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Microb Biotechnol. 2019 Dec 3. doi: 10.1111/1751-7915.13518. [Epub ahead of print]

The development and use of Actiphage® to detect viable mycobacteria from bovine tuberculosis and Johne's disease-infected animals.

Author information

1
Pathobiology and Population Sciences, Royal Veterinary College, Hawkshead, Herts, AL9 7TA, UK.
2
School of Biosciences, University of Nottingham, Sutton Bonington Campus, Loughborough, Leics, LE12 5RD, UK.
3
School of Veterinary and Medicine Science, University of Nottingham, Sutton Bonington Campus, Loughborough, Leics, LE12 5RD, UK.
4
Moredun Research Institute, Pentlands Science Park, Penicuik, EH26 0PZ, UK.

Abstract

Here, we describe the development of a method that exploits bacteriophage D29 as a lysis agent for efficient DNA extraction from low numbers of mycobacterial cells. This method (Actiphage® ) used in combination with PCR achieved rapid and sensitive (LOD ≤ 10 cell ml-1 ) detection and identification of viable, pathogenic mycobacteria in blood samples within 6 h. We demonstrate that mycobacteriophage D29 can be used to detect a range of mycobacteria from clinical blood samples including both Mycobacterium tuberculosis complex and Mycobacterium avium subsp. paratuberculosis without the need for culture and confirms our earlier observations that a low-level bacteraemia is associated with these infections in cattle. In a study of M. bovis-infected cattle (n = 41), the sensitivity of the Actiphage® method was 95 % (95 % CI; 0.84-0.99) and specificity was 100 % (95% CI; 0.92-1). We further used Actiphage® to demonstrate viable Mycobacterium avium subsp. paratuberculosis is present in the blood of Johne's infected cattle. This method provides a revolutionary new tool for the study of infections caused by these difficult to grow pathogens.

PMID:
31793754
DOI:
10.1111/1751-7915.13518
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