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Development and field validation of a rapid immunomagnetic separation - lateral flow (IMS-LF) test for detecting Mycobacterium bovis infection in badgers and/or badger setts - SE3271

Description
The proposed project aims to address the DEFRA research requirement in relation to development of a more rapid diagnostic method for identifying Mycobacterium bovis (Tuberculosis) infection in badgers, or identifying setts which contain infected badgers, that can be used in the field.

During the course of a previous DEFRA funded research project at Queen’s University Belfast (QUB) an immunomagnetic separation (IMS) method for selectively capturing and concentrating M. bovis, the cause of bovine tuberculosis (TB), from animal tissue samples was successfully developed. This technique involves mixing microscopic paramagnetic spheres coated with M. bovis-specific antibody and peptide with a sample that may contain M. bovis cells. Any M. bovis cells present will bind to the beads so that when a magnet is applied the spheres plus M. bovis cells can be pulled out of suspension and trapped on the side of the sample tube. After washing the beads and resuspending them in a smaller volume of buffer the M. bovis cells can be detected by a variety of methods, including culture, polymerase chain reaction (PCR), ELISA, and phage amplification assay.

The goal of this project will be to develop a rapid endpoint detection method that is simple enough to be applied in the field and sensitive enough to detect low level M. bovis infection in badgers/badger setts. Phase 1 of the proposed project will be concerned with producing a prototype lateral flow device (LFD, not dissimilar to a pregnancy test format) incorporating M. bovis antibodies or peptides (products of the earlier DEFRA funded project) and proving that it can sensitively detect low numbers of laboratory-grown M. bovis added to badger faeces. Subsequently the LFD test will be used in conjunction with the previously developed IMS technique, as an IMS-LFD assay, to test for M. bovis in badger faeces samples. Phase 2 of the proposed project will be concerned with proving that the novel IMS-LFD assay is capable of detecting M. bovis infection in badgers by testing 500 faeces samples collected from around 100 badger sett major latrines on 50 TB infected and 50 non-infected farms throughout Northern Ireland, and comparing its performance in the field with established detection methods (culture or phage assay to detect viable M. bovis, and PCR to detect M. bovis DNA) carried out subsequently on the same samples in the laboratory.

The proposed research is to be primarily carried out in Northern Ireland (NI). In relation to badger ecology the NI situation differs from Great Britain (GB) mainland situaiton in a number of respects (landscape, hydrology, land use, field size and topography) which in practice means that NI badger sett size tends to be smaller than GB. In order to ensure that the newly developed IMS-LFD test is equally applicable in the GB context, 50-100 well-characterised (in terms of TB status of animals) badger faeces samples from the Woodchester, York, badger study area will be supplied by the Food and Environment Research Agency (FERA) for IMS-LFD testing at QUB.

Outputs from the proposed project will be: a newly developed IMS-LFD test suitable for detecting M. bovis in the field; a standardised protocol for application of the new test in the field; statistical analysis of the IMS-LFD results to determine how well the new field test performs relative to existing lab-based methods of detecting M. bovis in the NI and GB contexts; and, potentially, new information on the location of TB infected badger setts in NI, identified as such in course of the study.
Objective
1. Produce a prototype lateral flow device, incorporating either a monoclonal antibody or specific recombinant peptides, capable of sensitively detecting Mycobacterium bovis.

2. Combine M. bovis-specific immunomagnetic separation (IMS) and LFD to produce a novel IMS-LFD assay capable of detecting low numbers of M. bovis cells in badger faeces.

3. Compare performance of the new IMS-LFD test applied in the field to test badger faeces and latrine samples for evidence of M. bovis infection with results of parallel laboratory testing using IMS in conjunction with MGIT culture, phage assay and PCR.

4. Verify that new IMS-LFD test is applicable to badger faeces samples from England and Wales.

Project Documents
• EVID4 - Final project report : SE3271   (659k)
Time-Scale and Cost
From: 2012

To: 2013

Cost: £316,856
Contractor / Funded Organisations
Queen’s University
Keywords
Animal Diseases              
Animal Health              
Diagnosis              
Plants and Animals              
Tuberculosis              
Fields of Study
Animal Health