THE EFFECT OF NATURAL AND ARTIFICIAL BREEDING USING BULLS INFECTED WITH, OR SEMEN CONTAMINATED WITH, INFECTIOUS BOVINE RHINOTRACHEITIS VIRUS

Ten cows and heifers (Group B) were inoculated into the uterus at oestrus with semen followed by IBR virus for the first insemination and semen alone if a second insemination was necessary. All animals developed infectious pustular vulvo-vaginitis (IPV), and 2 cows conceived to the first and 2 to the second insemination (pregnancy rate of 40 percent requiring 4.5 services per conception). This group was compared with 10 control animals (Group A) which were treated similarly but received tissue culture fluid instead of virus at the first insemination. Group A had a pregnancy rate of 90 percent requiring 1.7 services per conception. Natural mating of 4 bulls with preputial infections due to infectious bovine rhinotracheitis (IBR) virus with 9 susceptible cows and heifers (group D), resulted in the production of lesions of IPV. The IPV infection did not affect their fertility (pregnancy rate of 89 percent requiring 1.4 services per conception) when it was compared to a similar group of females (group C) mated to the same bulls prior to infection with IBR virus (pregnancy rate of 100 percent requiring 1.2 services per conception). The 6 animals in Group B that were not pregnant and returned to oestrus 3 times were found on slaughter to have endometritis, salpingitis and vaginitis. A high incidence, 5 out of 18 (28 percent), of shortened oestrous cycles (less than 18 days) was a feature of the breeding pattern of this group. The undesirable consequences of distributing semen contaminated with IBR virus from artificial insemination centres are apparent.     

Ovarian lesions in heifers exposed to infectious bovine rhinotracheitis virus by non-genital routes on the day after breeding

Twelve heifers were exposed to either a Colorado infectious bovine rhinotracheitis (IBR) virus isolate or an Iowa IBR isolate obtained from a bovine respiratory disease outbreak. All inoculations were made on the day after the heifers had been in estrus and bred by an IBR virus-negative bull. Pairs of heifers were inoculated with each virus isolate intravenously, intramuscularly or exposed by aerosol. The heifers were killed 11-15 days after inoculation and their reproductive tracts and ovaries subjected to virological and pathological study. Virus was isolated from the ovaries of all 4 heifers inoculated intravenously and from 3 of the 4 heifers inoculated intramuscularly, but not from the ovaries of heifers exposed by aerosol. Virus isolations and lesions were, with only 1 exception, confined to the ovary containing the corpus luteum. In ovaries from which IBR virus was isolated, lesions in the corpus luteum ranged from focal necrosis and infiltration of mononuclear cells to diffuse hemorrhage and necrosis. Most of these ovaries also had necrotic follicles and a diffuse mononuclear cell accumulation in the stroma. Lesions were not found in ovaries from which IBR virus was not isolated. It was concluded that lesions are readily induced in the ovaries of post-estrus heifers as a result of hematogenous spread of IBR virus and suggest that the differences in lesion development observed with the 3 routes are related to whether or not a viremia occurred.     

Neural changes in recurrent infection of infectious bovine rhinotracheitis virus in calves treated with dexamethasone.

Recurrent infection by infectious bovine rhinotracheitis (IBR) virus was induced in calves by dexamethasone (DM) treatment (given 5 days) at 5 months after primary infection. The virus appeared in nasal secretions of the calves on the 4th day after initiation of DM treatment and continued until the 9th day. The calves were killed on the 1st, 3rd, 4th, 5th, 6th, 7th, 8th, 10th, and 11th days after DM treatment was started for examination by histopathologic and immunofluorescent antibody techniques. The most significant neural change was trigeminal ganglionitis with neuronophagia, which was observed from the 3rd to the 11th day. Significantly, the extent of changes in the trigeminal ganglion and medulla oblongata corresponded to the amount of DM treatment administered. The IBR virus antigen was first observed in the trigeminal ganglion cells, and thereafter, it was detected in the Schwann cells, satellite cells, neuroglia cells, and nasal mucosa until the 10th day. These observations indicate that the IBR virus is capalbe of producing a persistent infection in the trigeminal ganglion and that trigeminal ganglionitis may be a characteristic lesion for inducing the reactivation of lagent IBR virus.     

Diagnosis of infectious bovine rhinotracheitis by direct immunofluorescence

Summary

Clinical signs, virus excretion and immunofluorescence in nasal smears were studied in nine susceptible steers during a two week period, following intranasal expo‐sure with IBR‐virus. All animals responded with fever (ay. 3.9 days) and nasal discharge. IBR‐virus was isolated from nasal swabs from 1 to 11 days after exposure (ay. 10 days), whereas fluorescence in nasal smears was observed from the second till the seventh day after infection (ay. 5.5 days).

Fluorescence was most distinct 3 to 5 days after infection, which coincided with the period of fever and a serous nasal discharge. Smears from animals with a mucopurulent or slightly haemorrhagic nasal discharge were nearly always negative. For a reliable diagnosis on live animals by immunofluorescence, it is necessary to take nasal smears from several healthy looking animals with fever and a slight, preferably serous discharge. Air dried smears should be fixed in acetone within 24 hours. Seven yearlings were autopsied 3 to 11 days after intranasal exposure and subjected to a detailed investigation by the cryostat‐immunofluorescence technique (IFT). The tonsils of all animals were positive, followed in declining frequency by the larynx, namharynx, nasal mucosa, and pharyngeal mucosa. Besides the organs already mentioned, fluorescence was often observed in the lungs and tracheal mucosa of animals that had suffered a fatal infection of IBR in the field. The tonsils should be regarded as the organ of choice. Fluorescent foci were localized in the epithelial lining of the tonsillar crypts and in the surface epithelium of the mucosae. The direct IFT on nasal smears of suspected animals and on cryostat sections of tissues collected at autopsy offers veterinary laboratories with no facilities for tissue culture a possibility of a rapid and reliable diagnosis of IBR infections.     

Recrudescence of infectious bovine rhinotracheitis virus and associated neural changes in calves treated with dexamethasone

Reactivation of infection bovine rhinotracheitis (IBR) virus in calves administered dexamethasone (DM) was studied in 2 experiments. At 2, 3, 5, 15, or 30 months after inoculation of the Los Angeles strain of IBR virus, IV injections of DM were given for 5 consecutive days to induce a recurrent infection (experiment 1). Three months after the 1st treatment, a 2nd recurrent infection was induced, using DM with the same doses as used in experiment 1. The virus was excreted from nasal secretions from the 4th to the 10th day after initial treatment with DM, and from the 6th to the 9th day after the 2nd treatment. On pathologic examination, trigeminal ganglionitis, consisting of many proliferated microglia and inflammatory cells, was observed in all DM-treated calves. Moreover, degeneration of the ganglion cells and neuronophagia were prominent features in the calves after the 2nd recurrent infection. These observations indicated that the trigeminal ganglion may be one of the latent sites of IBR virus in calves after intranasal infection and that calves can develop a recrudescent infection after DM treatment several times during their lifetime.     

Excretion and reexcretion of thermosensitive and wild-type strains of infectious bovine rhinotracheitis virus after co-infection or two successive infections

Twelve cattle were divided into 2 groups. The first was intranasally co-infected with 2 strains of infectious bovine rhinotracheitis virus (Bovine herpesvirus 1; BHV 1): the thermosensitive vaccine strain IBR/ts RLB106 and a Belgian field isolate IBR/Cu5. Reactivation of BHV 1 was induced by dexamethasone treatment 2 months later and again 5 months later for 3 animals that only reexcreted small quantities of virus during the first dexamethasone treatment. The second group was intranasally infected with IBR/Cu5. Two months later, an attempt to reinfect this group with IBR/ts RLB106 failed. Four months after the primary infection, these cattle were treated with dexamethasone. Except after reinfection and at the beginning or the end of the (re)excretion periods, excreted and reexcreted viruses replicated at 35, 37 and 40 degrees C, indicating the presence of the wild-type virus. Only one isolate, out of 116 cloned from the nasal exudates collected during the excretion and reexcretion periods, expressed the thermosensitive phenotype. This isolate was characterized by its mean plaque size as the IBR/ts RLB106 strain. The epizootiological significance of these findings is discussed, with emphasis on the weak spreading capacity of the ts vaccine strain and the possibility of emergence of recombinant viruses.     

Reproductive safety of vaccination with Vista 5 L5 SQ near breeding time as determined by the effect on conception rates

Replacement heifers (N=799; 10 to 13 months of age) were vaccinated with Vista 5 L5 SQ (Intervet; a reconstituted vaccine-bacterin product containing modified-live cultures of infectious bovine rhinotracheitis [IBR] virus, bovine viral diarrhea virus [BVDV; types 1 and 2], parainfluenza-3 virus, and bovine respiratory syncytial virus and inactivated cultures of Leptospira serovars canicola, grippotyphosa, hardjo, icterohaemorrhagiae, and pomona with a proprietary adjuvant) at either 40 plus/minus 5 days (control; n=399) or 3 days (test; n=400) before peak breeding day. By 40 plus/minus 5 days before peak breeding day, heifers in both groups had greater average titers to IBR, BVDV types 1 and 2, and four of the five Leptospira antigens assessed as compared with prevaccination titers on day -90 plus/minus 25 days. Conception rates were not affected by treatment. This study suggests that conception rates will not differ between heifers vaccinated with Vista 5 L5 SQ 3 days before breeding and those vaccinated approximately 40 days before breeding.     

Infectious bovine rhinotracheitis virus: studies on the venereal carrier status in range cattle.

Genital samples collected at autopsy from 193 beef cows representing 22 different herds in Northern Australia yielded only one isolate of infectious bovine rhinotracheitis (IBR) virus. Serological evidence showed 59 per cent of similar cows had prior infection with this virus and of 19 sero-positive cows tested, 11 (58-2 per cent) shed detectable IBR virus from the vagina after treatment with corticosteroids. Transitory lesions of the vagina and vulva developed in five of the treated cows. Twenty-six (65 per cent) of 40 sero-positive bulls shed detectable IBR virus into the prepuce after corticosteroid treatment. Except for one bull, virus was not isolated after corticosteroid treatment of sero-negative animals. IBR virus and mucosal disease (MD) virus were not isolated from nasal swabs before or after corticosteroid administration. No correlation was observed between initial circulating antiboyd titre and virus excretion after treatment. There were no significant changes in levels of serum antibody during the virus excretion period.     

Duration of protection of calves against rhinovirus challenge exposure by infectious bovine rhinotracheitis virus-induced interferon in nasal secretions

Six calves inoculated intranasally with a vaccinal strain of infectious bovine rhinotracheitis (IBR) virus and 6 control calves were given a placebo. All calves were subsequently challenge exposed (by aerosol) with rhinovirus--3 of the calves from each group at 2 days after they were inoculated with IBR virus or with placebo and the remaining calves at 6 days. Nasal excretion of viruses, interferon (IFN) concentrations in nasal secretions (NS), and neutralizing antibody in sera and NS were determined. All calves given the vaccinal IBR virus subsequently had IFN in their NS. Interferon was detected as early as 1 day, reached maximal titers at 2 to 4 days, and persisted in individual calves for 5 to 10 days after inoculation. Rhinovirus shedding was not detected from IBR virus-inoculated calves whose NS contained both rhinovirus antibody and IFN at the time of challenge exposure; such calves were protected at either 2 or 6 days after IBR virus inoculation. The outcome of rhinovirus challenge exposure of calves whose NS contained IFN, but not rhinovirus antibody, varied with the day of challenge exposure. Rhinovirus excretion was detected from 2 of these calves challenge exposed 2 days after IBR virus inoculation, but was not detected from a calf challenge exposed 6 days after inoculation. However, while IFN was present in NS from the former 2 calves, rhinovirus shedding was markedly reduced as compared with that from control calves without IFN or NS antibody at the time of challenge exposure. Consistent relationship was not observed between the rhinovirus neutralizing antibody titer of calves' sera and NS. The antibody titer of NS more closely correlated with protective immunity to rhinovirus infection than did the serum antibody titer.     

Isolation of infectious bovine rhinotracheitis virus from a latently infected feral pig

Infectious bovine rhinotracheitis (IBR) virus was isolated from the trigeminal ganglion of a feral pig after dexamethasone treatment. Three pigs inoculated intranasally with the IBR virus did not respond clinically or serologically. The virus was re-isolated from tonsillar swabs from two animals on Post-Infection Day (PID) 3.     

Active and Passive Immunity to Bovine Viral Respiratory Diseases in Beef Calves After Shipment

Fifteen steers were vaccinated after shipment with a modified live virus vaccine containing infectious bovine rhinotracheitis (IBR), bovine virus diarrhea (BVD), and bovine myxovirus parainfluenza-3 (PI3), and 16 unvaccinated steers were kept as controls. Geometric mean titers one month after vaccination were highest to BVD, followed by PI3 and IBR. Weight gains were higher during 30 days after vaccination in the controls. One case of acute respiratory disease developed in one vaccinated calf. Revaccination 79 days after the first dose increased antibody to PI3 and BVD virus but not IBR. In a second trial, no clinical respiratory disease developed after shipment of 13 heifers that received an antibacterial-antiviral antiserum or in the 12 controls. Weight gains 30 days after shipment were identical in both groups.     

Attempts to reactivate bovid herpesvirus-2 in experimentally infected calves

A study was carried out to determine whether bovid herpesvirus-2 (BHV-2) is able to induce a recurrent infection in experimentally infected calves. Twelve calves infected with the virus were treated with dexamethasone (DMS) beginning 69 days after the infection, ie, several weeks after the animals had recovered from the disease and were negative for BHV-2. The stress induced by DMS treatment failed to reactivate the clinical condition or to induce shedding of BHV-2. However, treatment with DMS reactivated a latent infectious bovine rhinotracheitis (IBR) virus infection in all calves previously inoculated with BHV-2, and also in 2 noninoculated controls. The reactivation of IBR virus occurred without any clinical evidence of the disease, but the virus was isolated from nasal and pharyngeal swabbings and from the organs. A proliferative ganglionitis of the trigeminal ganglion was also observed. Because of the interference by IBR virus, this study did not resolve the question as to whether BHV-2 can induce a recurrent infection.     

Eradication of infectious bovine rhinotracheitis in Switzerland: review and prospects

Ten years after the first outbreak of infectious bovine rhinotracheitis (IBR) in Swiss dairy cows, the national cattle herd is almost free from infection with IBR virus (bovine herpesvirus 1, BHV 1). The national programme for the eradication of IBR was divided into four phases: (1) Prevention of transmission of the infection by restrictions on trade of bovines and assessment of the prevalence of cattle with antibodies to BHV 1. (2) Slaughtering animals with antibodies to BHV 1 in order to eradicate BHV 1 from breeding herds. (3) Detection and eradication of further BHV 1 reservoirs (e.g. fattening cattle). (4) Monitoring programme and legal actions in order to maintain the favourable situation. Approximately 50,000 animals were slaughtered in the course of the eradication of IBR. The total costs amounted to approximately SFr. 110,000,000 over 10 years. The costs for maintaining the situation are estimated at approximately SFr. 5,000,000 per annum.     

Experimental Infection of Calves with Bovine Viral Diarrhoea Virus Type-2 (BVDV-2) Isolated from a Contaminated Vaccine

A non-cytopathic strain of BVDV-2 was isolated from a batch of live infectious bovine rhinotracheitis (IBR) vaccine, and inoculated intranasally into four 3-month-old calves. Severe signs of disease developed by days 4 and 6 in three of the calves, free of BVDV and antibodies to BVDV, that had been exposed to the virus. These calves survived the acute phase of the infection and progressively recovered. BVDV was consistently isolated, or the respective viral RNA was detected, in the buffy coats from blood samples collected starting from days 2 or 4 up to days 11 or 14 after the experimental infection. Viral RNA was also detected in sera from these infected calves until the presence in the serum of virus neutralizing antibodies was demonstrated. By contrast, the only calf having pre-existing neutralizing antibodies to BVDV at the start of the study was protected from the disease. No virus was detected at any time after experimental inoculation of this calf. Genomic characterization of the BVDV-2 isolated in cell cultures, or detected in sera from the experimentally infected animals, revealed 100% homology in the nucleotide sequence with the BVDV-2 detected as a contaminant of the live IBR virus vaccine. These findings provided evidence of the infective nature of the contaminant BVDV-2 and of its potential to generate disease outbreaks when inoculated into susceptible animals.     

A Controlled Field Study Using Live Virus Vaccines and an Antiserum in a Preconditioning Program

Thirty-five vaccinates and 29 control beef calves from five farms were studied. Vaccinates in group 1 received a modified live virus vaccine against infectious bovine rhinotracheitis (IBR) and bovine virus diarrhea (BVD) 30 days after shipment; vaccinates in groups 2, 3 and 4 received live virus vaccines agains IBR and bovine parainfluenza 3 (PI3) seven to 17 days before shipment. Half of group 5 were given bovine origin antiserum containing antibodies against IBR, BVD and PI3. Three weeks later, the animals that had received serum were given a live modified vaccine containing IBR, BVD and PI3. In group 1, WBC counts were lower in the vaccinates than in the controls for two weeks after vaccination. WBC counts in groups 3 and 4 were higher in vaccinates than in controls after addition to the feedlot. Seroconversions to BVD virus occured in all groups. Clinical disease apparently due to BVD affected one vaccinated calf in group 2 and eight calves in group 5. Combined weight gains were significantly higher in three groups of calves vaccinated before shipment compared to unvaccinated control animals after addition to the feedlot. Vaccination with IBR and PI3 live virus vaccines should be given at least 17 days before shipment to feedlots containing infected cattle. Antiserum containing antibodies against the three viruses showed no apparent advantage in preventing clinical respiratory disease over control calves not receiving the serum.     

Diagnosis of infectious bovine rhinotracheitis by direct immunofluorescence

Summary Clinical signs, virus excretion and immunofluorescence in nasal smears were studied in nine susceptible steers during a two week period, following intranasal expo-sure with IBR-virus. All animals responded with fever (ay. 3.9 days) and nasal discharge. IBR-virus was isolated from nasal swabs from 1 to 11 days after exposure (ay. 10 days), whereas fluorescence in nasal smears was observed from the second till the seventh day after infection (ay. 5.5 days). Fluorescence was most distinct 3 to 5 days after infection, which coincided with the period of fever and a serous nasal discharge. Smears from animals with a mucopurulent or slightly haemorrhagic nasal discharge were nearly always negative. For a reliable diagnosis on live animals by immunofluorescence, it is necessary to take nasal smears from several healthy looking animals with fever and a slight, preferably serous discharge. Air dried smears should be fixed in acetone within 24 hours. Seven yearlings were autopsied 3 to 11 days after intranasal exposure and subjected to a detailed investigation by the cryostat-immunofluorescence technique (IFT). The tonsils of all animals were positive, followed in declining frequency by the larynx, namharynx, nasal mucosa, and pharyngeal mucosa. Besides the organs already mentioned, fluorescence was often observed in the lungs and tracheal mucosa of animals that had suffered a fatal infection of IBR in the field. The tonsils should be regarded as the organ of choice. Fluorescent foci were localized in the epithelial lining of the tonsillar crypts and in the surface epithelium of the mucosae. The direct IFT on nasal smears of suspected animals and on cryostat sections of tissues collected at autopsy offers veterinary laboratories with no facilities for tissue culture a possibility of a rapid and reliable diagnosis of IBR infections.     

Effects in calves of mixed infections with bovine viral diarrhea virus and several other bovine viruses

The objective of this study was to verify whether a mixed infection in calves with bovine viral diarrhea virus (BVDV) and other bovine viruses, such as bovid herpesvirus-4 (BHV-4), parainfluenza-3 (PI-3) and infectious bovine rhinotracheitis (IBR) virus, would influence the pathogenesis of the BVDV infection sufficiently to result in the typical form of mucosal disease being produced.

Accordingly, two experiments were undertaken. In one experiment calves were first infected with BVDV and subsequently with BHV-4 and IBR virus, respectively. The second experiment consisted in a simultaneous infection of calves with BVDV and PI-3 virus or BVDV and IBR virus.

From the first experiment it seems that BVDV infection can be reactivated in calves by BHV-4 and IBR virus. Evidence of this is that BVDV, at least the cytopathic (CP) strain, was recovered from calves following superinfection. Moreover, following such superinfection the calves showed signs which could most likely be ascribed to the pathogenetic activity of BVDV. Superinfection, especially by IBR virus, created a more severe clinical response in calves that were initially infected with CP BVDV, than in those previously given the non-cytopathic (NCP) biotype of the virus. Simultaneous infection with PI-3 virus did not seem to modify to any significant extent the pathogenesis of the experimentally induced BVDV infection whereas a severe clinical response was observed in calves when simultaneous infection was made with BVDV and IBR virus.     

Validation of betapropiolactone (BPL) as an inactivant for infectious bovine rhinotracheitis (IBR) virus

Infectious bovine rhinotracheitis (IBR) virus causes vulvovaginitis, abortion and respiratory disease in cows and heifers. Betapropiolactone (BPL) is a disinfectant, effective against bacteria, fungi and viruses. It is also used to prepare inactivated vaccines because it destroys the nucleic acid core of viruses but does not damage the capsid. For the validation of BPL when used as an inactivant, it is more important to assure the quality of inactivating agent and the validity of the inactivation process. In the present study, the inactivation kinetics of IBR virus was determined with different concentration of BPL (1:250, 1:500, 1:1000, 1:1500, 1:2000 and 1:2500) at 4 and 37 °C. The result indicated that the BPL at 4 °C was able to inactivate the IBR virus within 4, 5 and 12 h with the concentration of 1:250, 1:500 and 1:1000, respectively. BPL at 37 °C was able to inactivate virus within 30 min with the concentration of 1:250. BPL with the concentration of 1:500 and 1:1000 were able to inactivate the virus within 120 min at 37 °C. Based on the kinetic study seven formulations were prepared and a sero conversion study of IBR inactivated vaccine was carried out. Serological response in animals to different formulations did not differ significantly (P > 0.05).     

Evidence for defective neutrophil function in lungs of calves exposed to infectious bovine rhinotracheitis virus

Calves were exposed to an aerosol of infectious bovine rhinotracheitis (IBR) virus followed five days later by an aerosol of Pasteurella haemolytica. The animals were subjected to bronchoalveolar lavage before IBR and four days after, and again at 0, 4, 24, and 48 hours following Pasteurella haemolytica challenge. The results of these experiments suggest that neutrophil infiltration into the lung, in response to the presence of the bacteria was delayed thereby allowing the bacteria to become established in the lung. Neutrophils in infected animals displayed little random migration in vitro and did not respond to a chemotactic stimulus. It was also found that alveolar macrophages from virus-infected animals were not able to produce neutrophil chemotactic factors. These data suggest that the decrease in neutrophil chemotaxis and the lack of chemotactic factor production by the alveolar macrophage following infection with infectious bovine rhinotracheitis virus may predispose infected cattle to a secondary bacterial infection.     

Investigation of causative agents of bovine respiratory tract disease in a beef cow-calf herd with an early weaning program.

Serum samples were collected from early weaned fall calves shortly after the onset of respiratory tract disease. Antibody titers to infectious bovine rhinotracheitis (IBR) virus, parainfluenza type 3 (PI-3) virus, bovine viral diarrhea (BVD) virus, bovine adenovirus type 3 (BAV-3), and bovine respiratory syncytial virus (BRSV) were determined on paired (acute and convalescent) serums. Seroconversion rate (a fourfold or greater rise in antibody titer) for IBR virus was 4.3%, PI-3 virus--16.3%, BVD virus--9.6%, and BAV-3--2.2%. Seroconversion for BRSV was 45.4%. An increased rate of seroconversion for IBR, PI-3, and BVD viruses and BAV-3 was observed in the presence of BRSV seroconversion. These results suggest that BRSV may facilitate infection by other viruses. Results of virus isolation procedures from these calves were negative.