Maria Stokstad

Bluetongue: a historical and epidemiological perspective with the emphasis on South Africa

Bluetongue (BT) is a non-contagious, infectious, arthropod transmitted viral disease of domestic and wild ruminants that is caused by the bluetongue virus (BTV), the prototype member of the Orbivirus genus in the family Reoviridae. Bluetongue was first described in South Africa, where it has probably been endemic in wild ruminants since antiquity. Since its discovery BT has had a major impact on sheep breeders in the country and has therefore been a key focus of research at the Onderstepoort Veterinary Research Institute in Pretoria, South Africa. Several key discoveries were made at this Institute, including the demonstration that the aetiological agent of BT was a dsRNA virus that is transmitted by Culicoides midges and that multiple BTV serotypes circulate in nature. It is currently recognized that BT is endemic throughout most of South Africa and 22 of the 26 known serotypes have been detected in the region. Multiple serotypes circulate each vector season with the occurrence of different serotypes depending largely on herd-immunity. Indigenous sheep breeds, cattle and wild ruminants are frequently infected but rarely demonstrate clinical signs, whereas improved European sheep breeds are most susceptible. The immunization of susceptible sheep remains the most effective and practical control measure against BT. In order to protect sheep against multiple circulating serotypes, three pentavalent attenuated vaccines have been developed. Despite the proven efficacy of these vaccines in protecting sheep against the disease, several disadvantages are associated with their use in the field.

Indirect transmission of bovine viral diarrhoea virus at calving and during the postparturient period.

Two trials were designed to investigate whether bovine viral diarrhoea virus (BVDV) could be transmitted after the birth of persistently infected calves, even if they were removed immediately after birth. In trial 1, 11 calves were actively exposed to fetal fluids and uterine lochia collected from cows that had delivered calves persistently infected with type 1 BVDV. One calf that was exposed to a sample taken on the day of calving seroconverted. In trial 2, six calves were housed in stables where persistently infected calves were being born and then removed immediately from their dams and from the stable unit within two to three hours. One of four calves in close contact with the cows after delivery seroconverted and one of two calves housed within the same stable unit became infected.

Transplacental infection in goats experimentally infected with a European strain of bluetongue virus serotype 8

The capability of the recently emerged European strain of bluetongue virus serotype 8 (BTV-8) to cross the ruminant placenta has been established in experimental and field studies in both sheep and cattle. Seroprevalence rates in goats in North-Western Europe were high during the recent outbreak of BTV-8; however the capability of the virus to infect goats through the transplacental route has not been established. In the present study, four Saanen goats were inoculated with the European strain of BTV-8 at 62 days of gestation; this resulted in mild clinical signs, however gross lesions observed post mortem were more severe. Viral RNA was detected by real-time RT-PCR in blood and tissue samples from three fetuses harvested from two goats at 43 days post infection. Conventional RT-PCR and genome sequencing targeting viral segment 2 confirmed infection of brain tissue with BTV-8 in two of these fetuses. In total, five of six fetuses demonstrated lesions that may have been associated with transplacental infection with BTV. Infected fetuses did not demonstrate neurological lesions. Low viral RNA concentrations in fetal blood and tissue further suggest that the infected fetuses would probably not have been born viraemic. The implications of these findings with regards to the epidemiology and overwintering of BTV-8 in Europe remains unclear.

Bovine respiratory syncytial virus: infection dynamics within and between herds

The infection dynamics of bovine respiratory syncytial virus (BRSV) were studied in randomly selected Norwegian dairy herds. A total of 134 herds were tested twice, six months apart. The herds were classified as positive for BRSV if at least one animal between 150 and 365 days old tested positive for antibodies against BRSV, thereby representing herds that had most likely had the virus present during the previous year. The prevalence of positive herds at the first and second sampling was 34 per cent and at 41 per cent, respectively, but varied greatly between regions. Negative herds were found in close proximity to positive herds. Some of these herds remained negative despite several new infections nearby. Of the herds initially being negative, 42 per cent changed status to positive during the six months. This occurred at the same rate during summer as winter, but a higher rate of animals in the herds was positive if it took place during winter. Of the herds initially being positive, 33 per cent changed to negative. This indicates that an effective strategy to lower the prevalence and the impact of BRSV could be to employ close surveillance and place a high biosecurity focus on the negative herds.

A review of experimental infections with bluetongue virus in the mammalian host

Abstract Experimental infection studies with bluetongue virus (BTV) in the mammalian host have a history that stretches back to the late 18th century. Studies in a wide range of ruminant and camelid species as well as mice have been instrumental in understanding BTV transmission, bluetongue (BT) pathogenicity/pathogenesis, viral virulence, the induced immune response, as well as reproductive failures associated with BTV infection. These studies have in many cases been complemented by in vitro studies with BTV in different cell types in tissue culture. Together these studies have formed the basis for the understanding of BTV-host interaction and have contributed to the design of successful control strategies, including the development of effective vaccines. This review describes some of the fundamental and contemporary infection studies that have been conducted with BTV in the mammalian host and provides an overview of the principal animal welfare issues that should be considered when designing experimental infection studies with BTV in in vivo infection models. Examples are provided from the authors’ own laboratory where the three Rs (replacement, reduction and refinement) have been implemented in the design of experimental infection studies with BTV in mice and goats. The use of the ARRIVE guidelines for the reporting of data from animal infection studies is emphasized.

Occurrence and phylogenetic analysis of bovine respiratory syncytial virus in outbreaks of respiratory disease in Norway.

BackgroundBovine respiratory syncytial virus (BRSV) is one of the major pathogens involved in the bovine respiratory disease (BRD) complex. The seroprevalence to BRSV in Norwegian cattle herds is high, but its role in epidemics of respiratory disease is unclear. The aims of the study were to investigate the etiological role of BRSV and other respiratory viruses in epidemics of BRD and to perform phylogenetic analysis of Norwegian BRSV strains.ResultsBRSV infection was detected either serologically and/or virologically in 18 (86%) of 21 outbreaks and in most cases as a single viral agent. When serology indicated that bovine coronavirus and/or bovine parainfluenza virus 3 were present, the number of BRSV positive animals in the herd was always higher, supporting the view of BRSV as the main pathogen. Sequencing of the G gene of BRSV positive samples showed that the current circulating Norwegian BRSVs belong to genetic subgroup II, along with other North European isolates. One isolate from an outbreak in Norway in 1976 was also investigated. This strain formed a separate branch in subgroup II, clearly different from the current Scandinavian sequences. The currently circulating BRSV could be divided into two different strains that were present in the same geographical area at the same time. The sequence variations between the two strains were in an antigenic important part of the G protein.ConclusionThe results demonstrated that BRSV is the most important etiological agent of epidemics of BRD in Norway and that it often acts as the only viral agent. The phylogenetic analysis of the Norwegian strains of BRSV and several previously published isolates supported the theory of geographical and temporal clustering of BRSV.

Bovine respiratory syncytial virus and bovine coronavirus antibodies in bulk tank milk - risk factors and spatial analysis.

Abstract Bovine respiratory syncytial virus (BRSV) and bovine coronavirus (BCoV) are considered widespread among cattle in Norway and worldwide. This cross-sectional study was conducted based on antibody-ELISA of bulk tank milk (BTM) from 1347 herds in two neighboring counties in western Norway. The study aims were to determine the seroprevalence at herd level, to evaluate risk factors for BRSV and BCoV seropositivity, and to assess how these factors were associated with the spatial distribution of positive herds. The overall prevalence of BRSV and BCoV positive herds in the region was 46.2% and 72.2%, respectively. Isopleth maps of the prevalence risk distribution showed large differences in prevalence risk across the study area, with the highest prevalence in the northern region. Common risk factors of importance for both viruses were herd size, geographic location, and proximity to neighbors. Seropositivity for one virus was associated with increased odds of seropositivity for the other virus. Purchase of livestock was an additional risk factor for BCoV seropositivity, included in the model as in-degree, which was defined as the number of incoming movements from individual herds, through animal purchase, over a period of five years. Local dependence and the contribution of risk factors to this effect were assessed using the residuals from two logistic regression models for each virus. One model contained only the x- and y- coordinates as predictors, the other had all significant predictors included. Spatial clusters of high values of residuals were detected using the normal model of the spatial scan statistic and visualized on maps. Adjusting for the risk factors in the final models had different impact on the spatial clusters for the two viruses: For BRSV the number of clusters was reduced from six to four, for BCoV the number of clusters remained the same, however the log-likelihood ratios changed notably. This indicates that geographical differences in proximity to neighbors, herd size and animal movements explain some of the spatial clusters of BRSV- and BCoV seropositivity, but far from all. The remaining local dependence in the residuals show that the antibody status of one herd is influenced by the antibody status of its neighbors, indicating the importance of indirect transmission and that increased biosecurity routines might be an important preventive strategy.

Association between the level of antibodies in bulk tank milk and bovine respiratory syncytial virus exposure in the herd.

Antibody levels in bulk tank milk (BTM) against bovine respiratory syncytial virus (BRSV) are used to classify BRSV status of herds. The aim of this study was to investigate how these levels correspond with the time at which the herds were infected. Bulk tank milk, individual milk and serum samples from cows and young stock were investigated using an indirect ELISA. Screenings of BTM from 89 dairy herds during two winter seasons revealed a prevalence of positive herds from 82 per cent to 85 per cent. Eleven herds showed a marked increase in antibody levels between two screenings, indicating new infection. However, two of these herds had been free from BRSV for the last five to seven years. Two newly infected herds were monitored for four years and did not appear to get reinfected. Surprisingly, the BTM antibody levels in these herds remained high throughout the study period, but fluctuated significantly. This shows that the levels of antibodies in BTM can remain high for several years, even in herds where reinfection does not occur. BTM serology is a useful tool in the monitoring of infectious diseases in dairy herds, but has limitations as a diagnostic tool for BRSV infections.

A cohort study of the effect of winter dysentery on herd-level milk production

ABSTRACT Winter dysentery (WD) is a contagious disease caused by bovine coronavirus. It is characterized by acute onset of diarrhea, fever, depression, and reduced milk yield in adult cattle. Although production loss is a well-known consequence of WD, large-scale studies estimating the effect on milk production are lacking. The objective of this study was to estimate the effect of farmer-reported WD on herd-level milk production and milk composition. A cohort study was performed based on reports of herd outbreaks of winter dysentery during a regional epidemic in Norway during the winter of 2011–2012. Reports were made by farmers, and diagnosis was based on a herd outbreak of acute diarrhea in adults. Milk shipment data were retrieved from the dairy company, and information on herd size and milking system were retrieved from the Norwegian Dairy Herd Recording System. We compared milk production in herds with reported outbreaks of WD (n = 224) with all herds in the same area without a reported outbreak (n = 2,093) during the same period. The outcome variable in the analysis was milk volume per cow per day, and the main predictor was whether the herd had a reported outbreak of WD or not. We assessed the effect of WD on milk production by fitting a linear mixed model, adjusting for milk production in the herd before the outbreak. Similarly, we assessed the effect of WD on milk composition using linear regression, adjusting for the levels of milk components before the outbreak. This study estimated a total loss of 51 L/cow during the study period, from 7 d before to 19 d after a reported outbreak. The lowest estimated production was 2 d after the outbreak was reported, when the average milk yield was 19.4 L/cow per day, compared with 23.0 L/cow per day 7 days before notification (i.e., a difference of 3.6 L/cow, or 15%). The effect gradually declined with time. The estimated effect on milk composition was modest, but an increase of 11% in free fatty acids and a small increase in fat/protein ratio indicated that WD might put cows into negative energy balance. Descriptive analysis indicated that herd milk yield was still reduced 4 mo after an outbreak. This cohort study showed that WD causes considerable decreases in milk production, and it alters milk composition. These findings highlight the important negative consequences of WD, and should motivate actions to prevent between-herd spread of bovine coronavirus.

Evaluation of a multiplex immunoassay for bovine respiratory syncytial virus and bovine coronavirus antibodies in bulk tank milk against two indirect ELISAs using latent class analysis

Abstract Bovine respiratory syncytial virus (BRSV) and bovine coronavirus (BCV) are responsible for respiratory disease and diarrhea in cattle worldwide. The Norwegian control program against these infections is based on herd-level diagnosis using a new multiplex immunoassay. The objective of this study was to estimate sensitivity and specificity across different cut-off values for the MVD-Enferplex BCV/BRSV multiplex, by comparing them to a commercially available ELISA, the SVANOVIR® BCV-Ab and SVANOVIR® BRSV-Ab, respectively. We analyzed bulk tank milk samples from 360 herds in a low- and 360 herds in a high-prevalence area. As none of the tests were considered perfect, estimation of test characteristics was performed using Bayesian latent class models. At the manufacturers’ recommended cut-off values, the median sensitivity for the BRSV multiplex and the BRSV ELISA was 94.4 [89.8–98.7 95% Posterior Credibility Interval (PCI)] and 99.8 [98.7–100 95% PCI], respectively. The median specificity for the BRSV multiplex was 90.6 [85.5–94.4 95% PCI], but only 57.4 [50.5–64.4 95% PCI] for the BRSV ELISA. However, increasing the cut-off of the BRSV ELISA increased specificity without compromising sensitivity. For the BCV multiplex we found that by using only one of the three antigens included in the test, the specificity increased, without concurrent loss in sensitivity. At the recommended cut-off this resulted in a sensitivity of 99.9 [99.3–100 95% PCI] and specificity of 93.7 [88.8–97.8 95% PCI] for the multiplex and a sensitivity of 99.5 [98.1–100 95% PCI] and a specificity of 99.6 [97.6–100 95% PCI] for the BCV ELISA.