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LABORATORY DIAGNOSIS OF Mycobacterium bovis INFECTION AND STRAIN TYPING OF ISOLATES

This article discusses the laboratory methods for diagnosing Mycobacterium bovis infection in animals and typing the different strains of the bacteria. It includes information on histopathology, culture, and DNA typing methods, as well as the accuracy of each technique. The genetic relationship between strains in different countries is also examined.

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LABORATORY DIAGNOSIS OF Mycobacterium bovis INFECTION AND STRAIN TYPING OF ISOLATES

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  1. LABORATORY DIAGNOSIS OF Mycobacterium bovis INFECTION AND STRAIN TYPING OF ISOLATES • Eamon Costello • Central Veterinary Research Laboratory, Backweston, Celbridge, Co. Kildare

  2. Central Veterinary Research Laboratory Mycobacteriology section • National reference laboratory for tuberculosis in animals • Diagnostics Histopathology and culture of tissue samples Anamnestic antibody ELISA • Research • Tuberculin potency assays

  3. Samples submitted for TB Diagnostic tests • Lymph node lesions detected at abattoir post mortem examination of non-reactor cattle: - approx. 5,000 per year • Selected ‘singleton reactor’ cattle: - approx. 600 per year • Reactors from herds where DVO is undertaking epidemiological investigation: - approx. 800 per year

  4. Diagnostic methods • Macroscopic examination • Histopathology • Rapid diagnosis • Good correlation with culture (94%) • 85% of lesions diagnosed by histopathology • Culture • Solid media (Stonebrinks + LJ) • Liquid media (MGIT 960) • Blood agar

  5. Positive by culture Negative by culture Positive by Histopathology (N = 200) 188 (94%) 12 (6%) Negative by Histopathology (N = 100) 0 100 Accuracy of histopathological examination

  6. MGIT 960 Liquid Culture System • Liquid medium • Growth of mycobacteria leads to depletion of oxygen • Oxygen sensitive fluorescent chemical indicator in the culture tube • System for simultaneous incubation and monitoring of culture tubes

  7. Isolate identification proceduresGrowth characteristics • Growth rate and effect of temperature on growth • Colony colour and morphology • Cording

  8. DNA typing methods • RFLP analysis • 2000 isolates typed • Good differentiation of strains • Spoligotyping • PCR based method • 50% of isolates SB140 • VNTR • Results can be expressed in a • digital format

  9. SB 140 clonal complex 1101101000001110111111111111111111111100000 1101101000001110111111111111111100111100000 Ireland Great Britain Australia Canada Argentina SB120 clonal complex 1101111101111110111111111111111111111100000 1101111101111110111101111111111111111100000 Continental Europe Brazil Cameroon Chad Genetic relationship between strains in different countriesBased on spoligotyping

  10. Acknowledgements • Kevin Kenny • Frances Quigley • Don O’Grady • Orla Flynn • Tony Gogarty • Gillian Madigan • Jack McGuirk • Joanne McLernon • Damien O’Meara • Jane O’Rourke • Maureen Padden • Elvira Sovsic • Marion Teeling • Dermot Yearsley

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