Indoor activity of Culicoides associated with livestock in the bluetongue virus (BTV) affected region of northern France during autumn 2006

In August 2006, bluetongue virus (BTV) was detected in the Netherlands, Belgium, western Germany, Luxembourg and northern France for the first time. Consequently, a longitudinal entomological study was conducted in the affected region of northern France (Ardennes) throughout the autumn of 2006. Data on the spatio-temporal distribution of Culicoides (Diptera: Ceratopogonidae) associated with livestock were collected and an attempt was made to identify the vector(s) involved in BTV transmission by means of virus detection in wild-caught biting midges. Weekly sampling using standardized Onderstepoort-type blacklight traps were performed simultaneously both outdoors and indoors in one BTV-free and three BTV-affected farms between September and December 2006. Culicoides were sorted according to farm, location (outdoors vs. indoors), time point (in weeks), species and physiological stage. BTV detection was conducted by RT-PCR on monospecific pools of non-bloodfed parous female Culicoides. The principal results showed: (i) the absence of the Mediterranean vector, C. imicola, (ii) the relatively low abundance of C. dewulfi and C. pulicaris, (iii) the widespread occurrence and abundance of C. obsoletus/C. scoticus with longevity and behaviour compatible with BTV transmission, and (iv) all Culicoides pools tested for BTV were negative. In France, the very low levels of BTV-8 circulation were probably due to the limited introduction of the virus from affected neighbouring countries, and not due to the absence of local vector populations. A key finding has been the substantiation, for the first time, that Culicoides, and particularly the potential vectors C. obsoletus/C. scoticus and C. dewulfi, can be active at night inside livestock buildings and not only outside, as originally believed. The endophagic tendencies of members of the Obsoletus group are discussed in light of the prolonged period of BTV transmission during the autumn of 2006 and the risk of BTV overwintering and resurgence in the spring of 2007. Overall, there is an urgent need to improve our knowledge on the ecology of local Culicoides species before any clear, effective and reliable recommendations can be provided to the veterinary authorities in terms of prevention and control.     

The 2006 outbreak of bluetongue in northern Europe--the entomological perspective

After bluetongue (BT) appeared in northern Europe in August 2006 entomological studies were implemented in all five affected Member States (MSs) to establish which species of Culicoides had acted as vectors. The findings can be summarised as follows: (i) C. imicola the principal southern European/African vector of BTV has not penetrated into northern Europe, (ii) three pools of C. obsoletus/C. scoticus and one of C. dewulfi assayed RT-PCR-positive to BTV-8, (iii) in support of these results it was found that both potential vectors had also high parity rates (approximately 40%) indicating increased longevity favouring BTV virogenesis and transmission, (iv) furthermore, C. obsoletus/C. scoticus and C. dewulfi occurred also widely and abundantly on sheep and cattle holdings across the entire affected region, (v) and during the latter part of the season showed strong endophily readily entering livestock buildings in significant numbers to bite the animals inside (endophagy), (vi) which demonstrates that housing at best offers only limited protection to livestock from Culicoides attacks, (vii) in contrast the potential vector C. pulicaris sensu stricto was restricted geographically, was captured rarely, had a low parity rate (10%) and was exophilic indicating it played no role in the outbreak of BT, (viii) the incrimination of C. dewulfi as a novel vector is significant because it breeds in cattle and horse dung this close association raising its vectorial potential, but (ix) problems with its taxonomy (and that of the Obsoletus and Pulicaris species complexes) illustrates the need for morphological and molecular techniques to become more fully integrated to ensure progress in the accurate identification of vector Culicoides, (x) midge densities (as adjudged by light traps) were generally low indicating northern European Culicoides to have a high vector potential and/or that significant numbers of midges are going undetected because they are biting (and transmitting BTV) during the day when light traps are not effective, and (xi) the sporadic capture of Culicoides in the winter of 2007 invites re-examination of the current definition of a vector-free period. The re-emergence of BT over a wide front in 2007 raises anew questions as to precisely how the virus overwinters and asks also that we scrutinise our monitoring systems in terms of their sensitivity and early warning capability.     

Seasonal dynamics of biting midges (Diptera: Ceratopogonidae: Culicoides), the potential vectors of bluetongue virus, in Sweden

The outbreak of bluetongue (BT) in northern Europe 2006 initiated the monitoring of vectors, biting midges of the genus Culicoides in Sweden. In order to determine the diversity, distribution and seasonal dynamics of Culicoides, weekly collections were made during 2008 and during March-December 2009 using the Ondestepoort Veterinary Institute black light trap. Twenty sampling sites were selected in 12 provinces. In total of 30,704 Culicoides were collected in 2008 and 32,252 in 2009. The most abundant species were the potential vectors of BTV Culicoides obsoletus/C. scoticus that comprised of 77% of the total catches. Other biting midges collected were Culicoides impunctatus (9%), Culicoides grisescens (3%), Culicoides punctatus (2%), Culicoides chiopterus (2%) and Culicoides pulicaris (2%). Culicoides obsoletus/C. scoticus were most abundant during May-June and August-September. The majority of the species were active from March to November in 2008 and April to October in 2009. Species considered as potential vectors of bluetongue virus (BTV) occurred as far north as latitude 65°N (Kalix).     

Culicoides trapping with Rothamsted suction traps before and during the bluetongue epidemic of 2006 in Belgium

The collection of biting midges was taking place some months before the first bluetongue outbreak in Belgium in August 2006. The Walloon Agricultural Research Centre had been monitoring aphid populations at two sites annually in Belgium (Gembloux and Libramont), using two stationary '12-m' Rothamsted suction traps. For the Gembloux trap, collections of insects captured daily from 11 May 2006 onwards were already available at the time of the outbreak. An examination of these samples revealed the presence of Culicoides, some species of which are considered as potential vectors of the bluetongue virus (BTV). The trapping was therefore extended beyond the normal aphid activity period and the Culicoides captured were identified to species level. From 11 May to 31 December 2006, the Gembloux trap caught 664 Culicoides specimens belonging to 19 species comprising known BTV-vectors. The second trap, at Libramont, was reactivated from 12 September to 13 October and caught 97 specimens belonging to nine species, all of which had been found at the Gembloux site. Among the 19 species identified, four were new to Belgian fauna: Culicoides achrayi, C. deltus, C. lupicaris and C. newsteadi. This paper examines the overall phenology and the physiological status of Culicoides in 2006 before and during the bluetongue epidemic. It discusses the potential of the Rothamsted suction trap to monitor Culicoides.     

Indoor activity of Culicoides associated with livestock in the bluetongue virus (BTV) affected region of northern France during autumn 2006

In August 2006, bluetongue virus (BTV) was detected in the Netherlands, Belgium, western Germany, Luxembourg and northern France for the first time. Consequently, a longitudinal entomological study was conducted in the affected region of northern France (Ardennes) throughout the autumn of 2006. Data on the spatio-temporal distribution of Culicoides (Diptera: Ceratopogonidae) associated with livestock were collected and an attempt was made to identify the vector(s) involved in BTV transmission by means of virus detection in wild-caught biting midges. Weekly sampling using standardized Onderstepoort-type blacklight traps were performed simultaneously both outdoors and indoors in one BTV-free and three BTV-affected farms between September and December 2006.Culicoides were sorted according to farm, location (outdoors vs. indoors), time point (in weeks), species and physiological stage. BTV detection was conducted by RT-PCR on monospecific pools of non-bloodfed parous female Culicoides.The principal results showed: (i) the absence of the Mediterranean vector, C. imicola, (ii) the relatively low abundance of C. dewulfi and C. pulicaris, (iii) the widespread occurrence and abundance of C. obsoletus/C. scoticus with longevity and behaviour compatible with BTV transmission, and (iv) all Culicoides pools tested for BTV were negative.In France, the very low levels of BTV-8 circulation were probably due to the limited introduction of the virus from affected neighbouring countries, and not due to the absence of local vector populations. A key finding has been the substantiation, for the first time, that Culicoides, and particularly the potential vectors C. obsoletus/C. scoticus and C. dewulfi, can be active at night inside livestock buildings and not only outside, as originally believed.The endophagic tendencies of members of the Obsoletus group are discussed in light of the prolonged period of BTV transmission during the autumn of 2006 and the risk of BTV overwintering and resurgence in the spring of 2007. Overall, there is an urgent need to improve our knowledge on the ecology of local Culicoides species before any clear, effective and reliable recommendations can be provided to the veterinary authorities in terms of prevention and control.     

The phenology and population dynamics of Culicoides spp. in different ecosystems in The Netherlands

The Netherlands has enjoyed a relatively free state of vector-borne diseases of economic importance for more than one century. Emerging infectious diseases may change this situation, threatening the health of humans, domestic livestock and wildlife. In order to be prepared for the potential outbreak of vector-borne diseases, a study was undertaken to investigate the distribution and seasonal dynamics of candidate vectors of infectious diseases with emphasis on bluetongue vectors (Culicoides spp.). The study focused primarily on the relationship between characteristic ecosystems suitable for bluetongue vectors and climate, as well as on the phenology and population dynamics of these vectors. Twelve locations were selected, distributed over four distinct habitats: a wetland area, three riverine systems, four peat land areas and four livestock farms. Culicoides populations were sampled continuously using CO(2)-baited counterflow traps from July 2005 until August 2006, with an interruption from November 2005 to March 2006. All vectors were identified to species level. Meteorological and environmental data were collected at each location. Culicoides species were found in all four different habitat types studied. Wetland areas and peat bogs were rich in Culicoides spp. The taxonomic groups Culicoides obsoletus (Meigen) and Culicoides pulicaris (Linnaeus) were strongly associated with farms. Eighty-eight percent of all Culicoides consisted of the taxon C. obsoletus/Culicoides scoticus. On the livestock farms, 3% of Culicoides existed of the alleged bluetongue vector Culicoides dewulfi Goetghebuer. Culicoides impunctatus Goetghebuer was strongly associated with wetland and peat bog. Many Culicoides species were found until late in the phenological season and their activity was strongly associated with climate throughout the year. High annual variations in population dynamics were observed within the same study areas, which were probably caused by annual variations in environmental conditions. The study demonstrates that candidate vectors of bluetongue virus are present in natural and livestock-farm habitats in the Netherlands, distributed widely across the country. Under favourable climatic conditions, following virus introduction, bluetongue can spread among livestock (cattle, sheep and goats), depending on the nature of the viral serotype. The question now arises whether the virus can survive the winter conditions in north-western Europe and whether measures can be taken that effectively halt further spread of the disease.     

Culicoides vectors of bluetongue virus in Chester Zoo

On four nights in June 2008, light traps were operated for Culicoides biting midges, the vector species for bluetongue virus (BTV), at five sites in Chester Zoo in north-west England. Over 35,000 Culicoides midges, of 25 species, were captured, including high densities inside animal enclosures. Over 94 per cent of all the Culicoides trapped were females of the Obsoletus group, which is implicated as the vector of BTV serotype 8 in northern Europe. The mean catch of this group per trap per night was over 1500, suggesting a potential risk of BTV transmission if the virus is introduced to Chester Zoo in the animals or midges in the summer.     

Efficacy of Flectron-eartags (cypermethrin) for control of midges (Culicoides) as the vectors of bluetongue virus in cattle: field studies and biossays

When in 2006 infection with bluetongue for the first time occurred in Germany the registered and already against flies and tabanids in cattle proofed Flectron ear tags were used against the blood feeding vector midges (Culicoides) also. However, the efficacy against gnats was not yet proofed. The efficacy of 1 and 2 ear tags (1,067 g cypermethrin per ear tag) per animal was investigated in North Germany with 237 heifers and dairy cows. Midges were caught in suction light traps close to the cattle on pasture or became trapped by mouth operated aspirators directly at the skin of the animal bodies. Within 12,051 specimens of midges 12 species of Culicoides could be identified. On grasslands 3 species, C. obsoletus, C. pulicaris and C. dewulfi were found to be dominant. These 3 species are also known to be vectors of BTV. The toxic efficacy was found for 14 days with 1 ear tag and up to 21 days with 2 ear tags. This duration of efficacy was confirmed in the laboratory with hair clippings from the dorsal line and the ventral abdomen (bioassay). In accordance with workers in the U.S.A. it is concluded that insecticide-impregnated ear tags will reduce the number of biting midges, and by this way the risk of infection with BTV in herds of treated cattle will be reduced as well as in other cattle of a particular region. It is concluded that ear tags are of considerable value as part of an integrated control program for BT, e.g. vaccination.     

A stochastic predictive model for the natural spread of bluetongue

In recent years the vector-borne diseases (VBD) are (re)-emerging and spreading across the world having a profound impact on human and veterinary health, ecology, socio-economics and disease management. Arguably the best-documented example of veterinary importance is the recent twofold invasion of bluetongue (BT) in Europe. Much attention has been devoted to derive presence-absence habitat distribution models and to model transmission through direct contact. Limited research has focused on the dynamic modelling of wind mediated BT spread. This paper shows the results of a stochastic predictive model used to assess the spread of bluetongue by vectors considering both wind-independent and wind-mediated movement of the vectors. The model was parameterised using epidemiological knowledge from the BTV8 epidemic in 2006/2007 and the BTV1 epidemic in 2008 in South-France. The model correctly reflects the total surface of the infected zone (overall accuracy=0.77; sensitivity=0.94; specificity=0.65) whilst slightly overestimating spatial case density. The model was used operationally in spring 2009 to predict further spread of BTV1. This allowed veterinary officers in Belgium to decide whether there was a risk of introduction of BTV1 from France into Belgium and thus, whether there was a need for vaccination. Given the far distance from the predicted infected zone to the Belgian border, it was decided not to vaccinate against BTV1 in 2009 in Belgium.     

A wind density model to quantify the airborne spread of Culicoides species during north-western Europe bluetongue epidemic, 2006

Increased transport and trade as well as climate shifts play an important role in the introduction, establishment and spread of new pathogens. Arguably, the introduction of bluetongue virus (BTV) serotype 8 in Benelux, Germany and France in 2006 is such an example. After its establishment in receptive local vector and host populations the continued spread of such a disease in a suitable environment will mainly depend on movement of infected vectors and animals. In this paper we explore how wind models can contribute to explain the spread of BTV in a temperate eco-climatic setting. Based on previous work in Greece and Bulgaria filtered wind density maps were computed using data from the European Centre for Medium-Range Weather Forecasts (ECMWF). Six hourly forward wind trajectories were computed at pressure levels of 850hPa for each infected farm as from the recorded onset of symptoms. The trajectories were filtered to remove wind events that do not contribute to possible spread of the vector. The suitable wind events were rastered and aggregated on a weekly basis to obtain weekly wind density maps. Next to this, cumulated wind density maps were also calculated to assess the overall impact of wind dispersal of vectors. A strong positive correlation was established between wind density data and the horizontal asymmetrical spread pattern of the 2006 BTV8 epidemic. It was shown that short (<5km), medium (5-31km) and long (>31km) distance spread had a different impact on disease spread. Computed wind densities were linked to the medium/long-distance spread whilst short range spread was mainly driven by active Culicoides flight. Whilst previous work in the Mediterranean basin showed that wind driven spread of Culicoides over sea occurred over distances of up to 700km, this phenomenon was not observed over land. Long-distance spread over land followed a hopping pattern, i.e. with intermediary stops and establishment of local virus circulation clusters at distances of 35-85km. Despite suitable wind densities, no long range spread was recorded over distances of 300-400km. Factors preventing spread Eastwards to the UK and Northwards to Denmark during the 2006 epidemic are discussed. Towards the east both elevation and terrain roughness, causing air turbulences and drop down of Culicoides, were major factors restricting spread. It is concluded that the proposed approach opens new avenues for understanding the spread of vector-borne viruses in Europe. Future developments should take into consideration both physical and biological factors affecting spread.     

Vector monitoring at Belgian outbreak sites during the bluetongue epidemic of 2006

In response to the first bluetongue outbreak in Belgium a monitoring programme was started at the end of August 2006 to identify possible vectors transmitting the disease. Black light traps were deployed at 36 outbreak sites and captured 1959 Culicoides specimens belonging to 16 different species. Eighty four percent of the biting midges captured belonged to the C. obsoletus complex, among them C. obsoletus s.s., C. dewulfi and C. scoticus, three suspected bluetongue vectors. The Veterinary and Agrochemical Research Centre detected viral RNA in pools of individuals belonging to this complex. Culicoides pulicaris, a potential bluetongue vector in Italy, should yet not be excluded as a possible vector in Belgium as this species was frequently found around outbreak sites, notwithstanding this species is not easily captured with the trapping techniques used during this survey.     

The vector potential of British Culicoides species for bluetongue virus

Two species of British Culicoides, C. nubeculosus and C. impunctatus were found to support bluetongue virus (BTV) multiplication after ingestion of the virus. Both species were infected by membrane feeding and C. nubeculosus also became infected after feeding on viraemic sheep. This species was shown to transfer the virus across a membrane after 8 days incubation at 25 degrees C and could therefore presumably act as a BTV vector. Six other British species of Culicoides supported BTV multiplication after intrathoracic inoculation of the virus.     

Bluetongue and epizootic hemorrhagic disease virus isolation from vertebrate and invertebrate hosts at a common geographic site

One serotype of bluetongue virus (BTV) and two serotypes of epizootic hemorrhagic disease virus (EHDV) were isolated from vertebrate and invertebrate hosts on a farm in Colorado. The isolations were from blood samples collected a week apart from a dairy heifer with stomatitis and laminitis; EHDV serotypes 1 and 2 were isolated from the first blood sample, and BTV serotype 13 and EHDV serotype 1 were isolated from the second. Antibodies to EHDV and BTV were detected in the serum from this heifer. Both EHDV serotypes and BTV serotype 13 were isolated from pools of female biting gnats (Culicoides variipennis) that had not had a recent blood meal. The BTV insect isolate was biologically transmitted by female gnats from an infected donor sheep to a recipient host sheep. Culicoides variipennis was the predominant insect collected during three nights of light trap captures at the farm.     

环状病毒的流行与传播

环状病毒是牲畜常见的重要病原体,主要包括有蓝舌病病毒、非洲马瘟病毒、马器质性脑病病毒和流行性出血热病毒等。这些病毒能够通过吸血性的库蠓传播。本文主要介绍了这几种病毒在世界各地的流行与传播情况。     

In vivo and in vitro propagation and transmission of Toggenburg orbivirus

The Toggenburg orbivirus (TOV), a recently discovered virus related to bluetongue virus (BTV), has been identified in goats in Switzerland, Italy and Germany. Isolation of TOV in vitro has not yet been achieved and the transmission mechanisms are still unknown. In the experimental infection of pregnant goats described here, TOV could not be detected in secretion/excretion samples or fetal blood. Material from the goat experiment was used as inoculum for propagating the virus in vitro. To enhance the infectivity of TOV several modified protocols, e.g. pretreatment of the virus with trypsin, polyethylene glycol-mediated infection and lipofection were applied. Isolation of TOV, attempts to infect Culicoides nubeculosus by feeding TOV-positive blood and intracerebral inoculation of newborn mice were unsuccessful. The results of these studies suggest that TOV requires specific but different factors than other BTVs for infection and replication outside of its natural caprine host.     

No evidence for involvement of sheep in the epidemiology of cattle virulent epizootic hemorrhagic disease virus

Epizootic hemorrhagic disease virus (EHDV) is an Orbivirus. While not previously considered as an important disease in cattle, several EHDV serotypes (EHDV-6 and 7) have recently been implicated in disease outbreaks. The involvement of sheep in the epidemiology of EHDV is still not understood. In this study we compared the prevalence of antibodies to EHDV and bluetongue virus (BTV) in sheep to their prevalence in cattle after an outbreak of EHDV that occurred in Israel during 2006. Sixty-six sheep and lambs scattered in seven herds were compared to 114 cows and calves scattered in 13 dairy cattle herds, matched to the sheep herds by location. While antibody prevalence to EHDV was high in cattle (35.2% within the outbreak zone) no evidence of exposure to EHDV was found in sheep (p<0.0001). Antibodies to BTV were apparent in both cattle and sheep though in the former it was significantly higher (63.2%, 16.7% respectively, p<0.0001), suggesting higher exposure of cattle to biting Culicoides midges. Taken together, these results imply that sheep have a negligible role in the epidemiology of EHDV.     

The phenology and population dynamics of Culicoides spp. in different ecosystems in The Netherlands

The Netherlands has enjoyed a relatively free state of vector-borne diseases of economic importance for more than one century. Emerging infectious diseases may change this situation, threatening the health of humans, domestic livestock and wildlife. In order to be prepared for the potential outbreak of vector-borne diseases, a study was undertaken to investigate the distribution and seasonal dynamics of candidate vectors of infectious diseases with emphasis on bluetongue vectors (Culicoides spp.). The study focused primarily on the relationship between characteristic ecosystems suitable for bluetongue vectors and climate, as well as on the phenology and population dynamics of these vectors.Twelve locations were selected, distributed over four distinct habitats: a wetland area, three riverine systems, four peat land areas and four livestock farms. Culicoides populations were sampled continuously using CO₂-baited counterflow traps from July 2005 until August 2006, with an interruption from November 2005 to March 2006. All vectors were identified to species level. Meteorological and environmental data were collected at each location.Culicoides species were found in all four different habitat types studied. Wetland areas and peat bogs were rich in Culicoides spp. The taxonomic groups Culicoides obsoletus (Meigen) and Culicoides pulicaris (Linnaeus) were strongly associated with farms. Eighty-eight percent of all Culicoides consisted of the taxon C. obsoletus/Culicoides scoticus. On the livestock farms, 3% of Culicoides existed of the alleged bluetongue vector Culicoides dewulfi Goetghebuer. Culicoides impunctatus Goetghebuer was strongly associated with wetland and peat bog. Many Culicoides species were found until late in the phenological season and their activity was strongly associated with climate throughout the year. High annual variations in population dynamics were observed within the same study areas, which were probably caused by annual variations in environmental conditions.The study demonstrates that candidate vectors of bluetongue virus are present in natural and livestock-farm habitats in the Netherlands, distributed widely across the country. Under favourable climatic conditions, following virus introduction, bluetongue can spread among livestock (cattle, sheep and goats), depending on the nature of the viral serotype. The question now arises whether the virus can survive the winter conditions in north-western Europe and whether measures can be taken that effectively halt further spread of the disease.     

Estimating front-wave velocity of infectious diseases: a simple, efficient method applied to bluetongue

ABSTRACT: Understanding the spatial dynamics of an infectious disease is critical when attempting to predict where and how fast the disease will spread. We illustrate an approach using a trend-surface analysis (TSA) model combined with a spatial error simultaneous autoregressive model (SARerr model) to estimate the speed of diffusion of bluetongue (BT), an infectious disease of ruminants caused by bluetongue virus (BTV) and transmitted by Culicoides. In a first step to gain further insight into the spatial transmission characteristics of BTV serotype 8, we used 2007-2008 clinical case reports in France and TSA modelling to identify the major directions and speed of disease diffusion. We accounted for spatial autocorrelation by combining TSA with a SARerr model, which led to a trend SARerr model. Overall, BT spread from north-eastern to south-western France. The average trend SARerr-estimated velocity across the country was 5.6 km/day. However, velocities differed between areas and time periods, varying between 2.1 and 9.3 km/day. For more than 83% of the contaminated municipalities, the trend SARerr-estimated velocity was less than 7 km/day. Our study was a first step in describing the diffusion process for BT in France. To our knowledge, it is the first to show that BT spread in France was primarily local and consistent with the active flight of Culicoides and local movements of farm animals. Models such as the trend SARerr models are powerful tools to provide information on direction and speed of disease diffusion when the only data available are date and location of cases.     

Seasonal abundance and parity of Culicoides biting midges associated with livestock at Roma, Lesotho (Diptera: Ceratopogonidae)

A light-trap survey was undertaken of the species composition, seasonal abundance and parity of Culicoides at Roma, Lesotho, to establish whether the likely vectors for bluetongue and African horse sickness occur in this area as well as the chance of transmission. A total of 34 catches was made between 21 September 1985 and 24 September 1986; 32,819 Culicoides were caught belonging to 19 species. Culicoides numbers rapidly built up from December to a peak in February which implies that this may also be the optimum time for virus transmission. The number of Culicoides dropped sharply in April with the onset of cooler conditions. C. zuluensis was the dominant species forming 69.6% of the totalled catches, followed by C. pycnostictus with 11.7%. C. imicola, the only proven vector of bluetongue, was never abundant representing only 4.4% of the midges caught. The parous rate for each of the 2 commonest species was low, implying a low vector capacity.     

Genetic variation of bluetongue virus serotype 11 isolated from host (sheep) and vector (Culicoides variipennis) at the same site

Genetic relatedness of 2 strains of bluetongue virus (BTV) serotype 11 that were isolated from the same geographic site--one from host (sheep) and the other from the vector Culicoides variipennis during an enzootic of bluetongue at Bruneau, Idaho, in August 1973--was determined by comparing the oligonucleotide fingerprint analyses of the individual double-stranded RNA segments of the genomes. It was observed that the 2 strains of BTV-11 exhibit considerable differences in their genotypes, the percentage of diversity being different for each of the corresponding RNA species of the 2 strains of BTV-11. These results indicate that more than one genotype of BTV can circulate in juxtaposition in a given geographic site. The observed genotypic diversity might be due to the accumulation of point mutations on specific RNA species or antecedent reassortment of RNA segments between different BTV in nature or both.