Development of diagnostic systems, reference collections and molecular epidemiology studies for important arboviral pathogens of livestock in India

Lead Research Organisation: The Pirbright Institute
Department Name: Vector-borne Viral Diseases


This project "Development of diagnostic systems, reference collections and molecular epidemiology studies for important arboviral pathogens of livestock in India" will build on established links between colleagues at The Pirbright Institute, the University of Glasgow, The John Innes Institute and the Royal Veterinary College (in the UK) with The University of Veterinary and Animal Sciences at Hisar in India, creating links with colleagues working with arboviruses in southern India.
Outbreaks of arboviral diseases are a significant burden on livestock / farming communites in India. Serological data show that 21 of the 26 bluetongue virus (BTV) seroptypes exist in India, one of the highest levels of BTV diversity anywhere in the world. The severity of BT outbreaks in Indian sheep has increased significantly in recent decades, reaching 30% in some areas, possibly due to introduction of exotic strains. However, very few Indian BTV isolates have been genetically or phenotypically characterised, creating a major barrier to local validation of diagnostics and to control strategies, including development of vaccines.
During the past 25 years, multiple strains of BTV and other arboviruses have emerged in Europe, possibly linked to changes in climate and global trade. However, a lack of data concerning global strain-diversity and distribution limits our ability to understand, respond to and control these events. The incomplete nature of the global database for BTV limits our ability to Identify the geographic origin of BT outbreak-strains. Similar problems exist for many endemic arboviruses in India (Myers et al 1971; Padbidri et al 2002). We will address these knowledge-gaps by isolating, identifying and characterising arboviruses from India, to study their distribution, abundance, relationships, and movements that result in disease outbreaks. The primary focus of the project is BTV, although diagnostic samples used to detect and isolate viruses, will also provide materials and potentially isolates of other arboviruses from the region.
The reference collection at Pirbright provides different orbiviruses and sequence data, for development and evaluation of diagnostic assays (by RT-PCR). These assays, which identified the BTV types that invaded Europe since 1998, have become widely accepted 'front-line' tools for diagnosis, surveillance and typing of BTV around the world. However they have not yet been widely validated or deployed in India. BTV Isolates from the Pirbright collection were also used to develop / evaluate inactivated vaccine strains that successfully eradicated BTV type 8 from the UK and northern Europe .
Well characterised and documented virus isolates will be generated in India, as a basis for a reference collection of 'Indian arboviruses', focussing particularly on BTV and the other orbiviruses. This will provide materials for local development of relevant vaccines (seed stocks and challenge strains) and validation / further-development of diagnostic assays for identification, detection and surveillance of these viruses on the Indian subbcontinent. Linking 'Indian-reference-collections', to the existing collection at Pirbright, will add to the global resources and our knowledge concerning these viruses.
Diagnosis, isolations, sequencing, phylogenetics and virus storage will be undertaken in India, with assistance from Pirbright. Phylogenetic and evolutionary analyses will be performed at Pirbright and Glasgow. Pathology studies will be carried out at Pirbright, at the RVC and in India. The project will take advantage of recent developments in plant-based expression technologies at John Innes (JIC) (P3), to generate low-cost reagents for BTV serotyping. This will create links between plant and animal science in the UK and in India. Development of next generation serological assays will take place at JIC and Pirbright, for evaluation at Pirbright and in India.

Technical Summary

Outbreaks of arboviral diseases represent an important burden on livestock / farming communites in India as well as important threats to livestock industries and human health in Europe. Bluetongue virus (BTV) has spread to both southern and northern Europe ( including the UK) and has been followed by the novel Schmallenberg virus.
Twenty one BTV seroptypes (out of 26) exist in India, one of the highest levels of BTV diversity anywhere in the world. Phylogenetic analyses show clear relationships between two European BTV isolates with strains from India. Pilot studies also show introduction of exotic (western) strains into India, possibly linked to more severe disease in local sheep. However, few Indian BTV isolates have been sequenced or phenotypically characterised, creating barriers to local validation of diagnostic assays, development of control strategies, and our knowledge of transboundary virus movements / threats.
The project will make multiple arbovirus isolations, focussing particularly on southern India where disease outbreaks (particularly Bluetongue) are more frequent and severe. Based on expertise at the Pirbright Institute we will establish a well documented arbovirus reference collection (at Hisar and Hyderabad), with data links to existing collections at Pirbright. Sequencing of indian viruses will support phylogenetic, phylogeography and reassortment analyses. These Indian 'reference strains' will provide materials for vaccine development / testing, as well histpathology studies to help map phenotypic changes in strain virulence onto viral phylogenies.
Established RT-PCR based assays developed at Pirbright for detection, diagnosis and typing of BTV, related orbiviruses and other arboviruses will be validated in India. Relevant viral proteins will be transiently expressed in plant, from an Agrobacterium tumefaciens binary vector, for development of novel serological assays using ELISA and chip/bead (MagPix) based technologies.

Planned Impact

Impact summary
The Vector-borne Viral Diseases Programme (VVDP) plays a central role in research, diagnostic surveillance and control of the arboviral diseases, including zoonoses that threaten Europe and the UK. By helping to control and eradicate arboviral diseases of livestock, including those caused by high hazard exotic and zoonotic agents, the VVDP contributes to global food security, animal welfare, economic prosperity and human health. These activities have been of particular value during recent outbreaks of arboviral diseases (notably bluetongue in ruminants) in Europe, contributing directly to disease surveillance, advising on control strategies and contributing to development and evaluation of vaccines.
This project will transfer relevant technologies to set up similar diagnostic capabilities in India, ensuring that assay systems will work effectively with Indian virus strains, establishing virus reference collections to act as a source of materials for further study, development of diagnostic assays and development/evaluation of vaccines.
The project will generate nucleotide sequence data for local reference virus isolates as well as epidemiological data concerning the distribution, prevalence, movements and virulence of Indian strains. These data will be of importance to both local animal Health authorities, as well as (in view of the trans-boundary nature of these diseases) the international authorities involved in disease control and legislation (including those in Europe).

Beneficiaries in both the UK and India include the livestock industry (farmers, food industry, vaccine manufacturers / pharma companies), research organisations (e.g. BBSRC, MRC, institutes, universities), the international scientific community, disease control agencies (Defra, EU, OIE), developing countries and their supporters (FAO, DfID, GALVmed), and the general public. In particular the project will address diseases (notably bluetongue) that are an important problem for the farming industries in India (particularly in less wealthy communities) and to Europe. Staff at Pirbright has previously worked closely with both vaccine manufacturers and diagnostics companies in Europe and will generate/provide information and materials that would also be of value to similar organisations in India. It is anticipate that development of better diagnstics and control programmes for arboviral diseases in India, will be of direct benefit to the farming community particularly in less wealthy communities.

Monitoring impact
Impacts of the project will be monitored through formal annual reviews, which will document impacts from the past year. Regular telephone conference call will help to maintain links betrween specific Indian and UK partners (particularly Pirbright and Hisar).


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Description The research funded by this project has established state of the art technologies (developed at TPI) for isolation of bluetongue viruses (BTV) circulating in India. As a result the number of BTV isolates from India has increased from approximately 60 to over 700. These materials form a basis for an Indian BTV reference collection, providing materials to study the origins and movement of the viruses circulating in the Indian sub-continent.
The resulting reference collection and sequence database will support epidemiology studies to determine virus origins and movement, as well as vaccine matching to ensure the relevance of vaccines being used in the region.

As a result of these studies several papers have been published showing that strains of BTV derived from Africa and north America (western strains), have been introduced to India. These exotic strains are associated with an increase in the severity of the disease and increased mortality rates in local Indian breeds of sheep.

One of the genes, encoding a non-structural protein derived from western BTV strains, has spread rapidly through the Indian BTV population (by reassortment), and is associated with increased virulence in sheep, particularly in the South of the sub-continent. This indicates that this protein would be a valuable component of future subunit vaccines to help reduce the severity of the disease in sheep, both in India and elswhere.
Exploitation Route The Pirbright Institute has signed a Memorandum of Understanfing (MOU) to promote research links and collaboration with the Veterinary University at Hisar in Haryana, India.
The isolation technologies and diagnostic assay systems that have now established in Hisar and other laboratories in India will ehance the speed, and accuracy of surveillance for BTV.

The virus isolates and sequence data generated for India provide a basis for molecular epidemiology studies and can be used to ensure that the vaccines being used are relevant to the dominant serotypes of BTV circulating in the field (in India). This will help to reduce the severity of clinical disease, fatalities in sheep, and economic losses that are particularly important to rural Indian communities.
Sectors Agriculture, Food and Drink

Description This grant is still ongoing but it has already contributed significantly to the development of surveillance and control strategies for bluetongue outbreaks in India. In particular the more effective and more rapid virus isolation and diagnostic technologies that have been established there have identified ways in which the BTV vaccines in current use could be made more relevant and more effective against current strains circulating in the region.
Sector Agriculture, Food and Drink
Impact Types Economic

Description "Understanding pathogen, livestock, environment interactions involving bluetongue virus" (PALE-Blu)
Amount € 6,300,000 (EUR)
Funding ID 727393-2 
Organisation European Commission 
Sector Public
Country European Union (EU)
Start 06/2016 
End 11/2020
Description Development of diagnostic systems, reference collections and molecular epidemiology studies for important arboviral pathogens of livestock in India
Amount £338,317 (GBP)
Funding ID BB/L004690/1 
Organisation Biotechnology and Biological Sciences Research Council (BBSRC) 
Sector Public
Country United Kingdom
Start 04/2014 
End 06/2017