Control Tools

• Diagnostics availability

• Commercial diagnostic kits available worldwide

  Many different antibody detection ELISA kits are available commercially. These include competitive and double antigen ELISAs.Real-time RT-PCR detection assays (group specific) are available commercially from many companies.Real-time RT-PCR serotyping assays for European serotypes 1, 2, 4, 6, 8, 9, 11, 16, 25 are also available commercially.

  List of commercially available diagnostic kits (Diagnostics for Animals)

• Commercial diagnostic kits available in Europe

  Many different antibody detection ELISA kits are available commercially. These include competitive and double antigen ELISAs.Real-time RT-PCR detection assays (group specific) are available commercially from many companies.Real-time RT-PCR serotyping assays for European types 1, 2, 4, 6, 8, 9, 11, 16, 25 are also available commercially.

  List of commercially available diagnostic kits (Diagnostics for Animals)

  GAPS:

  • Real-time RT-PCR assays are needed for remaining serotypes. Although these are currently under development, further access to variants of each serotype (from around the world) and further validation are required.
  • Real-time and conventional RT-PCR assays have been developed for all BTV serotypes. However, they require further evaluation and their commercial availability depends on demand.
  • These assays will potentially need updating as new isolates/variants are detected in the field.
  • Other diagnostic systems could potentially also be developed including a RNA microarray and a single tube/bead/chip based typing assay.

• Diagnostic kits validated by International, European or National Standards

  Host: Agar-gel-immunodiffusion and competitive ELISA; virus neutralization test.Pathogen: virus isolation in embryonated chicken eggs and insect or mammalian cell cultures. Virus neutralisation test.Real-time and gel based RT-PCR for viral genome detection

  GAPS:

  • RT-PCR amplification and sequence analyses, followed by phylogenetic comparison to existing datasets, gives the most accurate identification of each virus strain and can identify reassortants strains, which may be biologically important. It is used to provide important data on individual outbreak strains.
  • Next Generation Sequencing (NGS) is becoming the most prevalent sequencing technique. Harmonization of procedures devoted to amplify the full-length of each genome segments of BTV isolates or straight from biological sample by NGS is required.

• Diagnostic method(s) described by International, European or National standards

  Host: competitive ELISA; virus neutralization test.Pathogen: virus isolation in embryonating chicken eggs and cell culture. Virus neutralisation test. Real-time and gel based RT-PCR for viral genome detection.

  GAP:

  RT-PCR amplification and sequence analyses , followed by phylogenetic comparison to existing datasets, gives the most accurate identification of virus strain that is possible. Although this technology may be unsuited to routine diagnosis it is used to provide important data on individual outbreak strains

• Commercial potential for diagnostic kits in Europe

  Test systems for antibody detection are commercially available. No antigen detection tests are commercially available.Bluetongue group specific real-time RT- PCR tests are commercially available. Type specific real-time RT- PCR tests are available for European types (1, 2, 4, 6, 8, 9, 11, 16 and 25). There is potential for the remaining types (under development).Real time RT- specific PCR for BTV-3 and other serotypes. Type specific c-ELISA

  GAP:

  Real-time RT-PCR assays are needed for remaining types. Although these are currently under development, further access to variants (topotypes) of each serotype (from around the world) and further validation are required.

• DIVA tests required and/or available

  No serological DIVA tests are currently available but are needed for international trade in animals, and as an important part of control measures to distinguish infected from vaccinated animals when there is widespread vaccination.RT-PCR assays can be used in this way in combination with inactivated vaccines. In particular in surveillance programs and longitudinal studies, RT-PCR-diagnostics can be very beneficial. These assays have played a very important role in surveillance during recent vaccination campaigns with inactivated vaccines in Europe. A continued process for the development of more sensitive tests (with the possibility of pooling) and automation/robotizing makes this technique very reliable and competitive to ELISAs.The live virus vaccines generate NS proteins and the current inactivated vaccines also contain NS-proteins, so it may therefore be difficult or even impossible to develop reliable DIVA assays for either of these vaccine categories. Next generation vaccines allow DIVA strategies / assays.

  GAPS:

  • DIVA assay required, and need to be used in combination with DIVA vaccines, when available.
  • There is a need for serological as well as molecular DIVA assays
  • Currently RT-PCR assays can be used to identify animals that do not represent a transmission risk.

• Opportunities for new developments

  GAPS:

  • Further development of existing real-time assays, may be required to maintain effectiveness to detect new BTV isolates/variants.
  • Further validation of RT-PCR assays is required for the serotype-specific real-time RT-PCR assays that are designed detect specifically the remaining BTV types.
  • Initial studies indicate that chip based technologies to detect viral RNA, can be used to identify members of the BTV group and each serotype. However, commercialisation will depend on cost and ease of use. This technology will need further development.
  • It may be possible to develop a pen-side assay for BTV. Although in view of the relatively long term nature of BTV outbreaks this may be more important for differential diagnosis (e.g. compared to FMDV) than directly valuable as a tool against BTV.
  • By expressing individual BTV VP2 proteins from different serotypes, and generating either polyclonal or monoclonal type-specific antibodies to them, it may be possible to develop ‘type-specific ELISA’ to detect antibodies to each BTV serotype. These may be useful to track movements of new types in areas that have previously been vaccinated against an existing serotype.
  • Micro-arrays technology in order to identify circulating BTVs on a genome segment level.

• Vaccines availability

• Commercial vaccines availability (globally)

  Live attenuated vaccines:Available and produced in India, Turkey, South Africa, North Africa, Russia and Europe. Modified virus vaccines are cheap, easy to produce. A single dose is usually effective in controlling clinical BTV outbreaks in areas of endemic disease and in the face of outbreaks.

  Traditional attenuated vaccines bare the risk of transmission and reassortment (exchange of genetic segments), when used during the vector season and do not conform with the European Pharmacopeia.Vaccines against almost all BTV serotypes are available. Multi-valent (up to penta) vaccines are available.None of the attenuated vaccines have a European Marketing Authorisation. Some countries issue a ‘Temporary Authorisation for Use’ (TAU), which isn’t a license but a permission to use. TAU’s are valid for one year with possibility for renewal (see below).

  Inactivated vaccinesProduced by a number of companies in Europe. Some contain partially purified and inactivated virus.

  GAPS:

  • Live vaccines:LicenseConformity with European PharmacopeiaNew serotypes do not have vaccines available. Cross reactivity of vaccines needs further development/studySafetyProduction capacity in EuropeQuality standards
  • Inactivated vaccinesEuropean Commission long-term planning of management of antigen and vaccine stocks (see also vaccine bank)Also: see licensingNo subunit vaccines commercially available as yet.Needed: Incentives for producers to develop and produce ‘ahead’ of crisis. Vaccine producers need incentives to develop, test and produce vaccine for a non-existing market.
  • Subunit vaccines are not yet commercially available, but have been developed and concept proven experimentally.
  • Other possibilities based on the recently developed reverse genetics.Novel antigen delivery methods, e.g. recombinant virus carriers. DNA, better adjuvants.

• Commercial vaccines authorised in Europe

  License:Five types of ‘licences’ are known:

  1. Marketing Authorisation (MA)Issued by the European Medicines Agency (EMEA)Validity: Europe, unlimited time.
  2. European MA under exceptional circumstances:A MA under exceptional circumstances is a marketing authorisation with annual renewal, [Reg 2004/28 Art.39.7; Dir 2001/82 Art. 26.3.] Once commitments are fulfilled, a Full MA is granted.Validity: European Union
  3. National MA:A country may issue a national MA (for example Switzerland in 2008 for a vaccine against BTV8)Validity: national
  4. Provisional MA (PMA):If a product is not ready to obtain a full MA, some countries issue a PMA. In general, a PMA is granted for 1 to 2 years, renewable. Validity: national
  5. Temporary Authorisation of Use (TAU):A TAU is not a marketing authorisation but the authorisation to use a non-registered product.Validity: nationally; one year

  Several European manufacturers introduced other inactivated vaccines in Europe under the same regulatory framework: temporary authorizations of use, conditional licenses, or registrations under exceptional circumstances. These temporary authorizations were later converted into full marketing authorizationsBy 2018, no subunit vaccine is commercially available / licensed.It is not expected that subunit vaccines will be commercially available soon.Inactivated serotype specific vaccines are produced by a number of companies.Some bivalent inactivated vaccines are also available

  GAPS:

  • Registration process long, heavy and complex.
  • Clear guidelines/legislation by the European Commission are missing (No European wide standard): each country chooses the type of license it would like to issue even though preference should be given to a product holding a MA or MA under exceptional circumstances.
  • Legal background not ready for:o Multi-strain license*o Rapid licensing process for vaccines used in emergency**o Impact of vaccination on international trade…(EU adjusted legislation several times)*There is now the possibility to create a multi-strain dossier. If a company launches a multi strain dossier additional serotypes can be added.** No written standards/comparisons for Bluetongue so that the experience gained in bluetongue could shorten the process for other vector borne diseases. This gap analysis could used to identify parallels with other diseases
  • Vaccine market potential: Legislation for the use of vaccines varies by country; EU process: long and heavy. Funding for developmento No existing EC legislation/Guidelines how to proceed with a ‘new’ or exotic serotype (vaccinate or not)Currently separate authorisations are required for vaccines for each serotype. It may be possible to develop a generic authorisation that would apply to essentially similar vaccines but for different ‘types’ reducing the amount of data required prior to licensing.

• Marker vaccines available worldwide

  No.

  The “next generation” strategies, many with DIVA capability, did not have the chance to be launched on the market or tested on a large scale. They include non-replicating subunit vaccines and virus-like particles, heterologous microbial expression vectors (e.g. using poxviruses as expression vectors for immunogenic BTV proteins), and genetically engineered LAVs, including replicating but non-transmittable virus-based vaccines (DISC and DISA).

  GAPS:

  • Need for efficacious and safe DIVA vaccine
  • Further work is needed to commercialise experimental vaccine candidates and to develop cross reactive vaccine reagents/ strategies. These approaches would all be amenable to DIVA assay development.

• Marker vaccines authorised in Europe

  No.

• Effectiveness of vaccines / Main shortcomings of current vaccines

  Live attenuated are effective and provide long lasting immunity with a single dose but animals vaccinated by live vaccines cannot be differentiated from infected animals. There is also potential for some live vaccine strains to cause disease, particularly in naive animals/populations. Transmission and reassortment of some live vaccine strains can/has also occurred in the field.Inactivated or recombinant vaccines may need two injections to afford effective protection. The duration of immunity may also be shorter, requiring annual re-vaccination. No reliable DIVA assay is currently available for the inactivated BTV vaccines, and it may be difficult or even impossible to develop one.All of the current monovalent live or inactivated vaccines are type specific. Cross-protection can be generated by serial vaccination with multiple serotype vaccines.

  GAPS:

  • Ideally: one shot application for inactivated vaccine that gives a long-lasting protection from vireamia and clinical signs.
  • Work is needed to explore the nature of cross-serotype specific protection and epitopes, and cross topotype protection and the viral antigens involved. This may lead to development of truly cross-reactive vaccines, offering protection against multiple serotypes. These would be particularly welcome in areas where multiple types are circulating and causing disease. They could also be used in a wider eradication campaign.

• Commercial potential for vaccines in Europe

  Even if many countries decided to move from compulsory, government driven vaccination policy to voluntary, and private driven vaccination schedules, the commercial potential for future vaccines against BTV is still appealing. Although not reaching the same peak of 2008 and 2009, sales of inactivated vaccines against BTV will have a stable trend. Incursions of new virulent strains continue to take place and the vaccine is the only tool that protects from clinical disease and helps to avoid losses due to trade barriers.However, the future BTV vaccine market is a highly unpredictable and thus poses a great risk for vaccine producers. Consequently, vaccine producer are reluctant to take risks to develop new vaccines for a ‘non-existing market’.If there is a move towards disease control or even eradication inEndemic areas (such as India) there may be greater demand for truly effective and cross-reactive vaccines. Any future incursions into northern Europe could also increase the demand for relevant vaccines.

  Inactivated vaccines offer significant advantages over attenuated vaccines because absence of replicating virus eliminates concerns about viraemia, vector transmission and reversion to virulence.Recent reverse genetics techniques have provided novel approaches to developing safe vaccines. This technology offers substantial advantages both in terms of safety and the potential of developing a marker vaccine. The latter could be used as a prophylaxis in areas at risk, without endangering the “free” status of the region. An accompanying serological test would allow the distinction between vaccinated and infected animals. A platform of DISA/DIVA vaccine for BTV has been developed.Subunit vaccines are under development but not commercially available at the present time. With the highly unpredictable market and high development costs it is doubtful whether subunit vaccines against BTV will reach the market soon.

  Inactivated vaccines:

  Market (risks):'Endemic’ BTV serotypes in Europe: absence of BTV vaccine market vaccine producer may stop producing antigens and

  GAPS:

  Inactivated vaccines & subunit vaccines:

  • European BTV strain bank where vaccine producer have quick access to strains with full description of all details (eg origin, organ, location) is needed
  • Antigen bank containing a minimum stock of antigens and serotypes for the future
  • Vaccination ring around Europe: Protect Europe by vaccinating North Africa, Israel and Greece => European Commission supports financially or by providing high quality BTV vaccine the vaccination programs in those countries, which pose a risk to Europe (known as entry doors
  • It may be difficult for companies to provide vaccines to cope with a fluctuating market and provide appropriate vaccine for arrival of new serotypes.
  • If vaccines with longer shelf life can be developed a central vaccine bank might be feasible.
  • Diva assays needed for next-generation sub-unit vaccines.

• Regulatory and/or policy challenges to approval

  To allow authorisation of vaccines in a more streamlined way provided the seed vaccine meets the regulatory requirements. This is particularly important for the current type specific vaccines, as a new set of reagents/approval is needed for each type. A truly cross-reactive vaccine might avoid such problems.

  If a generic set of procedures and materials for vaccine production could be approved, then a change of only the seed virus/antigen might allow / lead to a more rapid approval.

  The development of more appropriate/effective vaccine adjuvants/ carriers. The use of toll-like receptors to direct the immune response may lead to more effective immune responses to single-shot / non-replicating or virus-vectored vaccines.

  GAP:

  Studies of the target host immune system in order to engineer more effective vaccine delivery mechanisms (cross-serotype / single shot).

• Commercial feasibility (e.g manufacturing)

  High.

  GAPS:

  • Commercial feasibility is in general high which does not necessarily mean that vaccine is available in a short time if there is no stock available (endemic and exotic strains), see also antigen/vaccine bank and market risks.
  • New BTV vaccines can however be developed relatively rapidly where close links between laboratories, international organizations, governments and industry exist. One clear area for improvement may also be in government action as placing pre-orders for vaccines is likely to result in more rapid delivery and earlier vaccine deployment.
  • Meanwhile, problems with vaccine uptake can be reduced by improving the provision of information to livestock keepers about the effectiveness and safety of a given vaccine and the economic risk of not vaccinating.

• Opportunity for barrier protection

  Low.

• Opportunity for new developments

  Multivalent vaccines with longer shelf life

  Multivalent or cross-reactive vaccines with longer shelf life and associated DIVA assay. Development of single shot inactivated or sub-unit vaccines. Non-replicating vaccines that generate a longer lived immune response.

  GAP:

  Registration process and acceptance by registration bodies (EMEA) of multi-valent vaccines.There is now the possibility to create a multi-strain dossier. If a company launches a multi strain dossier additional serotypes can be added.

• Pharmaceutical availability

• Current therapy (curative and preventive)

  Symptomatic treatment and care includes Non Steroidal Anti-inflammatory drugs (NSAID) to reduce pain, fluid therapy to treat or prevent dehydration, provision of shade against the sun, protection against extremes of temperature, use of mild disinfectants to irrigate affected areas, general nursing care, the provision of a trough of cool water in which affected animals can cool their muzzle, provision of soft green foods during the period when the mouth lesions are painful, and long-acting antibiotics against secondary bacterial diseases.Livestock can be treated directly with synthetic pyrethroids, generally in the form of pour-ons. Insecticides may reduce biting rates and therefore vectorial capacity of midges. Even the most effective repellents are likely to reduce biting rates for only a few hours. Most of these products also have a withdrawal period which makes difficult their use when moving animals for slaughtering.

  GAP:

  Possibility of developing therapies targeting specific viral proteins/ functions, e.g. RNA silencing or specific inhibition of specific viral enzymes.

• Future therapy

  There is a need of repellent products with long acting efficacy and shorter withdrawal period.

• Commercial potential for pharmaceuticals in Europe

  There is a need of repellent products with long acting efficacy and shorter withdrawal period.

• Regulatory and/or policy challenges to approval

  None needed.

• Commercial feasibility (e.g manufacturing)

  Not applicable at present.

• Opportunities for new developments

  A range of possibilites including husbandry modification, vector control, habitat alteration, adult vector insecticides, use of larvicides.

• New developments for diagnostic tests

• Requirements for diagnostics development

  Kinetics of antibody response.Kinetics of virus replication at the genome and protein level.BTV genome database.

  GAP:

  Better knowledge of antigenicity / cross reactivity of viral proteins between different strains / topotypes.

• Time to develop new or improved diagnostics

  If required can be done fast, depends on priorities. The step that takes the time is a full evaluation/validation of the assay with a wide range of strains.

• Cost of developing new or improved diagnostics and their validation

  If a company has to start from scratch in the development and validation of new or improved diagnostics it will take long time and the cost will be significant. Cooperation between diagnostic/research labs and commercial companies is absolutely necessary in order to pool resources, take advantage of relevant expertise, and reduce cost.

• Research requirements for new or improved diagnostics

  Develop diagnostic reagents based on viral genes and proteins to specifically distinguish viral infection from vaccination.Cooperation and serious involvement of public and private labs with a combination of research and diagnostics is necessary.Set up processing protocol and data analysis workflow for the diagnostic use of NGS platforms of second (e.g. Illumina) and third generation (Oxford nanopore technology).

  GAP:

  • International sequence database.
  • Harmonization of NGS protocols.

• Technology to determine virus freedom in animals

  C-ELISA and PCR-diagnostics.Serotype-specific ELISAs and serotype-specific PCRs for all serotypes are useful to determine freedom for a respective serotype.

• New developments for vaccines

• Requirements for vaccines development / main characteristics for improved vaccines

  Safe, lifelong and broad protection.

  GAPS:

  • Better knowledge on the virus diversity and variability in different animal species and vectors.
  • Better knowledge on the function of viral genes and proteins.
  • Better knowledge on the virus biology and its capacity to persist in susceptible animals, or susceptible insect populations.
  • Better knowledge on the immune mechanisms (cellular and humoral) involved in protection.
  • Better knowledge on immune mechanisms in the insect vectors.

• Time to develop new or improved vaccines

  5 years.

• Cost of developing new or improved vaccines and their validation

  Expensive.

• Research requirements for new or improved vaccines

  Further research is required in the development of vaccines that are safe and lifelong protective for livestock against all BTV serotypes.Thanks to advanced molecular virology and the availability of genomic information on BTV, there are new opportunities to create novel concept vaccines. Focus should be on new vaccine development and improvement of existing vaccines for a broader and longer term protection. Improved diagnostic and vaccination strategies are certainly crucial to tackle the growing threat of BTV infection in order to effectively prevent and control the disease in the future.

  GAP:

  Explore plants as possible vaccine production platforms.

• New developments for pharmaceuticals

• Requirements for pharmaceuticals development

  None anticipated at present.

• Time to develop new or improved pharmaceuticals

  Not applicable.

• Cost of developing new or improved pharmaceuticals and their validation

  Not applicable.

• Research requirements for new or improved pharmaceuticals

  None at present apart from new methods of vector control both in terms of killing vectors but also preventing vectors form attacking hosts.

Disease details

• Description and characteristics

• Pathogen

  Bluetongue virus (BTV) is the type species of the genus Orbivirus, family Reovirdae. The bluetongue virus particle is approximately 80 nm in diameter, is non-enveloped and is composed of three concentric protein layers, which enclose one copy each of the ten linear segments of double-stranded RNA that form the virus genome. The viral genome segments code for 7 structural proteins (VP1-VP7) and 5 non structural proteins (NS1-NS5). All bluetongue isolates share common antigenic determinants, the most immunodominant of which is the core surface protein VP7. Detection of antibodies to VP7 can be used for identification of BTV infected animals. The outer-most capsid protein, VP2, is variable and determines the identity of the specific serotype of the virus. Including the recent identification of two novel BTV types, a total of 27 serotypes have been officially identified worldwide and more have been described. The highly conserved atomic structure of the BTV core has been resolved by x-ray crystallography. Although the structure of outer capsid proteins of a few serotypes has been explored by cryo-microscopy, no atomic structures are as yet available. Although the core proteins of BTV are highly conserved, they show sequence variations that reflect the geographic origins of the virus isolate, identifying at least two major ‘topotypes’ (east and west) along with a number of sub-groups.

  GAPS:

  • Identify and understand the mechanisms involved in cell binding and initiation of infection, genome replication, assembly and packaging, control of differential protein expression levels in mammalian and insect cells, release and transmission of BTV at the molecular, cellular and whole organism level.
  • Determine the atomic structure, / function relationships and molecular organisation of the outer capsid layer and proteins of selected viruses from different BTV serotypes, and different topotypes, using x-ray crystallography, cryo-electron microscopy, reverse genetics, biochemical and immunological / immuno-staining studies. These analyses would provide a better understanding of the structural and epitope basis for variability and cross-reactivity between BTV serotypes, potentially providing a basis for the design of cross-reactive vaccines.
  • The recent identification of several new BTV serotypes using the latest generation of diagnostic assays and sequencing technologies/databases, suggests that there are still other serotypes / viruses yet to be discovered. The biological/disease significance and transmission features of these new strains have not yet been fully explored. Furthermore, the remarkable genetic heterogeneity amongst field strains of BTV remains to be fully characterized, which is important as this genetic variation in turn translates into virus strains with different biological properties such as virulence and, perhaps, vector tropism.
  • Better understand the virus-related genetic control of horizontal BTV transmission in the ruminant host associated to atypical and typical BTV serotypes.
  • Investigate whether atypical and typical BTV strains could reassort in case of co-infection.

• Variability of the disease

  BTV has a wide pathogenic variability, which is partially virus strain dependent and partially host dependent. The neutralising antibody response generated during infection of the mammalian host is protective but it is BTV serotype specific. However an immune response is also generated against the more conserved (cross-reactive) core and non-structural proteins, which also appears to be at least partially protective against the clinical disease. This is likely to involve both antibody and cell-mediated responses. Consequently although susceptible hosts can be sequentially infected by multiple different BTV serotypes, they become progressively protected against the more severe aspects of the disease. Cross-reactions (usually low level or one way) may also occur with other orbiviruses especially Epizootic Haemorrhagic Disease virus (EHDV).Owing to the insect transmission, in temperate climates, the infection and disease are seasonal occurring in the late summer and autumn. Not all serotypes of BTV are present in each of the areas where it exists around the world.Different vector species also exist in different regions (episystems) around the world and variations may exist in the viruses present in a region that reflect adaptation to the vectors that are present.Several ‘western’ BTV strains (BTV-1, 4, 6, 8, 11 and 14) have been detected in northern-central Europe, suggesting that they can be transmitted effectively by Culicoides species present in the region. However, several additional BTV strains (eastern BTV-1, 4, 9, 16 and western BTV-2 and some strains of western BTV-4) have persisted in Southern Europe without (so far) spreading to the north. This has provided a unique opportunity for these viruses to exchange genome segments, generating novel virus strains, some of which appear to have distinct biological characteristics. In addition, BTV-26 and BTV-27 have been shown to be transmitted by direct contact rather than vectors and also they do not replicate in Culicoides-derived KC cells. By reverse genetics, the genome segments of BTV-26 that restrict replication in KC cells have been identified.Reassortment is a frequent process that plays an important and on-going role in evolution of BTV. Reassortment is evident in all ten segments without a significant bias towards any particular segment.

  GAPS:

  • BTV genome sequence analyses to define the existence and global distribution of different virus serotypes / lineages / topotypes; determine significance of strain variations and genome-segment-reassortment, in infection of the mammalian host, infection of different insect vectors, and transmission between them.
  • Investigate why viruses emerge and persist and what determines their transmissibility, changes in their distribution, and evolution. Investigate the viral genomic constraints involved in the geographical clustering of BTV strains by reverse genetics and vector competence studies.
  • The identity of neutralising epitopes on the outer capsid proteins (VP2 and VP5) of BTV has not been fully resolved for the different serotypes. There is evidence for at least some cross-reactive sites between types, identification of which could help to support the design and development of cross-reactive vaccines.
  • The nature of the mammalian cell-mediated immune response against BTV and its role in protection against homologous and heterologous strains / topotypes has not been fully explored.
  • Use of new reverse genetics technologies to explore the genetic basis for biological characteristics (e.g. virulence or transmissibility, serotype, temperature dependence, vector competence, etc) of different virus strains.
  • The immune response of adult Culicoides to BTV infection has not been explored, but may have an important role in persistence and transmission of different viruses by different insect species / populations.
  • Continue to provide sequence data for multiple well documented isolates of BTV, to provide information concerning their molecular epidemiology and the processes of virus evolution. These studies will provide ‘reference’ sequences for each of the ten genome segments from specific BTV lineages, topotypes and strains around the world, to identify genome segment reassortment events and strain movements in the field.

• Stability of the agent/pathogen in the environment

  The bluetongue virus (BTV) is very stable. It survives essentially forever when frozen, although the process of freezing and thawing will itself reduce the titre of virus by approximately 1 log10. It can survive for years in whole blood kept in a refrigerator. The virus survives as long as 60 days in the circulation after infection of a ruminant, and infection persists life-long in vector insects. (BTV-25 infection of goats clearly is an exception to this general property because viraemia in BTV-25 infected goats can persist for several years and potentially is lifelong). The virus apparently survives freezing winters. However, the mechanism behind this survival or ‘over-wintering’ remains unknown although vertical transmission in the mammalian host has been demonstrated and may contribute although this is disputed. It has been proposed that BTV “overwinters” in temperate areas through low level circulation of the virus in animals and vectors, including infected adult insects that survive for relatively long periods even in winter. There is also some evidence for detection of BTV RNA in Culicoides larvae collected in the field from outbreak sites in North America. This suggests that in some cases the virus may be transmitted vertically in the insect vector, However, attempts to recover infectious virus were unsuccessful and the epidemiological significance of these observations is uncertain.

  GAP:

  Understanding the over wintering mechanism(s) in the host, vector and/or environment including potential differences between species and distinguished ecological zones.

• Species involved

• Animal infected/carrier/disease

  Bluetongue virus infects many domesticated, zoo and wild ruminants including sheep, goats, cattle, South American camelids, buffalo, bison, deer, antelope, bighorn sheep and North American elk. Clinical disease is most often seen in sheep, occasionally in goats, but rarely in cattle. However, with BTV-8 in the EU, clinical disease in cattle was reported from several countries. Severe disease can also occur in some wild ruminants including white-tailed deer (Odocoileus virginianus), pronghorn (Antilocapra americana) and desert bighorn sheep (Ovis canadensis). Disease also has been described in an extensive variety of non-African ungulates in zoos in Europe, and in South American camelids present in the UK, France, Germany and USA. In Africa, some large carnivores have antibodies to bluetongue and fatal disease has been described in Eurasian lynx. In North America a contaminated vaccine resulted in abortions and deaths in pregnant dogs.

  GAPS:

  • The role of wildlife species in the persistence of BTV in the environment.
  • The role of carnivores including domestic dogs in the transmission of BTV.
  • Understanding the over-wintering mechanisms including more investigation on the potential role of reservoir and vertebrate host in carrying the virus.
  • The potential role of the vertebrate host as a carrier for BTV – overwintering mechanism.

• Human infected/disease

  No.

• Vector cyclical/non-cyclical

  Biological vectors are Culicoides species.There have also been some reports of BTV infection/recovery from tick species, although the epidemiological significance of these observations is uncertain.

  GAPS:

  • Identification and impact of the different Culicoides species involved in the transmission of BTV in the different ecological zones.
  • Understanding of BTV transmission by other routes (e.g. via other biological vectors, by mechanical transmission involving other biting insect species, via an oral route such as ingestion of infected meat/placenta).

• Reservoir (animal, environment)

  Cattle are the main reservoir and amplification hosts. Sheep and possibly other ruminants are also a potential source of virus for transmission.In the last few years next to the classical 24 serotypes which are transmitted by vectors, new serotypes (BTV-25, BTV-26, BTV-27, BTVXITL2015) (Hofmann et al. 2008; Maan et al. 2011; Zientara et al., 2014; Savini et al. 2017) have been described. They are characterized by causing asymptomatic infections, being directly transmitted, at least BTV-26 (Batten et al.)2014), having long viraemia (BTV-25, Vogtlin et al., 2013) and using goats and sheep as reservoir hosts.

  GAP:

  Understanding the over-wintering mechanisms including more investigation on the potential role of reservoir and vertebrate host in carrying the virus.

• Description of infection & disease in natural hosts

• Transmissibility

  Insects: Transmission primarily by biting midges (adult females of certain Culicoides species). Non contagious disease, however, there is also evidence for oral infection / transmission, although this is uncommon and its epidemiological significance uncertain. Trans-placental transmission occurs with lab-adapted viruses (mostly live attenuated vaccine strains) and also some virulent field strains such as the current European BTV-8 strain. Minimally passaged (one passage on KC cells and/or followed by a single passage on mammalian cells) BTV-2 field strain has been shown to pass the placental barrier of experimentally infected ewes and infect offspring. Also BTV-1 passaged in vitro caused abortion in experimentally infected pregnant ewes. BTV-26 and BTV-27 have been shown to be transmitted by direct contact rather than vectors and that BTV-26 does not replicate also in Culicoides-derived KC cells. By reverse genetics, the genome segments of BTV-26 that restrict replication in KC cells have been identified.The proteases present in saliva from adult Culicoides can modify the outer capsid proteins of the virus, enhancing its infectivity for the vector insect and removing its hemagglutination activity. These changes and enzymes may therefore play a significant role in the infection processes in both the insect and mammalian host.

  GAPS:

  • Understand the processes and mechanisms which underlie transmission of BTV by arthropod vectors, including factors that promote, or act as barriers to transmission / vector competence with specific virus strains
  • Full genome analyses and comparison of genetic variations between vector and non-vector populations / species of Culicoides, to determine the mechanisms involved in barriers to infection / transmission.
  • Culicoides saliva contains a large number (undetermined) of individual proteins, including both enzymes (e.g. proteases) and inhibitors. The role of these proteins needs further investigation.
  • Define the thermal & vector requirements of BTV in both the field and laboratory, to predict incursion risk into the EU.
  • Investigate the effect of temperature on the extrinsic incubation period (EIP) in different species of Culicoides midges.
  • Mechanism of transplacental transmission:a) Determine the viral genetic factors involved in transplacental transmission. b) Determine why transplacental transmission occurs with some field strains (for example the northern European strain of BTV-8) but not others. c) Determine why transplacental transmission occurs in animals infected with laboratory adapted BTV strains far more readily than with field strains. d) Determine the molecular mechanisms controlling transplacental transmission. e) Determine whether viraemic newborns play a role in BTV spread/overwintering, and the importance of colostral antibodies in suppressing infection.
  • The genetic control of vector competence and horizontal transmission
  • Investigate further a possible role for oral transmission, through infected colostrum or placenta, in the spread and over-wintering of BTV.
  • Determine the significance of short distance (local) spread as compared to long distance spread?
  • During outbreaks investigate the proportion of spread by wind in relation to transmission by animal movement taking into account different regions of Europe and the world.
  • Investigate the potential role of ticks and other arthropod vectors in the transmission of BTV.

• Pathogenic life cycle stages

  During its life cycle the virus infects arthropod and vertebrate hosts. The virus has an enzootic cycle and is transmitted from arthropod vector to competent reservoir host.For the new serotypes (small ruminant adapted serotypes) or at least for BTV-26 and BTV-27, a direct life cycle (host-host transmission) has been proposed.

  Domain: Viral hosts belong to the Domain Eucarya.

  GAPS:

  • Vector competence and ability of different species of Culicoides vectors to transmit different serotypes / strains / topotypes of BTV.
  • Investigate the infection dynamic of atypical BTV serotypes: incubation period, infection route, viraemia, immune response, virus excretion etc
  • Investigate the occurrence of riassortment phenomena in Culicoides spp.

• Signs/Morbidity

  The vast majority of infections with bluetongue are clinically inapparent. In a percentage of infected sheep and occasionally other ruminants, more severe disease can occur. The severity of clinical signs depends on virulence of the BTV strain, breed and immune status of the host, and is greater in naive animals / populations. Acute form (sheep and some species of deer)Transient fever (up to 42°C), depressionInflammation, haemorrhages (particularly in the skin), ulceration, erosion and necrosis of the oral mucosa, swollen and sometimes cyanotic tongueSevere pulmonary oedema, serous effusions and subcutaneous and intermuscular oedemaLameness due to coronitis or pododermatitisTorticollisConjunctivitisComplications of pneumoniaEmaciationEither death within 8-10 days or long recovery with alopecia, sterility and growth delayAbortion of severely affected animals (often without virus-infection of the foetus)Can be teratogenic in cattle and sheep (depending on strain), and can lead to dummy calf syndrome. Early embryonic loss and decreased reproductive efficiency is a more frequently seen manifestation of the disease in cattle and can be devastating to their calf/milk production. Clinical signs in cattle also include hyperaemia and necrosis of the muzzle (“burnt muzzle”) and patchy dermatitis.

  GAPS:

  • Why are some strains of BTV more pathogenic than others?
  • What is the basis for the reduced severity of clinical signs in those animals that have previously been infected with heterologous serotypes. Is a cell-mediated or cross-reactive antibody response involved?

• Incubation period

  In sheep, the incubation period is usually around 5 to 10 days. Cattle can become viraemic starting at 3-4 days post-infection, but rarely develop clinical symptoms. Infected animals are usually infectious to the insect vector for several weeks.

  GAP:

  What is the maximum length of time that animals remain infectious to midges compared to persistence of viral RNA and virus (isolated in the lab)

• Mortality

  In sheep, the severity of disease varies with the breed of sheep, immune status, virus strain and environmental stresses. The morbidity rate can be as high as 75% in this species. The mortality rate is usually 0-20%, but can be up to 70% in highly susceptible sheep. Similar morbidity and mortality rates are seen occasionally in certain other species, including South American camelids, zoo and free ranging non-African ungulates, with a morbidity rate as high as 100% and a mortality rate of 80-90%.Most (but not all) infections in cattle, goats and North American elk are asymptomatic. In cattle, up to 5% of the animals infected with certain BTV serotypes (serotype 8 in particular) may become ill, but deaths are rare. In some animals, lameness and poor condition can persist for some time. The morbidity of the atypical BTV serotypes is much higher (over 70%). Infections with these serotypes in goats and sheep are usually asymptomatic.European BTV-8 strain isolated at the beginning of the bluetongue outbreak in 2006 was more virulent than a strain isolated toward the end of the outbreak and when the same strain re-appeared in France in 2015. There is a link between the variability of the BTV population as a whole and virulence, and Culicoides cells might function as an "incubator" of viral variants. BTV virulence seems to be determined by different viral genomic segments. Some BTV genome segments have been demonstrated to be important virulence factors.

  GAPS:

  • Expand the knowledge gained so far upon host and virulence factors determining the clinical outcome of infection.
  • Why are some strains of BTV more virulent than other in cattle and sheep?
  • Why are certain breeds of sheep resistant to BTV infection whereas other breeds are extremely susceptible?
  • Natural resistance - why are African ruminants in general more resistant than non-African ruminants.
  • It has been suggested that the introduction of novel exotic strains to endemic areas can result in more severe disease in native breeds (as observed in India). Is this due to differences in the virus topotype, resulting in a reduction in cross-reactive, non-neutralising antibody based or cell mediated cross-protection?

• Shedding kinetic patterns

  Although virus may be detected in the blood of cattle and sheep by real time RT-PCR for several months, infected animals usually only transmit virus to a competent biting vector for a number of weeks (at most) after infection. The OIE states that the infectious period is up to 60 days post infection although most animals are infectious to vectors for a shorter period. It has demonstrated that, at least for BTV-8, the infectious dose does not influence neither the length nor the magnitude of BTV-8 viraemia. The epidemiology of BTV-25, BTV-26 and BTV-27 infection of goats appears to be different than that of the other serotypes (BTV 1 – 24) and may not involve Culicoides midges. Recent studies also suggest direct contact transmission of BTV-26 and BTV-27, likely by aerosol, between livestock. In one report, serotype 25 viral RNA was found for at least 2 years in individual animals, and the blood of some goats was infectious at 12-19 months.

  GAPS:

  • Identification of the infectious period and period of detection by molecular methods under natural environmental conditions.
  • How long do sheep and cattle remain infectious and is the length of the infectious period dependent on the infectious dose for the remaining BTV serotypes?
  • Are there host genetic factors involved in controlling the length of the infectious period?
  • Are there genetic factors of BTV involved in controlling the infectious period?
  • Are there genetic factors of the vector involved in controlling the infectious period of the host (does the species of infecting Culicoides influence the length of the infectious period).
  • The role of Culicoides saliva and the significance of the local hypersensitivity reaction, in the mechanism and kinetics of infection and dissemination in the mammalian host.

• Mechanism of pathogenicity

  After infection via the saliva of a biting midge, BTV can infect and is transported by different cell types, including cellular components of the host’s immune system. Replication of BTV in target cells, notably mononuclear phagocytic cells (dendritic cells and macrophages) and endothelium, leads to the generation of the innate and adaptive immune responses that mediate both initial virus clearance and subsequent resistance to infection with the homologous virus serotype.The virus spreads to and multiplies in the regional lymph nodes and then disseminates to secondary organs of replication, followed by dissemination in the blood. This systemic multiplication and spread allows ample opportunity for humoral and cell-mediated immune responses to develop. As BTV is associated with the cellular fraction of the blood where it is protected from the effects of humoral antibodies, extended viraemia may occur and virus and antibody may circulate in the system at the same time.

  GAPS:

  • What are the molecular mechanisms related to pathogenicity?
  • The role of innate and acquired immune responses in the pathogenesis of disease.
  • The role of Culicoides saliva in the pathogenesis of disease.
  • Sites of transient persistence of BTV in the vertebrate host

• Zoonotic potential

• Reported incidence in humans

  Bluetongue is not a significant threat to human health. Only one suspect human case has ever been reported, but reasonable precautions should be taken while working with this virus.

• Risk of occurence in humans, populations at risk, specific risk factors

  None - Bluetongue does not affect humans, nor is there any risk of the disease being contracted or spread through meat or milk. (EFSA).

• Symptoms described in humans

  None.

• Estimated level of under-reporting in humans

  None.

• Likelihood of spread in humans

  None.

• Impact on animal welfare and biodiversity

• Both disease and prevention/control measures related

  Variable impact depending on species, immune status, the virulence, and genotype/topotype of the of the infecting virus strain.

• Endangered wild species affected or not (estimation for Europe / worldwide)

  Possibly if susceptible.There is evidence for BTV specific antibodies in large carnivores in Africa. Eurasian Lynx fed with infected meat also became fatally infected with BTV-8. It is possible that the incidence of BTV could represent a significant threat of infection and even the survival of endangered predator species.

  Certain deer species (e.g. white tailed deer) can also be very susceptible to BTV infection showing very severe clinical signs, including significant fatalities.

  GAPS:

  • Investigate the pathogenesis of BTV infection in different wildlife species to assess relative susceptibilities.
  • Understand the potential impact on endangered species of the introduction of exotic strains / topotypes in endemic areas.
  • The efficacy of vaccination against BTV, has not been assessed neither in wild life nor in predator species, (e,g, in dogs).

• Slaughter necessity according to EU rules or other regions

  Only for animals welfare reasons. BTV-8 caused severe clinical signs in sheep necessitating, for animal welfare reasons, slaughter.

• Geographical distribution and spread

• Current occurence/distribution

  The virus is present in a broad band of countries extending approximately between 40°N and 35°S although in some regions (e.g. China, North America, and more recently Europe) it may extend to 55^oN. With the notable exception of the recently identified, horizontally transmitted small ruminant adapted BTVs, classical BTV has been shown by serology and by virus isolation to be limited to regions where vector species of Culicoides are present and within these regions vector transmission is limited to those periods of the year when adult Culicoides are active (e.g. Africa, the Americas, Australia, the Middle East and some countries of southern Asia and Oceania).The virus is present in some regions with little associated clinical disease in the native ruminant populations. However introduction of exotic strains (topotypes) may lead to more severe disease in these native breeds. The introduction of exotic breeds may also lead to more severe disease caused by local strains of virus. In the United States, the distribution of vector usually limits infections to the southern and western states. In other countries the distribution of the vectors similarly limits the distribution of the virus.Until recently bluetongue had only been recorded in southern regions of the EU including parts of Italy, Spain, France, Greece and Portugal, matching the distribution of the known vector species Culicoides imicola . However, since 1998 the bluetongue situation in the EU has changed considerably, with incursions of six new serotypes into southern Europe and serotypes 1, 6, 8 and 11 into northern Europe. In late summer 2006, several northern European countries reported the first ever outbreaks of bluetongue in the region, caused by BTV serotype 8 (BTV-8), including Holland, Belgium, Germany, Luxembourg and France. Further outbreaks were reported in 2007 and 2008. In 2007, northern Europe experienced a dramatic increase of new cases in the existing infected areas, and cases numbered into the many tens of thousands, as the disease spread steadily across most of Europe, including an incursion in the UK.Affected countries began vaccination programmes in 2008 when, as expected, disease re-emerged in many of them. Vaccination with inactivated vaccines has proved highly successful, dramatically reducing the number of BTV cases in the majority of northern and western European countries in 2008. The exception was France which reported over 23,000 cases of BTV-8 and over 3,500 cases of BTV1 in 2008. However, the vaccination campaign had a dramatic effect in France during 2009, with a total of only three successful virus isolations. Compulsory and voluntary vaccination programmes have continued against BTV-1 and BTV-8 throughout 2009 and into 2010. These campaigns have been extremely successful with no cases of BTV-8 being reported throughout 2010 in Northern and Western Europe. In 2015, BTV-8 re-emerge in France and between 2018 and 2019 it spread in Germany, Switzerland and Belgium. Western BTV-1incursion was recorded in Italy in 2012-2014. In 2012, a reassortant BTV-4 strain was also shown to circulate in Sardinia whereas a multiple reassortant BTV-4 strain with gene segments originating from BTV-1, BTV-2, BTV-4, BTV-16 and BTV-24 parental strains emerged in the Balkans in 2014. The same BTV-4 strain was detected in Central Europe and Italy between 2014 and 2018. A multiple reassortant BTV-4 was also detected in Spain in 2014. Novel BTV serotypes have been discovered in healthy goats in Corsica and Italy, appointed as BTV-27 and BTV-X ITA2015, respectively. In 2017, BTV-3 was detected in sheep in Sicily. It was the first time that this serotype was detected in Italy and Europe. In 2018, the same strain was detected in Sardinia. More potential novel serotypes infecting sheep and goats have been reported in Europe in recent years.

  GAPS:

  • To understand and model the distribution, spread, persistence and risks represented by BTV including viral, vector and host dependent factors, and the importance of environmental factors, such as temperature (day and night), humidity, geographic conditions, etc.
  • Identification of apparent associations between some genotypes of virus and some vector species has resulted in a concept of viral-vector ’episystems’. A more complete understanding of these epidemiological aspects may further facilitate international trade in ruminants.
  • . Information is not always up to date. Several countries do not report test results or no testing is performed, so the disease status or changes in status therefore remain unknown.

• Epizootic/endemic- if epidemic frequency of outbreaks

  Regarding the epidemiological status in relation to BT, it has to be said that the status could be related to the occurrence of a single or multiple serotypes. According to the recent history of BT in Europe, it is reasonable to say that BT is endemic in the countries of Southern and Eastern Europe. BTV-8 is likely endemic in France. In the other countries of Central and Northern Europe BT is epidemic whereas sporadic in the countries of the Scandinavian peninsula. Transmission is seasonal and usually depends on the infection of the Culicoides vectors from the reservoir hosts such as cattle early in the season. When present, atypical BTVs are usually endemic.

  GAPS:

  • Continuous monitoring and overview of global BT situation.
  • Investigate the competence of the northern European species of Culicoides midges.
  • The nature of future climate change in Europe is uncertain. Hotter summers are considered likely to lead to an increased likelihood of further BTV incursions in Europe, and perhaps persistence of the virus.

• Seasonality

  Seasonal cycle related to the movement life cycle and seasonal abundance of the adults of Culicoides vectors. Culicoides peaks depend on season, local meteorological conditions and species involved. Culicoides abundance is also linked to livestock density and land use. Atypical BTV (at least BTV-26 and BTV-27) occurrence is not related to seasons

  GAP:

  Factors involved in vector survival throughout the winter period, and their re-emergence during the vector season (usually summer).

• Speed of spatial spread during an outbreak

  The rate of the BTV spread after introduction into a naïve population depends on many factors: infection rate in the Culicoides vectors, animal density, the time of the year, wind. The wind borne spread of infected Culicoides can be rapid, and can lead to dispersal of the virus over considerable distances across water (e.g. from Morocco to Iberia, across the English Channel, from Northern Africa to Sicily, Sardinia, Baleari Islands) Initial spread may involve small numbers of infected animals over large distances caused by movements of infected animals or vector insects (as seen in Europe during 2006), but spread in subsequent seasons (2007) involved very large numbers of animals across massive areas. BTV spread has been estimated from 6 to 30 km/week depending on outbreaks, country, methods of calculation etc.

  GAPS :

  • Understand in vivo replication and transmission routes, and the significance of climate, providing input into predicting and modelling viral dissemination / risks.
  • Identify risks associated with BT and develop epidemiological models for its spread & control.
  • Investigate the speed of spatial spread of BT depending on the different transmission modes under varying meteorological and environmental conditions.
  • Rates of spread through different Culicoides species taking into consideration host preference, efficiency of transmission etc.

• Transboundary potential of the disease

  Bluetongue virus has recently expanded its geographic range and is able to cross borders due to the wide distributions of vector species of Culicoides, their high mobility and their adaptability to various climatic conditions.

  Significantly, the route of introduction of BTV-6, 8 and 11 in northern Europe are all unknown. Although such events are rare, they are clearly of major epidemiological significance.Apart from possible spread of BTV by infected midges through the wind, one of the most important way of BTV spread is through the movement of viraemic animals.

  GAPS:

  • Impact of global trade, animal movement, cultural habits and travel on disease spread in relation to long distance dispersal mechanism like wind spread.
  • Understanding of gradual and apparently limited expansion versus “long distance incursions”. Both are not always related to the involved Culicoides species. Why?

• Route of Transmission

• Usual mode of transmission (introduction, means of spread)

  The usual route of BTV transmission to its animal (ruminant) hosts is via the bites of virus-infected haematophagous Culicoides midges that act as biological vectors of the virus. Vector midges play a crucial role to the natural epidemiology and spread of BTV. Midges of certain species in the genus Culicoides transmit BTV between susceptible ruminants, having become infected by feeding on viraemic animals (the vertebrate host). After a replication period of 6–8 days in the insect’s salivary glands (development time being dependent upon temperature) the virus can be transmitted to a new vertebrate host during feeding. Infected midges remain infective for life. Infection of the midge is a relatively inefficient process with less than ~10% of insects that ingest a viraemic blood meal becoming infected. An even lower percentage may become fully infected and capable of transmitting the virus, depending on the insect vector species. However, transmission of virus from a fully infected insect to mammalian host is an efficient process (possibly up to 100% efficient). This may help to explain the transmission of virus by wind borne insects over large distances. Vector midges can fly short distances of 1- 2 km, but they can be blown much farther by wind. Long distance spread of BTV from endemic regions to adjacent uninfected areas can occur via the wind-borne dissemination of virus-infected midges, especially over water.The movement of BTV-infected animals can be responsible for translocation of BTV, however, such occurrences are only important if the local vector population within the receiving region is able to efficiently acquire and transmit the introduced virus.

  GAPS:

  • Determination of likelihood of introduction of BT into new regions depending on the mode of transmission.
  • The Culicoides saliva contains multiple proteins, including proteases that can modify the virus capsid, influencing its cell binding properties and infectivity. Further information is required concerning the significance of these proteins/ modification during transmission from insect to host and from host to insect.

• Occasional mode of transmission

  Vector-independent transmission of BTV clearly can occur, although its significance is largely unknown:

  • direct contact transmission of BTV-26, likely by aerosol, between livestock
  • transplacental transmission by the northern European strain of BTV-8 and lab adapted vaccine strains (live attenuated vaccine strains).
  • the occasional occurrence of venereal transmission from infected male ruminants to females and their offspring.
  • oral transmission to ruminants (eating infected placenta, ingestion of virus-containing colostrum) and carnivores (eating infected meat)
  • contamination of biological products, notably those that utilize foetal bovine serum
  • direct human role in mediating the introduction of these viruses

  GAPS:

  • Further work is required in order to study oral / mechanical transmission of BTV and the significance of these transmission mechanisms in the field.
  • Investigate further a possible role for oral transmission, through infected colostrum or placenta, in the spread and over-wintering of BTV.
  • Explore possible role played by atypical BTV in the BT epidemiology.

• Conditions that favour spread

  Presence and abundance of Culicoides vectors and high livestock density in an area. High temperatures which enhance virus development in and transmission by vectors. High temperature and relative humidity, wind conditions which can blow the vector into new areas. Naive host populations; introduction of exotic BTV strains.

• Detection and Immune response to infection

• Mechanism of host response

  Complex.Humoral with neutralizing and non-neutralizing antibodies. Cell-mediated immunity.Inflammation.Immunity (naturally acquired after infection or induced by vaccination) against one serotype is often ineffective or less effective in case of infection due to a different serotype.BTV can persistently infect ovine γδ T-cells in vitro, a process that may also occur during infection in mammalian hosts, thus providing a mechanism for virus persistence.BTV can infect other cellular components of the mammalian immune system (dendritic cells), which may be involved in dissemination of the virus in the mammalian host. Cleavage of virus surface proteins by host protease enzymes associated with inflammation generates infectious sub-virus particles that have enhanced infectivity (100 times) for the insect vector.The nature of the antiviral response in the insect vector is almost entirely uncharacterised.

  GAPS:

  • Increase knowledge concerning host immune responses to BTV, identify correlates of protection in vaccinated animals and determine immune evasion mechanisms as well as the role of immunity in pathogenesis of disease.
  • Increase understanding of the immune responses to BTV to improve vaccination strategies: including identification of viral epitopes involved in type specific and cross-reactive immune reactions and protection;
  • Identification of mechanisms by which viruses evade host innate and adaptive immune responses.
  • Understanding the mechanism of prolonged viraemia. Where is the virus for each serotype? In which form (infectious or non-infectious virions)? Where does the virus replicate in mammals?
  • What are the receptors in mammals and insects for all serotypes
  • What is the importance of cell-mediated immunity and the precise mechanisms for protective immunity?
  • What viral proteins / sites are involved in a protective cell mediated response.
  • What are the activated signaling pathways during infection and/or vaccination?
  • In insects, what is the nature of the innate barriers? Are some viruses able to kill the vector? (this is the case for other arboviruses).
  • Are acquired immune mechanisms induced during infection in insects?

• Immunological basis of diagnosis

  Neutralizing antibodies against VP2 and VP5. Antibodies against VP7 are detected in the current commercial ELISA tests. Antibodies against other conserved viral proteins could also serve as a basis to indicate a previous infection. Cellular immune responses against NS1, VP-2, VP3, VP5 and VP7.

  GAPS:

  • Development to serotype-specific ELISA assays.
  • The ability of existing ELISA to detect novel virus types and topotypes requires further investigation.

• Main means of prevention, detection and control

• Sanitary measures

  Quarantine and serological surveillance; vector control; zoning

  The most important sanitary measure to avoid the introduction of BTV in a free country is the testing and safe importation of live animals (and semen and embryos).

  To control the disease and the spread of infection: prompt reporting of BT outbreaks; vaccination; presence of appropriate serological and entomological surveillance and monitoring programme

  GAPS:

  • Development of vaccines against all BTV serotypes / topotypes.
  • Development of cross reactive vaccines

• Mechanical and biological control

  The measures to control and eradicate the disease include vector control, (use of insecticides in the animal premises and in the areas where these insects live, insect repellents onto animals, mosquitoe nets, etc.), restriction of movements of live ruminants from affected areas to non-infected regions where the vector is present and the use of vaccines.

  Restriction of movements and the use of vaccines are the most important control measures.

  GAP:

  Development of more effective insecticides with shorter withdrawal period and protective measures like impregnated netting.

• Diagnostic tools

  Antibody detection: ELISAs, Virus Neutralisation.

  Antigen detection: Real time RT-PCR or conventional RT-PCR, type specific RT-PCR, virus isolation and direct ELISA.

  Serogroup specific conventional and real-time RT-PCR assays are available.

  Serotype-specific PCR assays exist for all serotypes.

  DIVA vaccine for BTV are not yet available.

  GAPS:

  • Development of serotype specific ELISAs.
  • Check cross reactivity between topotypes
  • Further testing and validation of DIVA serology assays in combination with DIVA vaccine for all BTV serotypes by exchanging the Seg-2 of a common genomic backbone.
  • Development of molecular-based DIVA assays in combination with DIVA vaccines.
  • RT-PCR assays need to be tested / validated against a wider range of strains and updated as required.
  • To develop innovative diagnostic tools, including microarrays and single tube/bead/chip based multiplex assays (able to identify different virus serotypes or vectors at the same time). Mixed samples, containing multiple serotypes, can be missed by current diagnostic methodologies/work flows.

• Vaccines

  Although a number of vaccine strategies have been investigated with promising results, nowadays, live attenuated and inactivated vaccines remain the only vaccines commercially produced and used to prevent BT.When vaccinating against BT, the serotypes included into the vaccine must be the same as those causing infection in the field.Different vaccines have been used since the disease appeared in the EU. In the first BT outbreaks, sheep in France were vaccinated with live attenuated vaccines against serotypes 2, 4 and 16 whereas in Spain they were vaccinated against serotypes 2 and 4. In Italy, from 2002-2005, domestic ruminant (cattle, goats and sheep) population was vaccinated with live attenuated vaccine against serotypes 2, 4, 9, 16. The use of these ‘live’ vaccines does have some drawbacks. They may revert to virulence or may already be virulent in naïve populations; they may induce abortion when given to pregnant females, they cause viraemia and can circulate in the field in Culicoides midge populations and the vaccine virus may undergo reassortment with circulating field strains of another serotype or topotype. However, despite these drawbacks, live attenuated vaccines, if produced and used properly, can successfully control and, in some circumstances (Balearic islands), eradicate the infection. These vaccines have been used successfully for many years to protect animals in endemic areas (e.g. in Southern Africa). Inactivated vaccines to BTV-2 and BTV-4 became available in 2005 – 2006 as an alternative to live attenuated vaccines. From 2007 / 2008 inactivated vaccines against BTV-1, BTV-8 and BTV-9 came onto the market and, during 2008, a mass vaccination campaign was introduced in many EU countries against BTV-8 and BTV-1, which has been very successful. Vaccination was efficacious also when used during the recent BTV-4 incursions. In the last few years, however, because of the incursions of BTV strains with minimal occurrence of disease, many infected countries use vaccination for animal movement only. No inactivated vaccines against BTV-3 are currently available.The “next generation” strategies, many with DIVA capability, did not have the chance to be launched on the market or tested on a large scale. They include non-replicating subunit vaccines and virus-like particles, heterologous microbial expression vectors (e.g. using poxviruses as expression vectors for immunogenic BTV proteins), and genetically engineered LAVs, including replicating but non-transmittable virus-based vaccines (DISC and DISA).

  GAPS:

  • Development of vaccines that protect against all BTV serotypes. This could be achieved through cross reactive vaccines, or serotype specific vaccines against all serotypes.
  • Validation of chimeric vaccines for each of the BTV serotypes.
  • Increase understanding of the immune responses to BTV to improve vaccination strategies: including identification of viral epitopes involved in type specific and cross-reactive immune protection.
  • Further understanding is needed of the significance of strain / topotype variation in the specificity, efficacy of neutralizing antibody and cell mediated responses.
  • More work is required in the development / evaluation of novel / appropriate antigen delivery platforms / adjuvants.

• Therapeutics

  No specific treatment is available, other than supportive care.

  GAP:

  A better understanding of the biochemistry and replication of the virus could lead to opportunities to develop antiviral agents. However, it is not considered likely that these will play a major role in protection against BTV infection in the field.

• Biosecurity measures effective as a preventive measure

  Council Directive 2000/75/EC lays down control rules and measures to combat bluetongue in the EU, including the establishment of protection and surveillance zones and a ban on animals of the susceptible species leaving those zones. To address these issues Commission Regulation (EC) No 1266/2007 on implementing rules for Council Directive 2000/75/EC as regards the control, monitoring, surveillance and restrictions on movements of certain animals of susceptible species in relation to bluetongue has been adopted.Since bluetongue is considered to be non-contagious, isolation of infected animals is not necessary or appropriate for disease control. Other measures could be adopted in order to reduce the contact between hosts and vectors.Ensuring no contact between vector and susceptible animals (insect -repellent, housing of livestock during the times when the vector midges are most active, screening of access points (windows, doors) using fine mesh capable of excluding midges, or coarser mesh impregnated with a suitable insecticide such as a synthetic pyrethroid.

  GAP:

  Development of more effective insecticides with shorter withdrawal period and protective measures like impregnated netting.

• Border/trade/movement control sufficient for control

  EU regulation based on animal movement and vector control.Vaccination, serological / entomological surveillance and monitoring programs.

  GAPS:

  • Appropriate size of control and surveillance zones in case of BT outbreaks.
  • The mechanism for long distance movement of BTV strains (e.g. BTV-8 to northern Europe) still needs to be understood. Without this information it is difficult to develop appropriate controls/regulations to prevent further similar events.

• Prevention tools

  Movement controlsVaccination

  GAP:

  See under “Vaccines”.

• Surveillance

  Member States should design surveillance program to confirm that BTV is not circulating in the country, to early detect possible new introduction and to identify infected areas. Susceptible animals should be tested before moving and clinical disease reported.Entomological surveillance should be put in place in order to acquire information on the species and abundance of Culicoides populations which will be useful to define 'vector-free' or 'low vector activity' season.

  GAPS:

  • Surveillance strategies taking into account limited resources and local conditions of disease and vector occurrence as well as climatic and environmental conditions.
  • All assays will need continued evaluation/validation, to ensure they are effective for target/novel virus strains.

• Past experiences on success (and failures) of prevention, control, eradication in regions outside Europe

  BT vaccines may be used for different purposes or vaccination strategies, depending on the epidemiological situation of the affected area and desired objectives (i.e. disease control or disease eradication). The main objectives of BT vaccination strategies are to (i) prevent clinical disease, (ii) reduce the spread of BTV, (iii) eradicate BT from the country or the region, and (iv) permit the safe movement of susceptible animals between BT-affected and BT-free zones. These goals have guided BT vaccination campaigns since the incursion of BTV into Europe, enabling the eradication of serotypes 1, 2 and 8 in some regions. It is therefore commonly accepted that vaccines can help limit the spread of BTV and eradicate the infection in some instances. A vaccination campaign aiming at eradication should target susceptible ruminant species, achieve a high degree of herd immunity (80%), and include extensive areas surrounding any active BT outbreak. Such campaigns should also take into account climate, geography, and the abundance of competent insect vectors and susceptible wildlife animals.A successful control in fact requires the contemporaneous appliance of both vaccination of susceptible animal and restricting movement of viraemic animals between BT-affected and BT-free zones.Vaccination is then effective in reducing mortality and morbidity. However, once BT is endemic, it is difficult to eradicate, although it does also spontaneously clear without any vaccination after incursions of single serotypes – e.g. Canada.Successful eradication has been achieved (Spain, BTV-2 and BTV-4, Balearic Islands) through high vaccine coverage (>80%) over an extended period of time (3-5 years) by using live modified vaccine.Vaccination with inactivated vaccines in the recent northern European outbreak has been extremely successful, for example the UK completely prevented re-emergence of the BTV outbreak in 2008. The number of infected farms in France was also reduced from 29,000 to <100 between 2008 and 2009, but BTV-8 re-emerge in 2015. This was achieved through high levels of vaccine coverage (>80%) using a compulsory vaccination programme. Similar reductions in the incidence of disease have been achieved in Holland, Germany, Belgium and the UK through vaccination.According to a recent opinion document by EFSA, even when the vaccination of 95% of the susceptible cattle and sheep is constantly applied for three consecutive years, BTV is not eradicated and may re-emerge after a couple of years. Only after 5 years of vaccination of 95% of susceptible cattle and sheep, the prevalence of infection is close to eradication levels.

  GAP:

  Modelling the use of vaccination as an effective control measure.

• Costs of above measures

  The total net cost of the BTV-8 surveillance and vaccination programs arising from the outbreak amounted to €22.8 million.

• Disease information from the WOAH

• Disease notifiable to the WOAH

  Yes.

• WOAH disease card available

  Yes.

• WOAH Terrestrial Animal Health Code

  Infection with Bluetonge Virus

• WOAH Terrestrial Manual

  Infection with Bluetongue Virus

• Socio-economic impact

• Zoonosis: impact on affected individuals and/or aggregated DALY figures

  None.

• Zoonosis: cost of treatment and control of the disease in humans

  None.

• Direct impact (a) on production

  A global estimate of the impact of Bluetongue was US$ 3 billion. Losses due to any livestock disease may be classified as losses in production (direct losses), expenditure and lost revenue (indirect losses). Direct economic losses which include loss of productivity, loss of milk yield, abortion, loss of fertility, depends on the severity of the outbreaks which, in turn, depends on many variables including virulence of the BTV strain, immune status of the animal population etc… The direct costs of the recent BTV-8 incursions ranged from 30 to 160 million of Euro depending on the country and year.

• Direct impact (b) cost of private and public control measures

  Cost of vaccine, costs of application, testing, impact of movement controls, reduced ability to export or to move within a country. As said before, the total net cost of the BTV-8 surveillance and vaccination programs arising from the outbreak amounted to €22.8 million.

• Indirect impact

  Indirect losses include costs of vaccines or lost revenue, such as through trade restrictions limiting access to higher value markets. The overall estimates on the financial impact of the BTV-8 2007 outbreak, in France and the Netherlands, were US$ 1.4 billion and US$ 85 million, respectively. The costs are largely ascribed to the trade restrictions that were present during the outbreak period.

• Trade implications

• Impact on international trade/exports from the EU

  Preventing the spread of disease through international trade is one of the primary objectives of the Office International des Épizooties (OIE), the World Organisation for Animal Health. This is accomplished by establishing international standards that facilitate trade while minimising the risk of introducing diseases such as BT. Chapter 2.1.9 of the Code outlines the requirements that should be met for a country or zone to be defined as free of BTV and the sanitary measures that should be applied when importing live animals, semen and embryos into a BTV-free country or zone. Cost estimates have been reported in the previous paragraphs.

• Impact on EU intra-community trade

  Control rules and measures to combat bluetongue in the EU, including the establishment of protection and surveillance zones and a ban on animals of the susceptible species leaving those zones have been included in the Council Directive 2000/75/EC and in the Commission Regulation (EC) No 1266/2007. Costs derived from this ban have been detailed in previous points. To avoid these economic losses MSs can stipulate ad hoc agreements on animal trade according to the Commission Regulation (EC) No 1266/2007 art. 8, par 1b.

• Impact on national trade

  The duration of BTV viraemia in domestic ruminants has been a critical issue in international trade and placement of trade barriers. The OIE currently recognizes a 60 day infective period.In case of presence of restriction zones for different BTV serotypes or the contemporaneous presence of restriction and BTV-free areas in a country, ban or restriction measures on animal movement can have a dramatic effect on its livestock sector. For this reason, National authorities prefer, if possible, to expand restriction areas in order to have a unique BTV homogeneous restriction zone within which animal movement is allowed.

• Main perceived obstacles for effective prevention and control

  Diversity and variability of virus strains / serotypes / topotypes. Potential for reassortment.Diversity of vector (many 100s of potentially competent Culicoides species).Domestic and wild ruminants act as reservoirs with a long-lasting viraemiaVector control almost impossibleLikely to be at least 27 serotypes, vaccines are serotype specific and are not available for all serotypes (live-attenuated vaccines are multi-serotype vaccines).Uncertain elements of the BTV life cycle, including potential role of vertical and/or oral transmission of animals.

  GAPS :

  • Further understanding the degree of reassortment that is occurring in the field in areas where BTV is endemic.
  • Further understanding of the diversity and global distribution of different virusstrains /serotypes / topotypes.
  • Further understanding of cross-reaction and potential for cross protective vaccines between different strains, and the underlying immune mechanisms involved.

• Main perceived facilitators for effective prevention and control

  There should be close links between laboratories, international organizations, governments and industry.Most introductions of BTV in the last decade have occurred through repetitive ‘gateways’, it would be sensible to make the most efficient use of limited surveillance resources by targeting known routes of viral introduction.Continuous monitoring of the epidemiological situation of these countries is then recommended to identify serotypes posing the greatest risk for livestock. Establishing a vaccine bank for those serotypes circulating in countries geographically close would be extremely valuable. This would advance development of vaccines against those serotypes.

  GAPS:

  • Studies on the duration of viraemia in susceptible species, the determination of the infective titre and the mechanisms of overwintering would help to facilitate prevention and control. Virus is relatively well known but there is a need to develop new safe killed / recombinant vaccines against all the BTV serotypes.
  • In particular the development of effective cross serotype / cross topotype vaccines would be extremely useful and could potentially lead to an effective wider eradication campaign.

• Links to climate

  Seasonal cycle linked to climate

  In many parts of the world, infection has a seasonal occurrence.Climate, particularly ambient temperature has an impact on the Culicoides life cycle and survival of the vector. It may also impact on the development of BTV in the vector.

  GAP:

  Culicoides life cycle, BTV development in the midge and seasonal distribution of BT outbreaks taking also into account environmental conditions, farm/animal density in different parts of Europe / the world.

• Distribution of disease or vector linked to climate

  For the typical BTV serotypes, presence and abundance of vectors are associated with climate and environmental conditions and, as a consequence, occurrence of disease is also linked to the vector. Warmer seasons may increase vectorial capacity.

  GAP:

  Modelling the potential effects of climate change on the distribution of culicoides vectors and BTV into the future.

• Outbreaks linked to extreme weather

  Under favourable conditions, however, some biting midges can live long enough to survive the period between two vector seasons . Warmer winters in Europe may enable the adult Culicoides midges to overwinter more frequently. However, vector Culicoides can travel by wind for 100s of kilometres, thus increases in wind movements may enable colonization of new habitats by vectors and the rapid spread of BTV to areas remote from the source.

• Sensitivity of disease or vectors to the effects of global climate change (climate/environment/land use)

  Environmental factors profoundly affect vectorial capacity, governing dynamics and intensity of vector-vertebrate contact in time and space (e.g. seasonal vector population densities, biting rates, and feeding frequencies). Temperature influences vector developmental rates and life history parameters, and may modify vector competence. Vector Culicoides breeding and survival is governed by ecological factors, such as rainfall, temperature, humidity and soil characteristics. BTV infection and transmission by vectors is enhanced by high temperature and by vector survival rates. Any impact of climate change on any of these factors will potentially alter the distribution and spread of the diseaseIncreasing environmental temperature (climate change) will also extend the vector season.Vector competence of Culicoides vectors for Orbiviruses is partly controlled by temperature. Orbivirus development in Culicoides vectors is unable to occur at temperatures below about 10°C to 15°C depending on the Orbivirus species and strain. Furthermore, there needs to be a minimum amount of time at suitable temperatures (expressed as “day degrees or hour degrees”) for completion of the development cycle in the Culicoides vector before virus transmission can occur.The replication of the BTV viral RNA is entirely mediated by the viral polymerase, which is active over the range 10 to 45 degrees, with an optima around 31 degrees. This may help to explain the temperature dependence of BTV replication in the vector insects.

  GAPS:

  • Investigate the vector competence of different Culicoides species and how this is affected by increases in temperature.
  • Investigate how the replication of different virus strains/lineages responds to changes in ambient temperature, and the molecular basis for such variations.

Risk

• The spread of BTV (such as BTV-8) across the whole of Europe confirms the presence of suitable vectors in each of the affected countries. The whole of Europe must therefore be considered at risk from further incursions of BTV and other Culicoides transmitted orbiviruses, such as AHSV, EEV, EHDV. Local climate change could lead to increasing local temperatures, exacerbating these risks.In infected areas, monovalent live virus vaccines or subunit vaccines could potentially allow/force selection of new variant strains of BTV through genetic drift or shift (reassortment).Co-circulation of different BTV strain as well as typical and atypical BTV strain could also potentially allow/force selection of new variant strains of BTV through genetic drift or shift (reassortment).Reassortment can occur between existing field strains in the field in Europe, including eastern and western lineages or typical and atypical BTV strains, potentially leading to the emergence of progeny strains with novel biological characteristics. Mechanisms for spread of BTV are thought to include illegal movement of viraemic animals and wind translocation of infected vector Culicoides spp.Either new or different serotypes/strains continue to invade Middle East and Northern African countries which represent the most important gateways for the access of BTV in Europe.Traditional control measures for BT include animal movement restrictions, vector control, slaughter of viraemic animals, and management to reduce animal:vector exposure.

  GAPS:

  • Additional research is needed regarding use of vaccines during outbreaks.
  • Emergence of a novel reassorted BTV strain during a mixed infection of two parental strains, implies that it has some selective advantage over both parents and other progeny strains generated in the same outbreak. This implies that the new virus could have enhanced transmission or replication characteristics that might make it more dangerous.
  • Several reassortant strains have been identified in Europe. However, the potential for existing BTV strains in north Africa / the Mediteranean region /Europe to generate new strains, and the threat that these novel viruses might pose has not been fully evaluated.

Conclusion

• No region is immune from the negative impact of climate change. The Mediterranean region is also vulnerable to climatic changes and it is expected that the incidence of vector borne diseases including BT in the region will increase in the next coming years. Incursions of new strains/serotypes are continuously observed in North Africa and Middle East. The presence of these new strains/serotypes adds further genetic variability to the pool of BTV circulating genomes increasing the risk of creating reassortant strains. In addition, as the recent BTV incursions have demonstrated, many BTV strains that have circulated in Northern Africa have, soon or later, spread to Europe causing severe economical losses to European livestock.For these reasons, BT remains a major health and trade problem for livestock industries. In infected countries, BT is a priority for Veterinary Services. Surveillance as well as vaccination remains principle tools for prevention and control, depending on the context. A number of vaccines and diagnostic tests are available in Europe and worldwide but technological advancement in both domains would be desirable. Due to a relatively high numbers of products on the market, it is unlikely that industry will invest in new technologies, unless external funding sources can be mobilized within the context of formal collaborations.

Sources of information

• Expert group composition

  G. Savini, OIE Reference Laboratory for Bluetongue, IZS dell’ Abruzzo e Molise “G. Caporale”, Italy – [Leader]

  J.N. MacLachlan, University of California, USA

  P. Hudelet, Boehringer Ingelheim, France

  A. Lubisi, OIE Reference Laboratory for Bluetongue Onderstepoort Veterinary Institute, South-Africa

  E. Ostlund, OIE Reference Laboratory for Bluetongue, National Veterinary Services Laboratories, VS/APHIS/USDA, USA

  A. Laddomada, IZS della Sardegna, « G. Pegreffi », Italy

  P. Calistri, IZS dell’ Abruzzo e Molise “G. Caporale”, Italy

  C. Batten, EURL and OIE Reference Laboratory for Bluetongue, The Pirbright Institute, UK

  B. Hoffman, Friedrich-Loeffler Institut, Germany

  S. Zientara, ANSES/INRA/ENVA, France

• Reviewed by

  Project Management Board.

• Date of submission by expert group

  5 April 2019

• References

  OIE

  http://www.oie.int/eng/normes/MMANUAL/A_index.htm

  http://www.oie.int/eng/maladies/fiches/a_a080.htm

  http://www.oie.int/eng/ressources/en_diseasecards.htm

  http://www.oie.int/eng/maladies/en_alpha.htm?e1d7

  OIE –BT-Labnet website

  http://oiebtnet.izs.it/btlabnet/

  Institute for Animal Health Websites:

  http://www.iah.bbsrc.ac.uk/

  http://www.bluetonguevirus.org/

  http://www.culicoides.net/culicoides

  http://www.reoviridae.org/dsRNA_virus_proteins/

  Defra

  http://www.defra.gov.uk/animalh/diseases/vetsurveillance/az_index.htm