Genetic comparison of ovine and bovine pestiviruses

Viral RNA oligonucleotide fingerprinting was used to compare genetic relationship among pestiviruses originating from ovine or bovine host species. Ovine pestiviruses, including reference border disease virus and 2 border disease isolates originating from natural pestivirus infections of sheep, appeared to have a more distant genetic relationship among themselves than with certain bovine pestiviruses. A closer genetic relatedness was evident between border disease virus and 3 noncytopathic bovine pestiviruses, including Draper bovine viral diarrhea virus (BVDV), a BVDV isolate that originated from aborted bovine fetuses, and a virus that was isolated from the serum of a calf that had a chronic BVDV infection. Four noncytopathic bovine viruses, including Draper BVDV and 3 field isolates, were closely related. Reference Oregon C24V BVDV, a cytopathic virus, was closely related to only 1 of the 7 noncytopathic viruses in this study.     

Identification of hog cholera viral isolates by use of monoclonal antibodies to pestiviruses

A collection of 90 field isolates of hog cholera virus (HCV) was used to test the specificity of four hybridoma cell lines secreting monoclonal antibodies against pestiviruses. Reaction of virus isolates and monoclonal antibodies was controlled by an indirect immunofluorescence assay (IFA). Two monoclonal antibodies which had been generated against HC virus strain "Alfort 187" were reactive only with HCV field isolates and an HCV reference strain but not with bovine viral diarrhoea virus (BVDV) reference strains. Two other monoclonal antibodies (generated against BVDV, strain NADL) reacted only with BVDV reference strains but not with HCV field isolates, although with 3 of these strains focal reactions involving only a few cells were detected. The ability to discriminate between both viruses is a diagnostic need which may be fulfilled by these monoclonal antibodies.     

Differential diagnosis of classical swine fever and border disease: seroepidemiological investigation of a pestivirus infection on a mixed sheep and swine farm.]

During recent years neutralizing antibodies against Border Disease Virus (BDV) were found repeatedly in German pig herds. Consequently there was a demand for a differential diagnostic system. A permanent sheep cell line and BDV reference strain Moredun were chosen and were applied in a could be used case study. A pestivirus could be isolated from piglets on a mixed farm and was characterised as 'non-Classical Swine Fever' (CSF) by using monoclonal antibodies. Due to a CSF suspicion the pig herd was destroyed immediately. Serum samples of sheep from the same farm were used for further characterisation of the new virus isolate. A neutralization test of the sheep sera was performed against different pestiviruses and the new isolate. Neutralizing antibody titres against the new virus pig isolate were significantly higher than against all other pestiviruses. BDV strain Moredun recognised the antibodies clearly, whereas CSF viral strain Alfort 187 and several isolates of bovine viral diarrhoea virus (BVDV) strains scored the lowest cross reaction.     

Variation in pestivirus growth in testicle primary cell culture is more dependent on the individual cell donor than cattle breed

The causes of bovine respiratory disease complex (BRDC) are multifactorial and include infection with both viral and bacterial pathogens. Host factors are also involved as different breeds of cattle appear to have different susceptibilities to BRDC. Infection with bovine pestiviruses, including bovine viral diarrhea virus 1 (BVDV1), BVDV2 and ‘HoBi’-like viruses, is linked to the development of BRDC. The aim of the present study was to compare the growth of different bovine pestiviruses in primary testicle cell cultures obtained from taurine, indicine and mixed taurine and indicine cattle breeds. Primary cells strains, derived from testicular tissue, were generated from three animals from each breed. Bovine pestivirus strains used were from BVDV-1a, BVDV-1b, BVDV-2a and ‘HoBi’-like virus. Growth was compared by determining virus titers after one passage in primary cells. All tests were run in triplicate. Virus titers were determined by endpoint dilution and RT-qPCR. Statistical analysis was performed using one way analysis of variance (ANOVA) followed by the Tukey’s Multiple Comparison Test (P?0.05). Significant differences in virus growth did not correlate with cattle breed. However, significant differences were observed between cells derived from different individuals regardless of breed. Variation in the replication of virus in primary cell strains may reflect a genetic predisposition that favors virus replication.     

Virus neutralising antibodies against 22 bovine viral diarrhoea virus isolates in vaccinated calves.

Seven of nine colostrum deprived calves, free from bovine viral diarrhoea virus (BVDV), were vaccinated with a commercially available vaccine containing two inactivated strains of BVDV, an inactivated strain of bovine herpesvirus-1 and modified-live strains of bovine respiratory syncytial virus and para-influenza-3 virus. The two other calves were kept as controls. The virus neutralising (VN) antibodies induced by vaccination were tested against 22 antigenically diverse BVDV isolates, including reference strains and field isolates, both cytopathic and non-cytopathic, as well as genotypes I and II. The strains were isolated in Belgium, France, Germany, the United Kingdom and the USA. While there were variations in the VN titres of the individual calves against all the strains, serum from the seven animals neutralised 20 or more of the strains tested. From the results, it can be concluded that the vaccine can stimulate the production of VN antibodies capable of neutralising a wide range of European and American isolates of BVDV, including genotypes I and II.     

The genetic basis for cytopathogenicity of pestiviruses

Two biotypes of pestiviruses, cytopathogenic (cp) and noncp viruses, can be distinguished by their effects on tissue culture cells. Identification of cp bovine viral diarrhea virus (BVDV) has been frequently reported since antigenically closely related noncp and cp BVDV can be isolated from cattle with fatal mucosal disease (MD) and are called a virus pair. In contrast to the BVDV system, only few cp border disease virus (BDV) and cp classical swine fever virus (CSFV) strains have been described. Serological analyses and sequence comparison studies showed that cp pestiviruses arise from noncp viruses by mutation. Elaborate studies during the last 10 years revealed that in most cases RNA recombination is responsible for the generation of the cp viruses. Recent results showed a second way for the development of a cp pestivirus which is based on the introduction of a set of point mutations within the NS2 gene.     

Kinetics of single and dual infection of calves with an Asian atypical bovine pestivirus and a highly virulent strain of bovine viral diarrhoea virus 1

Atypical bovine pestiviruses related to bovine viral diarrhoea virus (BVDV) have recently been detected in cattle from South America, Asia and Europe. The purpose of this study was to compare the clinical and virological aspects of dual infection with BVDV-1 (Horton 916) and an Asian atypical bovine pestivirus (Th/04_KhonKaen) in na?ve calves, in comparison to single infections. Milder clinical signs were observed in the animals infected with single Th/04_KhonKaen strain. Leukocytopenia and lymphocytopenia were observed in all infected groups at a similar level which correlated with the onset of viraemia. Co-infection with both viruses led to prolonged fever in comparison to single strain inoculated groups and simultaneous replication of concurrent viruses in blood and in the upper respiratory tract. Following the infections all the calves seroconverted against homologous strains. Atypical pestiviruses pose a serious threat to livestock health and BVDV eradication, since they may have the potential to be widely spread in cattle populations without being detected and differentiated from other BVDV infections.     

Genetic typing of pestiviruses from New Zealand

AIM: To investigate the genetic type of 20 pestiviruses collected from New Zealand over the period 1967-97. METHODS: The pestiviruses were genetically typed by the sequencing of polymerase chain reaction (PCR) products. The primers selected were from the 5'-untranslated region (5'-UTR) of the pestivirus genome and consistently amplified a 288 bp fragment from all samples tested. RESULTS: Sequencing and phylogenetic analysis of PCR products revealed that all samples obtained from cattle represented bovine viral diarrhoea (BVDV) type I. Two sheep isolates were characterised as border disease virus (BDV). A pestivirus isolated from foetal calf serum of USA origin was typed as BVDV type II. CONCLUSIONS: The findings show that the evolution of pestiviruses in New Zealand has been similar to Europe and North America, indicating the occurrence of a conservative phylogenetic branch of BVDV type I in cattle and the presence of BDV in the sheep population.     

Practical significance of heterogeneity among BVDV strains: impact of biotype and genotype on U.S. control programs

In the early 1990s research groups in North America noted that a newly recognized severe acute form of bovine viral diarrhea virus infection, referred to as hemorrhagic syndrome or severe acute BVDV (SA BVDV), was associated with a genetically distinct subgroup of BVDV strains. This new subgroup was named BVDV genotype 2 or BVDV2. All BVDV strains previously characterized in the literature belonged to a separate genotype, BVDV1. However, not all strains identified as BVDV2 were associated with severe acute infections. If I did this deletion, I did not mean to do it. I think it was already here, though. I see there are some other big edits that I did not do; fine. Hollis subsequent surveys of BVDV strains isolated from clinical submissions to diagnostic laboratories and contaminated fetal calf serum suggested that the ratio of BVDV2 to BVDV1 strains in the U.S. approached 50%. Further, while antigenic cross reactivity is seen between BVDV1 and BVDV2 strains, a log or more difference is typically observed in titers against viruses from different genotypes. These observations prompted vaccine manufacturers in North America to produce vaccines against BVDV that contained antigens from both BVDV1 and BVDV2 strains. Under experimental conditions, these new vaccines offered improved protection against type 2 strains, however field data are still insufficient to assess their efficacy in practice. The BVDV genotypes may also be segregated into subgenotypes. Two subgenotypes of both BVDV1 (BVDV1a and BVDV1b) and BVDV2 (BVDV2a and BVDV2b) have been reported in North American. BVDV2a predominates with BVDV2b isolation a rare event. In contrast, BVDV1a and BVDV1b are both commonly isolated. Antigenic differences observed between strains from the BVDV1a and BVDV1b subgenotypes have led to the suggestion that protection may be improved by inclusion of strains from both BVDV1a and BVDV1b in vaccines in addition to BVDV2. The cost to benefit ratio of this proposal is currently a matter of debate.     

Transmission of Bovine viral diarrhea virus 1b to susceptible and vaccinated calves by exposure to persistently infected calves

Bovine viral diarrhea virus (BVDV) persistently infected (PI) calves represent significant sources of infection to susceptible cattle. The objectives of this study were to determine if PI calves transmitted infection to vaccinated and unvaccinated calves, to determine if BVDV vaccine strains could be differentiated from the PI field strains by subtyping molecular techniques, and if there were different rates of recovery from peripheral blood leukocytes (PBL) versus serums for acutely infected calves. Calves PI with BVDV1b were placed in pens with nonvaccinated and vaccinated calves for 35 d. Peripheral blood leukocytes, serums, and nasal swabs were collected for viral isolation and serology. In addition, transmission of Bovine herpes virus 1 (BHV-1), Parainfluenza-3 virus (PI-3V), and Bovine respiratory syncytial virus (BRSV) was monitored during the 35 d observation period. Bovine viral diarrhea virus subtype 1b was transmitted to both vaccinated and nonvaccinated calves, including BVDV1b seronegative and seropositive calves, after exposure to PI calves. There was evidence of transmission by viral isolation from PBL, nasal swabs, or both, and seroconversions to BVDV1b. For the unvaccinated calves, 83.2% seroconverted to BVDV1b. The high level of transmission by PI calves is illustrated by seroconversion rates of nonvaccinated calves in individual pens: 70% to 100% seroconversion to the BVDV1b. Bovine viral diarrhea virus was isolated from 45 out of 202 calves in this study. These included BVDV1b in ranch and order buyer (OB) calves, plus BVDV strains identified as vaccinal strains that were in modified live virus (MLV) vaccines given to half the OB calves 3 d prior to the study. The BVDV1b isolates in exposed calves were detected between collection days 7 and 21 after exposure to PI calves. Bovine viral diarrhea virus was recovered more frequently from PBL than serum in acutely infected calves. Bovine viral diarrhea virus was also isolated from the lungs of 2 of 7 calves that were dying with pulmonary lesions. Two of the calves dying with pneumonic lesions in the study had been BVDV1b viremic prior to death. Bovine viral diarrhea virus 1b was isolated from both calves that received the killed or MLV vaccines. There were cytopathic (CP) strains isolated from MLV vaccinated calves during the same time frame as the BVDV1b isolations. These viruses were typed by polymerase chain reaction (PCR) and genetic sequencing, and most CP were confirmed as vaccinal origin. A BVDV2 NCP strain was found in only 1 OB calf, on multiple collections, and the calf seroconverted to BVDV2. This virus was not identical to the BVDV2 CP 296 vaccine strain. The use of subtyping is required to differentiate vaccinal strains from the field strains. This study detected 2 different vaccine strains, the BVDV1b in PI calves and infected contact calves, and a heterologous BVDV2 subtype brought in as an acutely infected calf. The MLV vaccination, with BVDV1a and BVDV2 components, administered 3 d prior to exposure to PI calves did not protect 100% against BVDV1b viremias or nasal shedding. There were other agents associated with the bovine respiratory disease signs and lesions in this study including Mannheimia haemolytica, Mycoplasma spp., PI-3V, BRSV, and BHV-1.     

Serological and molecular diagnosis of bovine viral diarrhoea virus and evidence of other viral infections in dairy calves with respiratory disease in Venezuela

An investigation based on 2 studies was carried out to assess the involvement of bovine virus diarrhoea virus (BVDV), bovine herpesvirus type 1 (BHV-1), and bovine respiratory syncytial virus (BRSV) in calf respiratory disease in dairy farms in Venezuela. In the first study, 8 farms were selected and paired serum samples from 42 calves with respiratory disease were tested by ELISA for antibodies to the 3 viruses. Seroconversion to BVDV, BHV-1, and BRSV was found to 5, 2, and 6 farms out of the 8, respectively. The proportion of calves that showed seroconversion to BVDV, BHV-1, and BRSV were 19%, 14%, and 26%, respectively. In the second study, another farm having previous serological evidence of BVDV infection was selected. The decline of maternal antibodies against BVDV was monitored in 20 calves and the half-life of maternal antibodies was 34 +/- 12 days presumably indicating an early natural infection with BVDV. Furthermore, sera free of BVDV antibodies that were collected in studies 1 and 2 and were assayed for the presence of BVDV by nested RT-PCR. Two BVDV strains were detected and compared to those of ruminant and porcine pestiviruses. Both strains were assigned to subgroup Ib of type I BVDV. This investigation provides information on BVDV genotypes circulating in Venezuela and may contribute to the establishment of official control programmes against the viruses studied.     

Nucleotide sequence homology to bovine viral diarrhea virus 2 (BVDV 2) in the 5′ untranslated region of BVDVs from cattle with mucosal disease or persistent infection in Japan

Cytopathogenic and non-cytopathogenic bovine viral diarrhea viruses (BVDVs) were isolated from cattle with mucosal disease or persistent infection in Japan. These isolates were compared for antigenic properties by cross-neutralization tests with Japanese reference strains of BVDV belonging to classical type 1. Significantly low cross-reactivity to reference strains was noted, indicating the viruses to possibly represent a new serotype in Japan. Thus, to determine the genotype of the isolates, nucleotide sequences of the 5′ untranslated region were determined and compared with those of previously reported BVDV 1 and 2. The isolates were clearly shown to belong to BVDV 2, not to BVDV 1.     

Genetic typing of bovine viral diarrhea virus isolates from Argentina

Genetic typing of 29 Bovine Viral Diarrhea Virus (BVDV) isolates from Argentina was carried out by sequencing 245 nucleotides of the RT-PCR products of the 5'-UTR region. Sequence analysis shows that these Argentinean BVDV include types 1 and 2. The majority (26/29) of the isolates are type 1, which comprises subtypes 1a and 1b, together with an additional subgroup within subtype 1a. This subgroup is close to the South African subgroup Ic of 1a viruses, and to the deer pestivirus strain "Deer". The three type 2 BVDV were isolated from fetal tissues or serum during the 7-8 years before a clinical outbreak in Argentina had been reported. Only inactivated vaccines are used in bovines of the country, thus the analysed viruses are authentic field strains. The long term circulation of type 2 BVDV (situation similar to that of North America before the epidemic of 1993), and the existence of viral populations which differ from the reference strains commonly used in vaccine elaboration should be considered by manufacturers of diagnostic reagents and vaccines.     

Virus neutralizing antibodies against a panel of 18 BVDV isolates in calves vaccinated with Rispoval RS-BVD

Seven of nine colostrum-deprived calves, free from infection with bovine virus diarrhoea virus (BVDV), were vaccinated with Rispoval RS-BVD on two occasions, 21 days apart, while the other two were kept as BVDV infection controls. The virus neutralizing (VN) serum antibodies induced by vaccination were tested for their ability to neutralize 18 European BVDV isolates, including laboratory reference strains and recent field isolates, both cytopathic and non-cytopathic biotypes as well as genotypes I and II. The strains were isolated in Belgium, France, Germany and the United Kingdom. While there were large variations in the vaccine-induced VN titres of the individual calves against all the strains, e.g. the titres against Osloss NCP, the European reference strain ranged from 1.7 to 6.7 (1:log2), serum from each animal was capable of neutralizing between nine and all 18 of the strains tested. Nevertheless, from the results of this study, it can be concluded that in colostrum-deprived BVDV seronegative calves, Rispoval RS-BVD can stimulate the production of VN antibodies capable of neutralizing a wide range of antigenically diverse European isolates of BVDV, including genotypes I and II.