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Optimisation of sampling strategies for improving sensitivity of M. bovis detection by PCR - SE3280

The proposed research aims to identify an optimal regime for badger faeces sampling, maximising the sensitivity of a previously validated PCR test for Mycobacterium bovis, the causative agent of bovine tuberculosis (bTB). Previous work by this team demonstrated that a quantitative real-time PCR test identified a proportion of putative positive setts at Woodchester Park when compared to Fera data for the social groups tested. Of 11 social groups covered by PCR testing and Fera trapping, nine were found to be infected using several diagnostic methods on samples taken from trapped animals. Of these nine, the previous PCR ring trial identified two with a third putative positive at one laboratory. The level of positivity identified by PCR was approximately 22% which is inline with conventional culture based methods although less than STATPAK which identified 78% of positive setts over four trapping episodes in 2009, and routinely identifies approximately 50% (Fiona Stuart, pers. comm.). Badger latrine sampling for the PCR ring trial was carried out in July and August 2008 on a single day for each social group, which was a suboptimal strategy for maximising test sensitivity but was adequate for validating the reproducibility of the test.

Further work will consist of intensive latrine sampling at periods of peak badger activity, combined with sampling over a one year period matching Fera trapping studies to provide a definitive data set for modelling optimal sampling regimes. The PCR test will also be used on faeces samples collected from trapped badgers, allowing exact comparisons with shedding rates estimated by cultivation at Fera.

Heterogeneity of M. bovis distribution with in single droppings will be investigated using material from captive badger populations at AHVLA which have tested positive by culture. This will allow modification of sampling strategy, with multiple samples from single droppings or homogenisation methods used, if heterogeneity in spatial distribution is shown.

These data sets will allow an improved sampling strategy to be calculated using computer simulations and will also demonstrate PCR test sensitivity in direct comparison with conventional testing methods at Fera.

Accurate investigation of PCR test sensitivity combined with matched comparisons with conventional trapping based testing will elucidate the utility of the PCR test as a practical non-invasive management tool for identifying badger social groups containing infectious animals.
The main aim is to improve sensitivity of the PCR test for M. bovis by optimising latrine sampling
strategies. The proposed research will also allow a rigorous comparison of the PCR test with
conventional trapping and diagnostics, elucidating the utility of the PCR test as a non-invasive tool to
identify infectious badger social groups:

1. Cross-sectional sampling over a 12 month period matched to live trapping periods.

2. Intense longitudinal sampling of the same territories above every 2 days over 10 days in peak badger activity periods- September/October 2011 and February/March 2012.

3. To identify the distribution of M. bovis within droppings to improve detection probability if high special heterogeneity is observed.

4. FERA will retain subsamples of faeces from trapped badgers which will be screened using the PCR assay for comparison with culture.

5. Simulated sub-sampling of data sets to identify optimal sampling strategy, in terms of time of year, numbers of samples, regularity of repeat sampling.
Project Documents
• FRP - Final Report : 20140207 SE3280 Final Report Feb 7th 2014   (1558k)
Time-Scale and Cost
From: 2012

To: 2014

Cost: £467,353
Contractor / Funded Organisations
Veterinary Laboratories Agency (VLA), University of Warwick
Animal Diseases              
Animal Health              
Plants and Animals              
Fields of Study
Animal Health