Introduction

Bluetongue, an arboviral disease, caused by the bluetongue virus (BTV) naturally infects domestic (sheep, goats, cattle) and wild ruminants (deer, pronghorn antelope, bighorn sheep), camelids and some other herbivores such as elephants (Calistri et al., 2015). Bluetongue can cause severe outbreaks and thus reasonably placed in one of the few (16 in total) diseases classified in List A by the Office International des Epizooties (OIE, 2000). BT was first described as “Malarial Catarrhal Fever” and “Epizootic Catarrah of Sheep”. The name “bluetongue” was given (Spreull 1905; Bhatnagar et al., 1995) to describe the distinctive cyanotic tongue of some severely affected sheep. Now, bluetongue is endemic in India. Exotic sheep are more susceptible than indigenous and cross-bred sheep (Bayliss and Mellor 2001, Samal et al., 1987). However, outbreaks do occur in indigenous breeds as seen in Indian local breeds due to the prevalence of highly virulent strains. A serological survey has indicated the presence of bluetongue virus (BTV) antibodies in cattle and buffalo in several states in India. However, clinical BT has not been frequently observing in cattle or buffaloes. The rate of positively in buffaloes was higher than in cattle, and the incidence was higher in males than in females.

Virus

BTV is a member of the Orbivirus genus, currently one of nine genera classified in the family Reoviridae (Jain et al., 1986; Sreenivasulu et al., 1999). Within the Orbivirus genus, 14 groups are differentiated on serological basis. The virion is about 80nm in diameter and contains a genome composed of ten segments of double stranded RNA. BTV is relatively heat resistant and is partially resistant to lipid solvents but it can be rapidly inactivated at pH levels below 6.5 and above 8.0 and virus is very stable in the presence of proteins like blood (OIE: Technical card database).

Vector

Midges of the genus Culicoides act as biological vectors for bluetongue virus (Clavijo et al.,2000). Culicoides vector transmit BTV to and from susceptible animals, having become infected by feeding on viraemic vertebrates. After a replication period of 6-8 days, and following its appearance in the salivary gland, the virus can be transmitted to a vertebrate host during a blood meal. In India, although disease is endemic but detail studies on vector species responsible for disease and virus-vector relationship are lacking. Other vectors responsible for transmission include C. clavipalpis, C. anophelis and C. imicola (C. minutus) in West Bengal, C. anophelis andC. actoni in Uttar Pradesh, C. simiklis in Uttaranchal and C. orientalis in Gujarat.

Epidemiology

Bluetongue virus requires both in vertebrate and invertebrate hosts essential for its existence. There is no evidence of vertical transmission of the virus in the invertebrate host. Observations on the placental transmission of virus in the vertebrate host are contradictory (Byregowda et al.,2003).

Host Factors

BTV naturally infects domestic and wild ruminants, camelids, and some other herbivores such as elephants. It is clear that sheep and wild ruminants such as white-tailed deer are the species that are most susceptible to BT disease than any other ruminant species. Experiment showed BTV infection of the bovine endothelial cells resulted in endothelial activation, with the increased transcription of genes encoding a variety of vasoactive and inflammatory mediators and increased expression of cell surface adhesion molecules, similar infection of sheep endothelial cells resulted in minimal activation of endothelial cells. Furthermore, the ratio of thromboxane to prostacyclin, which is indicative of enhanced coagulation and possible consumptive coagulopathy, was significantly greater in sheep than in cattle that were experimentally infected with BTV (Prasad et al., 1994; DeMaula et al., 2016).

Vector Factors

Culicoides vectors are critical to the survival and transmission of BTV as infection is not contagious and there is no credible evidence of long-term maintenance of BTV in ruminants. The central role of the insect in BT epidemiology ensures that prevalence of the disease is governed by ecological factors, such as high rainfall, temperature, humidity and soil characteristics, which favors insect survival (Bonneau et al., 2000; Dahiya et al., 2003). The geographic distribution of BTV is categorized into three ecologic zones where the presence of bluetongue virus year round or seasonal, depends on the climatic zone type: endemic, epidemic and incursion. Endemic zones are defined as typically tropical regions where BTV transmission occurs throughout the year and subclinical infection is common. Epidemic zones include temperate areas where outbreaks occur seasonally, generally in the late summer when vector population is peak. Incursion zones are areas that experience outbreaks infrequently when climatic conditions favor disease transmission by vectors as observed in greatest outbreak of Mediteranian basin.

Economic Importance and Impact

Bluetongue is an economically important disease of ruminants for several reasons. The losses economic due to this disease is principally attributed to mortality and high morbidity, abortion, decreased quantity & quality of milk, meat and fleece losses in sheep. In endemic areas vaccination is a recurring cost. Moreover restriction in the free trade of cattle and other animals across different countries resulted in huge economic losses. Existence of bluetongue in the United States has been an impediment to moving cattle to Canada, New Zealand, and the European Union, which are classified by the OIE as being free of bluetongue; and also parts of Australia that have been regionalized as bluetongue-free (Mellor and Boorman 1995; Deshmukh et al.,2003).

Pathogenesis

Pathogenesis of BT disease in sheep depends on the virulence of the virus strains as well as the nutritional status, immune status, age of the animal and environmental stresses such as high temperature and ultraviolet radiation. After getting infection from the viraemic cattle, sheep or other susceptible ruminant’s culicoides these vectors become competent to transmit the disease to other susceptible ruminants after extrinsic incubation period of 7-14 days. After this period, these vectors transmit the disease at each time of talking blood meal. Then virus multiplies primarily in the regional lymph nodes which are followed by the viraemia where all the cells of the blood become infected (Bonneau et al., 2002, Malik et al., 2001). Systems virtually affected in BT disease include nervous, bone, skin, blood and circulatory system, reproductive, respiratory and digestive system (Animal Health Disease Cards, 2004). Endothelial injury leads to increase capillary permeability resulting oedema of various parts of the body especially in face region, larynx and lungs (Pini 1976). Affected animals frequently suffer pharyngeal or esophageal myodegeneration leading to dysphagia, regurgitation, development of bronchopneumonia. In severe cases, death of the animal occurs either from pulmonary edema, bronchopneumonia, cardiac insufficiency or any of these combinations.

Diagnosis

Laboratory tests are mainly performed for isolation of virus in blood, antigen detection on various isolation in tissues by ELISA, PCR and serum analysis for antibody by serological tests.

Virus Isolation in Culture

The following culture techniques may be used (given in order of sensitivity)-

1. Sheep Inoculation: not economical method, so not routinely practiced.
2. Intravenous Inoculation of Embryonated Chicken Eggs: it is the most sensitive technique for isolation of BTV. It is a prescribed test for international trade. Bluetongue virus may be detected in the inoculated eggs by antigen or nucleic acid detection procedures.
3. Intracerebral Inoculation of Newborn Mice: not routinely practiced.
4. Tissue Culture Method: cell culture isolates obtained are identified as bluetongue viruses by tests based on group-specific antibodies and using fluorescent or enzyme conjugates. Isolates placed in the bluetongue serogroup are typed by virus neutralization tests using serotype-specific antisera. Commonly, baby hamster kidney (BHK-21) and African green monkey kidney (Vero) cell cultures are used.

Serotyping by Virus Neutralization

The recommended ‘gold standard’ test for the detection of virus serotype-specific neutralizing antibodies are directed against outer membrane proper VP2, is the virus neutralization test.Neutralization tests are type-specific for the currently recognized 24 BT virus serotypes and can be used to serotype a virus isolate, or can be modified to determine the specificity of antibody in sera. The following techniques may be used- Microtitre neutralisation (most widely used, requires specific antisera for all BT virus serotypes), Plaque reduction, Plaque inhibition and Fluorescence inhibition are generally used for serotyping of BTV.

Serogrouping of Viruses

This is based on the reactivity of group-specific epitopes such as BT VP7. The following techniques are used-

• Immunocapture ELISA: (most widely used) Orbivirus isolates are typically serogrouped on the basis of their reactivity with specific standard antisera that detect proteins, such as VP7, that are conserved within each serogroup. Recently, a BT serogroup-specific monoclonal antibody (MAb) is used to rate out cross readability between membersw of oribivirus termily.
• Immunofluorescence
• Immunospot test
• Indirect peroxidase-antiperoxidase test

Nucleic Acid Recognition Techniques

Primer-directed amplification of viral nucleic acid has revolutionised BT diagnosis (Katz et al., 1993; Mehrotra et al., 2001). Results to date indicate that polymerase chain reaction (PCR) techniques may be used, not only to detect the presence of viral nucleic acid, but also to ‘serogroup’ orbiviruses and provide information on the serotype and possible geographic source (topotype or genotype) of BT virus isolates within a few days of receipt of a clinical sample such as infected sheep blood. Traditional approaches, which rely on virus isolation followed by virus identification, may require at least 3-4 weeks to generate information on serogroup and serotype does not and yield data on the possible geographical origin of the isolated virus. Nested PCR (nPCR) assay is increasingly used for screening of ruminants for the presence of BTV nucleic acids in tissues because it is highly sensitive, specific and extremely conservative assay by which BTV nucleic acid may be detected in the blood of sheep and cattle long after the virus has been cleared. It is a prescribed test for international trade. The “real time” PCR technology is faster than traditional PCR methods, and is less susceptible to problem of contamination. However,further validation is required is this regard as there may be problems in the identification of new strains of BTV with existing “real time” probes.

Serological Tests

Competitive or Blocking ELISA

It is widely used to measure BT virus-specific antibody without detecting cross-reacting antibody to other orbiviruses (Afshar et al., 1989). In the competitive ELISA, antibodies in test sera compete with the MAbs for binding to antigen. It is rapid, sensitive, and can be easily standardized and therefore, prescribed for international trade.

Agar Gel Immunodiffusion

This test to detect anti-BT virus antibodies is simple to perform and the antigen used in the assay is relatively easy to generate. Since 1982, the test has been the standard testing procedure for the international movement of ruminants. However, one of the disadvantages of the AGID used for BT is its lack of specificity in that it can detect antibodies to other orbiviruses, particularly those in the EHD serogroup. It is also a prescribed tests for international trade.

Complement Fixation: no longer widely used.

Fluorescent Antibody Technique (FAT) has been found useful for the detection of group specific antigen, but, not widely used.

Differential Diagnosis

Differential diagnosis in sheep should include foot and mouth disease, vesicular stomatitis, sheep pox, peste des petits ruminants, pneumonia, contagious ecthyma, photosensitization, copper deficiency, oestris ovis, ulcerative dermatosis, and Rift Valley Fever. In cattle, it should include foot and mouth disease, vesicular stomatitis, mucosal disease, mycotic stomatitis, infectious bovine rhinotracheitisc, akabane, bovine herpes mammilitis, malignant catarrhal fever, and bovine papular stomatitis.

Disease Prevention and Control

The interruption of bluetongue virus cycle is the most effective method of prevention and control. This can be achieved by following methods-

1. Movement Restrictions, Testing and Hygiene

In disease free countries, controlled importation of susceptible animals, serological testing of animals during quarantine period in vector-free area and importation of semen from bulls shown to be serologically BTV negative should be practiced. In disease prevalent countries, restriction of animal movement, slaughter and burn of infected stock, serological surveillance in area round infected zone, spray neighboring animals with long-acting insect repellent, house animals at night, reduce insect breeding sites by spraying or improved drainage, and spray buildings with insecticide should be undertaken (Mehrotra et al., 2000; Udupa et al., 2003). Disinfecting premises is not appropriate since virus is mainly transmitted by insects. After blue tongue infection, infected animal can remove from the heard and accumulating area. In some state animal cannot slater so we can remove from the healthy animal.

2. Vaccination

Of several vaccine options available, namely live attenuated, killed or recombinant, only attenuated virus vaccines are most effective and in current use in several countries.

Live Attenuated Vaccines

They induce an effective and lasting immunity. However, they must be type-specific, and therefore multiple vaccinations may be needed. Colostral immunity in young sheep can interfere with the development of active immunity to the vaccine and breeding ewes and rams should be vaccinated before mating. Most disadvantages of live attenuated vaccines are possible dangers of return to virulence and transmission through insect bites can occur apart from inducing teratogenesity and congenital malformations in the pregnant animals (DeMaula et al., 2017).

Recombinant Expressed Vaccines

It is not in use recently. There are several options for the development of recombinant BT virus vaccines, including live virus delivery of BT virus neutralisation antigens and the virus-like particles generated in infected insect cells by recombinant baculoviruses expressing the four major BT virus coat proteins, VP2, 3, 5 and 7 (DeMaula et al., 2017). Only the latter has shown significant promise.

Inactivated Vaccines

Due to problems encountered in complete inactivation of the virus, inactivate vaccines are still to be used in field condition. Again, duration of immunity is a matter of concern.

Acknowledgement

The authors are thankful to Dean, College of Veterinary Science and Animal Husbandry, NDUAT, Kumarganj, Faizabad for providing the funding and facilities to carry out the work.

Conflict of Interest

None of the authors of this paper have a financial or personal relationship with other people or organization that could inappropriately influence or bias the content of the paper.

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