-live (MLV) BRSV or a similar vaccine containing inactivated BRSV. At 6 months of age, these calves and 2 controls that received only the MLV at 2 months of age were challenged with BRSV via aerosol. Two calves, 1 control, and 1 MLV-boosted, developed severe respiratory disease and required euthanasia; the remaining calves developed no or mild respiratory disease and recovered. Calves that received the inactivated booster had significantly higher arterial oxygen concentrations on Day 7 after challenge and had anamnestic BRSV-specific IgG and neutralizing antibodies after challenge; the MLV-boosted calves did not. These data suggest that adjuvanted inactivated parenteral BRSV vaccines administered at 2 months of age may provide better boosting for neonatally mucosally primed calves.

Efficacité comparée des vaccins vivants modifiés et des vaccins inactivés pour améliorer la réponse au virus respiratoire syncytial bovin après la sensibilisation active néonatale des muqueuses chez les veaux de boucherie. Le virus respiratoire syncytial bovin (VRS) est la cause principale de pneumonie virale chez les veaux, ce qui rend des jeunes veaux à immunité passive des candidats pour la vaccination et soulève des enjeux liés à l'amélioration de la réponse des nouveau-nés sensibilisés. Dans le but d'aborder cette situation, 18 veaux de boucherie de race croisée Angus âgés de 2 mois ayant une immunité passive, qui avaient été sensibilisés activement à la naissance à l'aide d'une combinaison de vaccins intranasaux viraux, ont reçu soit un vaccin combiné parentéral contenant le VRS modifié vivant (VMV) ou un vaccin semblable contenant le VRS inactivé. À l'âge de 6 mois, ces veaux et deux témoins qui avaient reçu seulement le VNV à l'âge de 2 mois, ont été exposés au VRS par voie aérosol. Deux veaux, un témoin et un animal ayant reçu le rappel VMV, ont développé une maladie respiratoire grave et ont dû être euthanasiés; les autres animaux ont développé une maladie respiratoire légère et se sont rétablis ou n'ont manifesté aucun symptôme. Les veaux qui avaient reçu le rappel inactivé affichaient des concentrations d'oxygène significativement supérieures dans le sang artériel le jour 7 après le test et présentaient des anticorps neutralisants et anamnestiques spécifiques aux VRS après le test, contrairement aux veaux ayant reçu le rapport VNV. Ces données suggèrent que les vaccins VRS parentéraux inactivés avec adjuvants administrés à l'âge de 2 mois peuvent offrir une meilleure protection pour les veaux sensibilisés activement à la naissance sur les muqueuses.(Traduit par Isabelle Vallières).

Bovine respiratory syncytial virus-specific IgG-1 in nasal secretions of colostrum-fed neonatal calves.

In order to determine whether nasal secretions of young calves contain passively derived antibodies to bovine respiratory syncytial virus (BRSV) and if there are differences in presence and/or subclass of these antibodies between calves fed different colostrum replacement products, 17 Holstein calves were fed 150 g of IgG in either a sprayed-dried colostrum-based (CR; n = 8) or a plasma-based colostrum replacement product (PR; n = 9) within 6 h of birth. Venous blood and nasal secretions obtained before feeding and at 24 h of age were assayed for total IgG (serum) by radial immunodiffusion and for BRSV-specific total IgG, IgG-1, and IgG-2 by indirect enzyme-linked immunosorbent assay (ELISA). Calves that were fed a CR had higher concentrations of BRSV-specific IgG and IgG-1 in their serum and nasal secretions compared to calves fed product PR; calves fed the PR had higher levels of serum BRSV-specific IgG-2. The only subclass of antibodies detected in nasal secretions was IgG-1. Re-secretion of passive IgG with neutralizing activity, onto the nasal mucosa could contribute to BRSV-associated disease-sparing observed in the laboratory and in the field. Use of PR will result in lower nasal antibodies since IgG-2 is not re-secreted.

IgG-1 spécifique au virus respiratoire syncytial bovin dans les sécrétions nasales des veaux néonataux nourris au colostrum. Afin de déterminer si les sécrétions nasales des jeunes veaux contenaient des anticorps dérivés passivement envers le virus respiratoire syncytial bovin (VRS) et s'il y a des différences dans la présence et/ou la sous-catégorie de ces anticorps entre les veaux nourris avec différents produits de remplacement du colostrum, 17 veaux Holstein ont été nourris avec 150 g d'IgG soit sous forme de produit vaporisé-séché à base de colostrum (CR; n = 8) ou d'un produit de remplacement de colostrum à base de plasma (PR; n = 9) au cours des 6 premières heures après la naissance. Du sang veineux et des sécrétions nasales obtenus avant le nourrissage et à l'âge de 24 h ont été analysés pour obtenir la quantité d'IgG totale (sérum) par immunodiffusion radiale et le total des quantités d'IgG, d'IgG-1 et d'IgG-2 spécifiques au VRS par ELISA indirecte. Les veaux qui avaient été nourris d'un CR avaient des concentrations supérieures d'IgG et dIgG-1 spécifiques au VRS dans leur sérum et les sécrétions nasales comparativement aux veaux nourris de produits PR; les veaux nourris d'un PR avaient des niveaux supérieurs d'IgG-2 sérique spécifique au VRS. La seule sous-catégorie d'anticorps détectée dans les sécrétions nasales était l'IgG-1. La re-secrétion passive d'IgG avec de l'activité neutralisante sur les muqueuses nasales pourrait contribuer à l'immunité associée au VRS observée en laboratoire et sur le terrain. L'usage de PR produira des anticorps nasaux inférieurs vu que l'IgG-2 n'est pas re-secrété.(Traduit par Isabelle Vallières).