Epidemiology of bluetongue virus serotype 8 outbreaks in the Netherlands in 2006

In August 2006 a major epidemic of Bluetongue (BT) occurred in north-western Europe, affecting The Netherlands, Belgium, Germany, Luxemburg, and the north of France. It was caused by Br virus serotype 8 (BTV-8), a serotype previously unknown to the EU. Although clinical disease is usually restricted to sheep, this virus also caused clinical disease in a small proportion of cattle. The last clinical outbreak of BT in The Netherlands occurred mid-December 2006. The delay between observation of the first clinical signs by the owner and reporting of a clinically suspect BT situation to the veterinary authorities was approximately 2 weeks. BTV-8-associated clinical signs were more prominent in sheep than in cattle, and the relative frequency of specific clinical signs was different in cattle and sheep. Morbidity and mortality rates were significantly higher among sheep than among cattle, and a higher proportion of cattle than sheep recovered from clinical disease.     

Possible routes of introduction of bluetongue virus serotype 8 into the epicentre of the 2006 epidemic in north-western Europe

In August 2006, bluetongue (BT) was notified in The Netherlands on several animal holdings. This was the onset of a rapidly spreading BT-epidemic in north-western Europe (latitude >51 degrees N) that affected cattle and sheep holdings in The Netherlands, Belgium, Germany, France and Luxembourg. The outbreaks were caused by bluetongue virus (BTV) serotype 8, which had not been identified in the European Union before. Bluetongue virus can be introduced into a free area by movement of infected ruminants, infected midges or by infected semen and embryos. In this study, information on animal movements or transfer of ruminant germ plasms (semen and embryos) into the Area of First Infection (AFI), which occurred before and during the onset of the epidemic, were investigated in order to establish the conditions for the introduction of this virus. All inbound transfers of domestic or wild ruminants, non-susceptible mammal species and ruminant germ plasms into the AFI during the high-risk period (HRP), registered by the Trade Control and Expert System (TRACES) of the EC, were obtained. Imports originating from countries with a known or suspected history of BTV-incidence of any serotype were identified. The list of countries with a reported history of BTV incidence was obtained from the OIE Handistatus II for the period from 1996 until 2004. No ruminants were imported from a Member State (MS) with a known history of BTV-8 or from any other country with a known or suspected history of BTV incidence of any serotype. Of all non-susceptible mammal species only 233 horses were transported directly into the AFI during the HRP. No importations of semen or embryos into the AFI were registered in TRACES during the period of interest. An obvious source for the introduction of BTV-8, such as import of infected ruminants, could not be identified and the exact origin and route of the introduction of BTV-8 thus far remains unknown. However, the absence of legal import of ruminants from outside the EU into the AFI and the absence of BTV-8 in southern Europe suggest that, the introduction of the BTV-8 infection into the north-western part of Europe took place via another route. Specifically, in relation to this, the potential for Culicoides to be imported along with or independently of the import of animals, plants or other 'materials', and the effectiveness of measures to reduce such a possibility, merit further study.     

Impact of human interventions on the spread of bluetongue virus serotype 8 during the 2006 epidemic in north-western Europe

Bluetongue virus (BTV) can be spread by movement or migration of infected ruminants. Infected midges (Culicoides sp.) can be dispersed with livestock or on the wind. Transmissions of infection from host to host by semen and trans-placental infection of the embryo from the dam have been found. As for any infectious animal disease, the spread of BTV can be heavily influenced by human interventions preventing or facilitating the transmission pathways. This paper describes the results of investigations that were conducted on the potential role of the above-mentioned human interventions on the spread of BTV-8 during the 2006 epidemic in north-western Europe. Data on surveillance and control measures implemented in the affected European Union (EU) Member States (MS) were extracted from the legislation and procedures adopted by the national authorities in Belgium, France, Germany, and The Netherlands. The impact of the control measures on the BTV-incidence in time and space was explored. Data on ruminant transports leaving the area of first infection (AFI) to other areas within and beyond the affected MS were obtained from the national identification and registration systems of the three initially affected MS (Belgium, Germany, The Netherlands) and from the Trade Control and Expert System (TRACES) of the European Commission. The association between the cumulative number of cases that occurred in a municipality outside the AFI and the number of movements or the number of animals moved from the AFI to that municipality was assessed using a linear negative binomial regression model. The results of this study indicated that the control measures which were implemented in the affected MS (in accordance with EU directives) were not able to fully stop further spread of BTV and to control the epidemic. This finding is not surprising because BT is a vector-borne disease and it is difficult to limit vector movements. We could not assess the consequences of not taking control measures at all but it is possible, if not most likely, that this would have resulted in even wider spread. The study also showed an indication of the possible involvement of animal movements in the spread of BTV during the epidemic. Therefore, the prevention of animal movements remains an important tool to control BTV outbreaks. The extension of the epidemic to the east cannot be explained by the movement of animals, which mainly occurred in a north-western direction. This indicates that it is important to consider other influential factors such as dispersal of infected vectors depending on wind direction, or local spread.     

Transplacental and oral transmission of wild-type bluetongue virus serotype 8 in cattle after experimental infection

Potential vertical transmission of wild-type bluetongue virus serotype 8 (BTV-8) in cattle was explored in this experiment. We demonstrated transplacental transmission of wild-type BTV-8 in one calf and oral infection with BTV-8 in another calf. Following the experimental BTV-8 infection of seven out of fifteen multi-parous cows eight months in gestation, each newborn calf was tested prior to colostrum intake for transplacental transmission of BTV by RRT-PCR. If transplacental transmission was not established the calves were fed colostrum from infected dams or colostrum from non-infected dams spiked with BTV-8 containing blood. One calf from an infected dam was born RRT-PCR positive and BTV-specific antibody (Abs) negative, BTV was isolated from its blood. It was born with clinical signs resembling bluetongue and lived for two days. Its post-mortem tissue suspensions were RRT-PCR positive. Of the seven calves fed colostrum from infected dams, none became infected. Of the six calves fed colostrum from non-infected dams spiked with infected blood, one calf became PCR-positive at day 8 post-partum (dpp), seroconverted 27 days later, and remained RRT-PCR and Abs positive for the duration of the experiment (i.e., 70 dpp). This work demonstrates that transplacental transmission in late gestation and oral infection of the neonate with wild-type BTV-8 is possible in cattle under experimental conditions.     

Molecular epidemiology of bluetongue virus serotype 1 isolated in 2006 from Algeria

This study reports on an outbreak of disease that occurred in central Algeria during July 2006. Sheep in the affected area presented clinical signs typical of bluetongue (BT) disease. A total of 5245 sheep in the affected region were considered to be susceptible, with 263 cases and thirty-six deaths. Bluetongue virus (BTV) serotype 1 was isolated and identified as the causative agent. Segments 2, 7 and 10 of this virus were sequenced and compared with other isolates from Morocco, Italy, Portugal and France showing that they all belong to a ‘western’ BTV group/topotype and collectively represent a western Mediterranean lineage of BTV-1.     

Modelling local dispersal of bluetongue virus serotype 8 using random walk

The knowledge of the place where a disease is first introduced and from where it later spreads is a key element for the understanding of an epizootic. For a contagious disease, the main method is back tracing. For a vector-borne disease such as the Bluetongue virus serotype 8 epizootic that occurred in 2006 in North-Western Europe, the efficiency of tracing is limited because many infected animals are not showing clinical signs. In the present study, we propose to use a statistical approach, random walk, to model local spread in order to derive the Area of First Infection (AFI) and spread rate. Local spread is basically described by the random movements of infected insect vectors. Our model localised the AFI centre, origin of the infection, in the Netherlands, South of Maastricht. This location is consistent with the location of the farms where the disease was first notified in the three countries (Netherlands, Belgium, and Germany) and the farm where retrospectively the earliest clinical signs were found. The derived rate of spread of 10-15km/week is consistent with the rates observed in other Bluetongue epizootics. In another article Mintiens (2008), the AFI definition has then been used to investigate possible ways of introduction (upstream tracing) and to study the effect of animal movements from this area (downstream tracing).     

Experimental infection of South American camelids with bluetongue virus serotype 8

Bluetongue (BT) is an infectious, non-contagious disease of wild and domestic ruminants. It is caused by bluetongue virus (BTV) and transmitted by Culicoides biting midges. Since 1998, BT has been emerging throughout Europe, threatening not only the na?ve ruminant population. Historically, South American camelids (SAC) were considered to be resistant to BT disease. However, recent fatalities related to BTV in captive SAC have raised questions about their role in BTV epidemiology. Data on the susceptibility of SAC to experimental infection with BTV serotype 8 (BTV-8) were collected in an animal experiment. Three alpacas (Vicugna pacos) and three llamas (Lama glama) were experimentally infected with BTV-8. They displayed very mild clinical signs. Seroconversion was first measured 6-8 days after infection (dpi) by ELISA, and neutralising antibodies appeared 10-13 dpi. BTV-8 RNA levels in blood were very low, and quickly cleared after seroconversion. However, spleens collected post-mortem were still positive for BTV RNA, over 71 days after the last detection in blood samples. Virus isolation was only possible from blood samples of two alpacas by inoculation of highly sensitive interferon alpha/beta receptor-deficient (IFNAR(-/-)) mice. An in vitro experiment demonstrated that significantly lower amounts of BTV-8 adsorb to SAC blood cells than to bovine blood cells. Although this experiment showed that SAC are generally susceptible to a BTV-8 infection, it indicates that these species play a negligible role in BTV epidemiology.     

Mortality attributable to bluetongue virus serotype 8 infection in Dutch dairy cows

In 2007, bluetongue virus serotype 8 (BTV-8) re-emerged in the Netherlands and a large number of farmers notified morbidity and mortality associated with BTV-8 to the authorities. All dead cows in the Netherlands are registered in one of the three age classes: newborn calves <3 days, calves 3 days to 1 year, and cows >1 year. These registrations result in a complete data set of dead cattle per herd per day from 2003 until 2007. In this study, the mortality associated with BTV-8 for the Dutch dairy industry was estimated, based on this census data. Default, mortality associated with BTV-8 was estimated for the confirmed notification herds. Moreover, an additional analysis was performed to determine if mortality associated with BTV-8 infection occurred in non-notification herds located in BTV-8 infected compartments. A multivariable population-averaged model with a log link function was used for analyses. Separate analyses were conducted for the three different age groups. Confirmed notification herds had an increased cow mortality rate ratio (MRR) (1.4 (95% CI: 1.2-1.6)); calf MRR (1.3 (95% CI: 1.1-1.4)); and newborn calf MRR (1.2 (95% CI: 1.1-1.3)). Furthermore, in non-notification herds in BTV-8 infected compartments, mortality significantly increased 1.1 times (95% CI: 1.1-1.1) in cows, 1.2 times (95% CI: 1.2-1.2) in calves and 1.1 times (1.1-1.1) in newborn calves compared with BTV-8 non-infected months. Using objective census data over a 5-year period, the MRRs indicated increased mortality associated with BTV-8 infection not only in herds of which the farmer notified clinical signs but also in non-notification herds in infected compartments.     

Economic evaluation of the surveillance and intervention programme for bluetongue virus serotype 8 in Switzerland

Empirical analyses founded on sound economic principles are essential in advising policy makers on the efficiency of resource use for disease mitigation. Surveillance and intervention are resource-using activities directed at mitigation. Surveillance helps to offset negative disease effects by promoting successful intervention. Intervention is the process of implementing measures (e.g. vaccination or medication) to reduce or remove a hazard in a population. The scale and ratios in which the two are combined affect the efficiency of mitigation, its costs, benefits, and thus net effect on society's well-being. The Swiss national mitigation programme for bluetongue virus serotype 8 was used as case study to investigate the economic efficiency of mitigation. In 2008, Switzerland implemented a vaccination programme to avoid and reduce disease and infection in its ruminant population. To monitor the vaccination programme and the vector dynamics, a surveillance system consisting of serological and entomological surveillance was established. Retrospective analyses for the years 2008-2009 and prospective analyses for the years 2010-2012 were conducted to investigate if the mitigation programme was economically beneficial. In the retrospective analysis, the implemented programme (=comparative scenario) was compared to a hypothesised baseline scenario of voluntary vaccination and surveillance. In the prospective analysis, the comparative scenario assumed to continue was compared to two baseline scenarios: one of voluntary vaccination combined with surveillance and one of no vaccination combined with surveillance. For each scenario, monetary surveillance, intervention and disease costs were calculated. The comparison of baseline and comparative scenarios yielded estimates for the total benefit (=disease costs avoided), margin over intervention cost and the net value of the programme. For 2008-2009, in aggregate, the mean biannual total benefit was 17.46 m Swiss francs (CHF) (1CHF=0.66€ at the time of analysis) and the mean net benefit after subtraction of the intervention and surveillance cost was 3.95 m CHF. For the three years 2010-2012, overall net costs were estimated at 12.93 m and 8.11 m CHF, respectively, for comparison of the implemented mitigation programme with the two baseline scenarios. It was concluded that the surveillance and intervention programme implemented in 2008-2009 was economically beneficial, while its continuation in the same form in 2010-2012 would produce net costs. These costs were due to the mean intervention cost remaining constant at a level of approximately 11 m CHF per year while the mean total benefit would be gradually reduced in 2010-2012 due to the reduced occurrence of disease in a fully vaccinated population.     

Evaluation of antibody-ELISA and real-time RT-PCR for the diagnosis and profiling of bluetongue virus serotype 8 during the epidemic in Belgium in 2006

In 2006 bluetongue (BT) emerged for the first time in North-Western Europe. Reliable diagnostic tools are essential in controlling BT but data on the diagnostic sensitivity (Se) and specificity (Sp) are often missing. This paper aims to describe and analyse the results obtained with the diagnostics used in Belgium during the 2006 BT crisis. The diagnosis was based on a combination of antibody detection (competitive ELISA, cELISA) and viral RNA detection by real-time RT-PCR (RT-qPCR). The performance of the cELISA as a diagnostic tool was assessed on field results obtained during the epidemic and previous surveillance campaigns. As the infectious status of the animals is unknown during an epidemic, a Bayesian analysis was performed. Both assays were found to be equally specific (RT-qPCR: 98.5%; cELISA: 98.2%) while the diagnostic sensitivity of the RT-qPCR (99.5%) was superior to that of the cELISA (87.8%). The assumption of RT-qPCR as standard of comparison during the bluetongue virus (BTV) epidemic proved valid based on the results of the Bayesian analysis. A ROC analysis of the cELISA, using RT-qPCR as standard of comparison, showed that the cut-off point with the highest accuracy occurred at a percentage negativity of 66, which is markedly higher than the cut-off proposed by the manufacturer. The analysis of the results was further extended to serological and molecular profiling and the possible use of profiling as a rapid epidemiological marker of the BTV in-field situation was assessed. A comparison of the serological profiles obtained before, during and at the end of the Belgian epidemic clearly showed the existence of an intermediate zone which appears soon after BTV (re)enters the population. The appearance or disappearance of this intermediate zone is correlated with virus circulation and provides valuable information, which would be entirely overlooked if only positive and negative results were considered.     

Trajectory analysis and bluetongue virus serotype 2 in Florida 1982.

Examination of Northern Hemisphere synoptic charts and computation of backward trajectories indicated that Culicoides infected with bluetongue virus serotype 2 could have been carried on the wind and brought the virus to Florida on the afternoon of August 19, 1982 after leaving northern Cuba the previous evening. Flight would have occurred at a height of 1-1.5 km at temperatures of 15-17 degrees C. The distance of 500 km from northern Cuba to Ona would have been covered in 20 h at an average speed of 25 km h-1. Computation of trajectories indicated that a second electropherotype, Ona B, was unlikely to have been introduced by infected Culicoides.     

Bovine infection with bluetongue virus with special emphasis on European serotype 8

Bluetongue virus (BTV) is an arthropod-borne virus infecting domestic and wild ruminants. Infection in cattle is commonly asymptomatic and characterised by a long viraemia. Associated with the emergence and the recrudescence of BTV serotype 8 (BTV-8) in Northern and Central Europe, remarkable differences have been noticed in the transmission and in the clinical expression of the disease, with cattle showing clinical illness and reproductive disorders such as abortion, stillbirth and fetal abnormalities. Several investigations have already indicated the putative ability of the European BTV-8 strain to cross the bovine placenta and to cause congenital infections. The current epidemiological and pathological findings present an unusual picture of the disease in affected bovines.     

Performance of clinical signs to detect bluetongue virus serotype 8 outbreaks in cattle and sheep during the 2006-epidemic in The Netherlands

The performance of clinical signs as a diagnostic test for the detection of BTV-8 outbreaks during the 2006-epidemic in The Netherlands was evaluated by constructing and analysing receiver operating characteristic (ROC) curves. The area under the ROC curve of the BT-associated clinical signs in cattle was 0.77. An optimal efficient test (maximising both sensitivity and specificity) in cattle herds combined a sensitivity (Se) of 67% with a specificity (Sp) of 72%, comprising the following clinical signs: ulcerations and/or erosions of oral mucosa or erosions of lips/crusts in or around nostrils or oedema of the nose or hyperaemic/purple coloration of tongue, tongue protrusion or coronitis or apathy/tiredness or muscle necrosis, stiffness of limbs or loathing or refusal to move, prostration or torticollis or anoestrus. The area under the ROC curve of the BT-associated clinical signs in sheep was 0.81. The optimal efficient test in sheep flocks combined a Se of 76% with a Sp of 72%, comprising the following clinical signs: ulcerations of oral mucosa or serous nasal discharge or erosions/ulceration of tongue mucosa or hypersensitivity of the skin or muscle necrosis, stiffness of limbs or coronitis or grinding of teeth or salivation or weakness/paresis.     

Susceptibility of in vitro produced hatched bovine blastocysts to infection with bluetongue virus serotype 8

ABSTRACT: Bluetongue virus serotype 8 (BTV-8), which caused an epidemic in ruminants in central Western Europe in 2006 and 2007, seems to differ from other bluetongue serotypes in that it can spread transplacentally and has been associated with an increased incidence of abortion and other reproductive problems. For these reasons, and also because BTV-8 is threatening to spread to other parts of the world, there is a need for more information on the consequences of infection during pregnancy. The aim of the present study was to investigate whether hatched (i.e. zona pellucida-free) in vitro produced bovine blastocysts at 8-9 days post insemination are susceptible to BTV-8 and whether such infection induces cell death as indicated by apoptosis. Exposure of hatched in vitro produced bovine blastocysts for 1 h to a medium containing 103.8 or 104.9 TCID50 of the virus resulted in active viral replication in between 25 and 100% of the cells at 72 h post exposure. The infected blastocysts also showed growth arrest as evidenced by lower total cell numbers and a significant level of cellular apoptosis. We conclude from this in vitro study that some of the reproductive problems that are reported when cattle herds are infected with BTV-8 may be attributed to direct infection of blastocysts and other early-stage embryos in utero.     

Scaling from challenge experiments to the field: Quantifying the impact of vaccination on the transmission of bluetongue virus serotype 8

Bluetongue (BT) is an economically important disease of ruminants caused by bluetongue virus (BTV) and transmitted by Culicoides biting midges. The most practical and effective way to protect susceptible animals against BTV is by vaccination. Data from challenge studies in calves and sheep conducted by Intervet International b.v., in particular, presence of viral RNA in the blood of challenged animals, were used to estimate vaccine efficacy. The results of the challenge studies for calves indicated that vaccination is likely to reduce the basic reproduction number (R(0)) for BTV in cattle to below one (i.e. prevent major outbreaks within a holding) and that this reduction is robust to uncertainty in the model parameters. Sensitivity analysis showed that the whether or not vaccination is predicted to reduce R(0) to below one depended on the following assumptions: (i) whether "doubtful" results from the challenge studies are treated as negative or positive; (ii) whether or not the probability of transmission from host to vector is reduced by vaccination; and (iii) whether the extrinsic incubation period follows a realistic gamma distribution or the more commonly used exponential distribution. For sheep, all but one of the vaccinated animals were protected and, consequently, vaccination will consistently reduce R(0) in sheep to below one. Using a stochastic spatial model for the spread of BTV in Great Britain (GB), vaccination was predicted to reduce both the incidence of disease and spatial spread in simulated BTV outbreaks in GB, in both reactive vaccination strategies and when an incursion occurred into a previously vaccinated population.     

A new bluetongue virus serotype isolated in Kenya.

An apparently new strain of bluetongue virus was first isolated in Kenya in 1965 and since, has been obtained on 7 further occasions from diseased sheep during clinical outbreaks of disease. It proved to be serologically different from the 16 bluetongue virus strains then held at this laboratory. The virus was modified by passage in embryonated hens eggs to produce a live virus strain suitable for inclusion in a polyvalent vaccine. Recent neutralisation tests, carried out with 24 guinea pig immune sera prepared at Pirbright against the currently known World serotypes, have confirmed the earlier results and show that it is different from any of the existing serotypes.