Abstract

Avian mycobacteriosis generally has a low prevalence in most zoological collections, but it can have devastating consequences for collection and conservation program management. The management consequences are based on the assumption that avian mycobacterial infections are readily transmissible (directly or indirectly) from bird to bird, placing all exposed birds at high risk of eventually developing disease. Review of the literature reveals that this is only an assumption, apparently based on the long-recognized potential for clustering of cases. In humans and other species, molecular epidemiologic studies have shown that Mycobacterium avium infections are environmentally acquired and are not readily transmissible.^1,2 Case clusters in non-avian species are attributed to exposure to the same environmental source, rather than horizontal transmission. In this study, we test the assumption that Mycobacterium avium and Mycobacterium intracellulare infections are transmissible directly or indirectly between birds in captive settings.

We obtained 95 mycobacterial isolates from 62 birds representing 44 different species from the collections of the San Diego Zoo and San Diego Zoo’s Wild Animal Park. Isolates were cultured on BACTEC liquid media and LJ and Middlebrook agar. The initial identification of culture isolates was made with chemiluminescent cDNA probes specific for M. avium and M. intracellulare (Accuprobe, GenProbe, San Diego, CA, USA) and confirmed with biochemical tests. DNA was extracted from each isolate and the species identity of each isolate was confirmed by using a multiplex PCR that amplifies a region of the 16S rRNA gene specific to all members of the Mycobacterium genus and regions of the 16SrRNA gene that distinguish M. avium from M. intracellulare.^3,4 Eighty isolates were confirmed to be M. avium and 15 were confirmed to be M. intracellulare. Monoclonality of isolates was confirmed by PCR amplification of the 16S-23S rRNA internal transcribed spacer (ITS) region of Mycobacterium avium complex,5 and analysis of the ITS region by denaturing gradient gel electrophoresis (DGGE). Molecular strain typing was performed by two methods on each isolate: randomly amplified polymorphic DNA (RAPD)⁶, and amplified fragment length polymorphism (AFLP; using AFLP Microbial Fingerprinting Kit from Applied Biosystems, Foster City, CA 94404 USA)⁷. Band patterns obtained by agarose gel electrophoresis (RAPD) and automated capillary gel electrophoresis (AFLP) were compared and analyzed using GelCompar II software (version 4.5, Applied Maths, Kortrijk, Belgium).

Analysis of the band patterns revealed differences sufficient to suggest that Mycobacterium avium and Mycobacterium intracellulare infections in birds are primarily randomly acquired from the environment, as in other species. If direct or indirect bird-to-bird transmission does occur, it appears to be rare. Case clusters likely represent exposure to a common environmental source harboring many molecular strain types. This has very significant implications for management, indicating that M. avium and M. intracellulare infections can be managed conservatively, as other environmentally acquired infections are.

Acknowledgments

The authors thank Shirley Sykes for financial support of this study, and Yvonne Cates, Julie Concha, April Gorow, Michele Jasura, Liz Keays, J.P. Montagne, and Janet Stagnaro for technical assistance.

Literature Cited

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