A Case of Acquired Rifampin Resistance in Mycobacterium Bovis Bacillus Calmette-Guérin-induced Cystitis: Necessity for Treatment Guidelines.

A case of presumed bacillus Calmette-Guérin (BCG) cystitis in an elderly female patient following direct intravesical BCG instillation treatment for papillary transitional cell carcinoma is reported. The organism cultured from urine samples was eventually identified as a rifampin-resistant Mycobacterium bovis BCG isolate. Because the patient had received rifampin monotherapy during the course of treatment for presumed BCG disease, the clinical picture favoured acquired rifampin resistance. Sequencing of the target gene for rifampin (rpoB) confirmed a known mutation responsible for conferring high levels of resistance to both rifampin and rifabutin (Ser531Tyr). To the authors' knowledge, this is the first reported case of M bovis BCG disease in a non-HIV patient where the organism had acquired drug resistance to rifampin, and the second reported case of M bovis BCG that had acquired drug resistance. The present case demonstrates the necessity to re-evaluate appropriate guidelines for the effective treatment of BCG disease.

Viability testing of material derived from Mycobacterium tuberculosis prior to removal from a containment level-III laboratory as part of a Laboratory Risk Assessment Program.

BACKGROUND: In the field of clinical mycobacteriology, Mycobacterium tuberculosis (MTB) can be a difficult organism to manipulate due to the restrictive environment of a containment level 3 (CL3) laboratory. Tests for rapid diagnostic work involving smears and molecular methods do not require CL3 practices after the organism has been rendered non-viable. While it has been assumed that after organism deactivation these techniques can be performed outside of a CL3, no conclusive study has consistently confirmed that the organisms are noninfectious after the theoretical 'deactivation' steps. Previous studies have shown that initial steps (such as heating/chemical fixation) may not consistently kill MTB organisms. METHODS: An inclusive viability study (n = 226) was undertaken to determine at which point handling of culture extraction materials does not necessitate a CL3 environment. Four different laboratory protocols tested for viability included: standard DNA extractions for IS6110 fingerprinting, crude DNA preparations for PCR by boiling and mechanical lysis, protein extractions, and smear preparations. For each protocol, laboratory staff planted a proportion of the resulting material to Bactec 12B medium that was observed for growth for 8 weeks. RESULTS: Of the 208 isolates initially tested, 21 samples grew within the 8-week period. Sixteen (7.7%) of these yielded positive results for MTB that included samples of: deactivated culture resuspensions exposed to 80 degrees C for 20 minutes, smear preparations and protein extractions. Test procedures were consequently modified and tested again (n = 18), resulting in 0% viability. CONCLUSIONS: This study demonstrates that it cannot be assumed that conventional practices (i.e. smear preparation) or extraction techniques render the organism non-viable. All methodologies, new and existing, should be examined by individual laboratories to validate the safe removal of material derived from MTB to the outside of a CL3 laboratory. This process is vital to establish in house biosafety-validated practices with the aim of protecting laboratory workers conducting these procedures.

A high proportion of novel mycobacteria species identified by 16S rDNA analysis among slowly growing AccuProbe-negative strains in a clinical setting.

Sequencing of the 16S ribosomal DNA (rDNA) for identification of nontuberculous mycobacteria (NTM) has contributed to the establishment of more than 35 new species during the last decade. Increasingly, NTM are accepted as potential or proven pathogens. We identified, by 16S rDNA sequence analysis, slowly growing NTM isolates negative by AccuProbe (GenProbe, San Diego, CA) that previously were identified by using conventional biochemical techniques, to determine the accuracy of reporting AccuProbe-negative NTM prior to sequence-based identification. Of 82 strains, 30 were deemed novel. An attempt was made to determine the clinical importance of previously misidentified novel species. Clinical cases are described for a number of strains previously identified as Mycobacterium terrae complex, Mycobacterium scrofulaceum, and Mycobacterium avium complex. As sequence-based identification methods become more commonplace in clinical microbiology laboratories, there is a need to understand the significance of previously undescribed species, which often mimic and subsequently are identified as well-established species.

Mycobacterium neoaurum bacteremia in a hemodialysis patient.

Bacteremia due to Mycobacterium neoaurum, a rapidly growing mycobacterium, is described in a diabetic woman on hemodialysis. This is the first reported case of M neoaurum bacteremia in Canada. The organism initially grew on standard BacT/Alert SA aerobic blood cultures, and was subsequently positively identified using 16S rRNA sequence analysis. The present case serves to reinforce the need for a high index of clinical suspicion of infections caused by unusual microorganisms in the context of an immunocompromised host.

A mutation in Mycobacterium tuberculosis rpoB gene confers rifampin resistance in three HIV-TB cases.

Rifampin is a key component of standard short-course first-line therapy against Mycobacterium tuberculosis (MTB). Rifampin monoresistant MTB, previously a rare phenomenon, is now being reported at increasing rates worldwide. We report a mutation in the rpoB region leading to low level rifampin monoresistance in a cluster of HIV-positive patients. All rifampin monoresistant isolates identified from 2004 to 2006 underwent susceptibility confirmation, sequencing of rpoB and genotyping. Three patients were found to have a previously undocumented 3 base pair insertion at codon 525 in the rpoB region. The earliest initial case was infected with fully susceptible MTB. Disease relapse occurred 7 months later with a genotypically identical MTB isolate, showing acquired rifampin monoresistance. MTB isolates from 2 subsequent patients showed primary rifampin monoresistance with an identical genotype to the index case. Patients with rifampin monoresistant MTB tend to have poorer outcomes than those with fully susceptible strains. Risk factors for the development of rifampin monoresistance include co-morbid HIV infection and previously treated tuberculosis. HIV infection has been associated with malabsorption of anti-tuberculous medications leading to sub-therapeutic levels of administered drugs. These factors may have played a role in the development of this previously undocumented mutation.

Soft tissue infection caused by a novel pigmented, rapidly growing mycobacterium species.

Molecular techniques are playing an important role in the diagnosis of nontuberculous mycobacterial infections. This case report describes a chronic soft tissue infection in an immunocompetent patient caused by a previously undescribed pigmented, rapidly growing Mycobacterium species, emphasizing the importance of clinical suspicion and effective laboratory techniques in the diagnosis and treatment of infection.