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.     

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).     

Bluetongue disease in Swiss sheep breeds: clinical signs after experimental infection with bluetongue virus serotype 8

Clinical disease of bluetongue (BT) in sheep may differ depending on breed, age and immunity of infected sheep and may also vary between serotype and strain of BT virus (BTV). Since there are no data available on the susceptibility of Swiss sheep breeds for BT, we performed experimental infection of the 4 most common Swiss sheep breeds and the highly susceptible Poll Dorset sheep with the BTV serotype 8 (BTV-8) circulating in Northern Europe since 2006. Clinical signs were assessed regarding severity, localisation, progression and time point of their appearance. The results clearly show that the Swiss sheep breeds investigated were susceptible to BTV-8 infection. They developed moderate, BT-characteristic symptoms, which were similar to those observed in Poll Dorset sheep. Regardless of breed, the majority of infected animals showed fever, swelling of the head as well as erosions of the mouth and subcutaneous haemorrhages.     

Impact of a natural bluetongue serotype 8 infection on semen quality of Belgian rams in 2007

In 2006, bluetongue (BT) virus serotype 8 emerged in northern Europe and numerous ruminants were affected in the following year. Infertility in males is one of the consequences of BT, although its severity and duration after natural infection has not been documented. In this report, the impact of BT-8 on clinical signs and semen quality of naturally infected rams is described through a longitudinal study of two Belgian ram populations (n = 12 and n = 24) and a cross sectional study in a further ram population (n = 43).Macroscopic semen characteristics, semen concentration, motility, percentage of living and dead spermatozoa were assessed in 167 semen samples collected on 1–6 occasions from 79 BT-8 infected rams within 5–138 days after onset of clinical disease. These were compared with healthy control animals. Significant changes in all variables were observed after natural BT-8 infection. Total recovery occurred around 85 days after clinical disease in animals undergoing a close follow-up of semen quality. Good correspondence between the results of the longitudinal and cross sectional studies suggests that semen quality of BT-8 affected rams reached normal references values 63–138 days after clinical diagnosis of BT. In addition, semen concentration seems to be a sound epidemiological indicator of ram semen quality.     

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.     

Active circulation of bluetongue vaccine virus serotype-2 among unvaccinated cattle in central Italy

Several seroconversions occurring in 2002 among sentinel cattle during the bluetongue-vaccination campaign in Lazio and Tuscany (central Italy) led to the suspicion of vaccine-virus circulation. Therefore in 2003, 17 seroconverting sentinel herds were investigated for the characteristics of the virus involved. From these farms, 91 unvaccinated animals and 57 Culicoides pools were tested for the presence of the bluetongue vaccine virus (serotype-2) or other strains. The presence of vaccine virus serotype-2 was confirmed by PCR followed by restriction analysis in the whole blood of 17 unvaccinated sentinel cattle and 12 pools of Culicoides imicola or C. obsoletus. Of the 17 herds, five were positive only for vaccine virus serotype-2, four were positive for other strains and two for both the vaccine and other strains; the remaining premises were virologicaly negative. The vaccine virus serotype-2 also was detected in areas not included in the vaccination campaign.     

Bluetongue virus serotype 26: Infection kinetics,pathogenesis and possible contact transmission in goats

The aim of this study was to assess the pathogenicity and infection kinetics of Bluetongue virus serotype 26 (BTV-26) in goats. Out of a group of six goats housed in insect free accommodation, five were experimentally infected with BTV-26 and one was kept uninfected as an in-contact control. Samples taken throughout the study were used to determine the kinetics of infection using a pan specific BTV real time RT-PCR assay and a group specific ELISA. The five infected goats did not show clinical signs of BTV, however high levels of viral RNA were detected and virus was isolated from the blood of all 5 goats. Antibodies against BTV were first detected between 7 and 11 dpi in all 5 experimentally infected goats. Interestingly at 21 dpi viral RNA was detected in, and virus was isolated from, the blood of the in-contact control goat, which also seroconverted. These results suggest that BTV-26 replicates to high levels in goats, causing no obvious clinical disease, suggesting that goats may be the natural host for this virus. Preliminary evidence also indicates that BTV-26 may be spread by contact transmission between goats, however a more detailed study is required in order to confirm this observation.     

Transplacental transmission of Bluetongue virus serotype 8 in ewes in early and mid gestation

The ability of Bluetongue virus serotype 8 (BTV-8) originating from the 2006 European outbreak to cross the ovine placenta during early and mid gestation was investigated in two separate experiments. In the first experiment, 16 ewes were infected with BTV-8 at 70-75 days gestation. The foetuses were collected at 18-19 days after infection (dpi). BTV-8 could be isolated from at least two organs of 19 out of 40 lambs and from 11 out of 16 infected ewes. In the second experiment, 20 BTV-8 infected ewes in early gestation (day 40-45) were euthanized at 10 days (10 ewes) or 30 days (10 ewes) after infection. The presence of BTV could be demonstrated in two foetuses from two ewes at 10 dpi and in 4 foetuses from four ewes at 30 dpi. The main pathological findings in the foetuses in mid gestation were meningo-encephalitis and vacuolation of the cerebrum. In the foetuses early at gestation, haemorrhages in various foetal tissues and necrosis and haemorrhages in the placentomes were found. These experiments demonstrate for the first time the presence of infectious BTV in lamb foetuses at different stages of gestation, combined with a difference in transmission rate depending on the gestation stage. The high transmission rate found at mid term gestation (69%) makes our model very suitable for further research into the mechanisms of transplacental transmission and for research into the reduction of this route of transmission through vaccination.     

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.     

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.     

Quantitative analysis of transmission parameters for bluetongue virus serotype 8 in Western Europe in 2006

ABSTRACT: The recent bluetongue virus serotype 8 (BTV-8) epidemic in Western Europe struck hard. Controlling the infection was difficult and a good and safe vaccine was not available until the spring of 2008. Little was known regarding BTV transmission in Western Europe or the efficacy of control measures. Quantitative details on transmission are essential to assess the potential and efficacy of such measures.To quantify virus transmission between herds, a temporal and a spatio-temporal analysis were applied to data on reported infected herds in 2006. We calculated the basic reproduction number between herds (Rh: expected number of new infections, generated by one initial infected herd in a susceptible environment). It was found to be of the same order of magnitude as that of an infection with Foot and Mouth Disease (FMD) in The Netherlands, e.g. around 4. We concluded that an average day temperature of at least 15°C is required for BTV-8 transmission between herds in Western Europe. A few degrees increase in temperature is found to lead to a major increase in BTV-8 transmission.We also found that the applied disease control (spatial zones based on 20 km radius restricting animal transport to outside regions) led to a spatial transmission pattern of BTV-8, with 85% of transmission restricted to a 20 km range. This 20 km equals the scale of the protection zones. We concluded that free animal movement led to substantial faster spread of the BTV-8 epidemic over space as compared to a situation with animal movement restrictions.     

Toggenburg Orbivirus, a new bluetongue virus: Initial detection, first observations in field and experimental infection of goats and sheep

A novel bluetongue virus termed “Toggenburg Orbivirus” (TOV) was detected in two Swiss goat flocks. This orbivirus was characterized by sequencing of 7 of its 10 viral genome segments. The sequencing data revealed that this virus is likely to represent a new serotype of bluetongue virus [Hofmann, M.A., Renzullo, S., Mader, M., Chaignat, V., Worwa, G., Thuer, B., 2008b. Genetic characterization of Toggenburg Orbivirus (TOV) as a tentative 25th serotype of bluetongue virus, detected in goats from Switzerland. Emerg. Infect. Dis. 14, 1855–1861].In the field, no clinical signs were observed in TOV-infected adult goats; however, several stillborn and weak born kids were reported. Although born during a period of extremely low vector activity, one of these kids was found to be antibody and viral genome positive and died 3.5 weeks postpartum.Experimental infection of goats and sheep, using TOV-positive field blood samples, was performed to assess the pathogenicity of this virus.Goats did not show any clinical or pathological signs, whereas in sheep mild bluetongue-like clinical signs were observed. Necropsy of sheep demonstrated bluetongue-typical hemorrhages in the wall of the pulmonary artery. Viral RNA was detected in organs, e.g. spleen, palatine tonsils, lung and several lymph nodes of three experimentally infected animals.Unlike other bluetongue virus serotypes, it was not possible to propagate the virus, either from naturally or experimentally infected animals in any of the tested mammalian or insect cell lines or in embryonated chicken eggs.In small ruminants, TOV leads to mild bluetongue-like symptoms. Further investigations about prevalence of this virus are needed to increase the knowledge on its epidemiology.     

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.     

Transplacental BVD virus transmission after experimental inoculation of goats in different pregnancy stages

Eight groups of altogether 25 goats without neutralizing antibodies against BVD virus, were inoculated either intranasally or intranasally and subcutaneously with two different BVD virus isolates during different stages of gestation. In all 18 goats inoculated within the first 78 days of gestation an abortion and foetal death rate of approximately 100% occurred. Only one goat gave birth to a clinically healthy kid. The other seven goats which were inoculated after the 78th day of gestation showed also a high foetal death rate. Only two of them gave birth to clinically healthy kids. Neutralizing antibodies against BVD virus could be detected in blood samples drawn from 14 kids born at normal term including stillborn and non-viable offsprings. BVD virus was reisolated from different organs taken from seven foetuses. It was not possible to isolate BVD virus from any of the normal offsprings.     

Probability of introduction of exotic strains of bluetongue virus into the US and into California through importation of infected cattle

Strategies designed to minimize the probability of bluetongue virus (BTV) introduction to new areas should be based on a quantitative assessment of the probability of actually establishing the virus once it is introduced. The risk of introducing a new strain of bluetongue virus into a region depends on the number of viremic animals that enter and the competency of local vectors to transmit the virus. We used Monte Carlo simulation to model the probability of introducing BTV into California, USA, and the US through importation of cattle. Records of cattle and calf imports into California and the US were obtained, as was seroprevalence information from the exporting countries. A simulation model was constructed to evaluate the probability of importing either a viremic PCR-negative animal after 14-day quarantine, a c-ELISA BTV-antibody-negative animal after 28-day quarantine, or an untested viremic animal after 100-day quarantine into California and into the US. We found that for animals imported to the US, the simulated (best to worst scenarios) median percentage that tested positive for BTV-antibody ranged from 5.4 to 7.2%, while for the subset imported to California, the simulated median percentage that tested positive for BTV-antibody ranged from 20.9 to 78.9%. Using PCR, for animals imported to the US these values were 71.8–85.3%, and for those imported to California, the simulated median that test positive ranged from 74.3 to 92.4%. The probability that an imported animal was BTV-viremic is very low regardless of the scenario selected (median probability = 0.0%). The probability of introducing an exotic strain of BTV into California or the US by importing infected cattle was remote, and the current Office International des Epizooties (OIE) recommendation of either a final PCR test performed 14 days after entry into quarantine, a c-ELISA performed 28 days after entry into quarantine or a 100-day quarantine with no testing requirement was adequate to protect cattle in the US and California from an exotic strain of BTV.     

Field observations during the bluetongue serotype 8 epidemic in 2006. I. Detection of first outbreaks and clinical signs in sheep and cattle in Belgium, France and the Netherlands

Starting August 2006, a major epidemic of bluetongue (BT) was identified in North-West Europe, affecting The Netherlands, Belgium, Germany, Luxemburg and the North of France. It was caused by BT virus serotype 8 (BTV-8), a serotype previously unknown to the European Union (EU). In this outbreak, the virus caused clinical disease in a few individual animals within cattle herds, whereas overt clinical disease was usually restricted to sheep. Investigations in Belgium suggested that the first clinical signs of BTV-8 appeared mid July 2006 in a cattle herd, while the first suspicion of a BT-outbreak in Belgium was reported on 17 August 2006. In the first 10 BTV-8 outbreaks in the Netherlands, the owners indicated that the first clinical signs started approximately 12-17 days before a suspicion was reported to the veterinary authorities via a veterinary practitioner. In BTV-8 affected sheep flocks, erosions of the oral mucosa, fever, salivation, facial and mandibular oedema, apathy and tiredness, mortality, oedema of the lips, lameness, and dysphagia were among the most frequent clinical signs recorded. The most prominent clinical signs in BTV-8 affected cattle herds were: crusts/lesions of the nasal mucosa, erosions of lips/crusts in or around the nostrils, erosions of the oral mucosa, salivation, fever, conjunctivitis, coronitis, muscle necrosis, and stiffness of the limbs. Crusts/lesions of nasal mucosa, conjunctivitis, hyperaemic/purple coloration and lesions of the teats, and redness/hypersensitivity of the skin were relatively more seen on outbreak farms with cattle compared to sheep. Mortality, oedema of the head and ears, coronitis, redness of the oral mucosa, erosions/ulceration of tongue mucosa, purple coloration of the tongue and tongue protrusion and dyspneu were relatively more seen on outbreak farms with sheep compared to cattle.