Molecular characterization and drug susceptibility profile of a Mycobacterium avium subspecies avium isolate from a dog with disseminated infection

Abstract

Mycobacterium avium-intracellulare complex (MAC) infections have been described in many mammalian species, including humans and pets. We isolated and molecularly typed the causative agent of a rare case of disseminated mycobacteriosis in a dog. We identified the pathogen as M. avium subspecies avium by sequencing the partial genes gyrB and rpsA. Considering the zoonotic potential of this infection, and in an attempt to ensure the most effective treatment for the animal, we also determined the drug susceptibility profile of the isolate to the most common drugs used to treat MAC disease in humans. The pathogen was tested in vitro against the macrolide clarithromycin, as well as against amikacin, ciprofloxacin, rifampicin, ethambutol and linezolid, by the resazurin microdilution assay. It was found to be sensitive to all tested drugs apart from ethambutol. Despite the fact that the pathogen was sensitive to the therapies administered, the dog's overall clinical status worsened and the animal died shortly after antimicrobial susceptibility results became available. Nucleotide sequencing of the embB gene, the target gene most commonly associated with ethambutol resistance, showed new missense mutations when compared to sequences available in public databases. In conclusion, we molecularly identified the MAC pathogen and determined its drug susceptibility profile in a relatively short period of time (7 days). We also characterized new genetic mutations likely to have been involved in the observed ethambutol resistance. Our results confirmed the usefulness of both the gyrB and the rpsA genes as biomarkers for an accurate identification and differentiation of MAC pathogens.

Isolation, Molecular Typing, and Antibiotic Susceptibility Testing of Mycobacterium avium Subspecies hominissuis From a Dog With Generalized Mycobacteriosis

“…Two μl of DNA from each sample was used as a template for the multiplex PCR assay previously described ( 16 ), with some modifications (also as previously described) ( 17 ), in order to detect and identify members of the genus Mycobacterium , and to differentiate between members of the M. tuberculosis complex (MTC), M. avium and M. intracellulare . Reactions were performed in a total volume of 50 μl using MyTaq Red DNA Polymerase (Bioline, London, UK) following the manufacturer's instructions and the cycling conditions previously reported ( 17 ). Field isolates of M. bovis and M. avium from our collection were used as controls.…”

“…The four colonies, as well as the primary broth culture, positive for M. avium were typed at subspecies level. Both a 353 bp fragment of the gyrB gene encoding the DNA gyrase B and a 933 bp fragment of the rpsA gene encoding the ribosomal protein S1 were PCR amplified and sequenced as previously described ( 17 ).…”

“…Seven samples were tested: one from the primary broth culture, and six from single colony-derived subcultures (three for each different rpsA nucleotide sequence here identified) previously typed by MIRU-VNTR. The drug susceptibility testing was performed using the resazurin microtitre assay (REMA) according to the protocol previously described ( 17 ).…”

Mycobacterium avium: an Emerging Pathogen for Dog Breeds with Hereditary Immunodeficiencies

“…However, according to recent studies and unpublished data by authors, MAH appears to be the predominant pathogenic subspecies [77•, 78•, 79•, 80•]. Occasionally, mycobacteriosis due to MAP [64] or MAA [81] has been reported in dogs. To date, there are over 43 published cases reporting MAC-related mycobacteriosis in dogs (Table 1); however, there are many more unreported cases particularly in the Miniature Schnauzer and Basset hound breeds.…”

Serological survey for antibodies against Mycobacterium avium complexin hunting dogs in East-Central Portugal

“…The Mycobacterium avium complex (MAC) belongs to the family of nontuberculous mycobacteria and it is a major health problem worldwide in humans and animals (1,2).…”