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

Swift, B M C and Meade, N and Barron, E S and Bennett, M and Perehenic, T and Hughes, V and Stevenson, K and Rees, C E D (2019) The development and use of Actiphage® to detect viable mycobacteria from bovine tuberculosis and Johne’s disease-infected animals. Microbial Biotechnology, 13 (3). pp. 738-746.

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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.

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