Pseudo-outbreak of septicemia due to rapidly growing mycobacteria associated with extrinsic contamination of culture supplement

Between April and December 1994, 23 blood cultures from human immunodeficiency virus-infected patients grew rapidly growing mycobacteria suspected to be Mycobacterium chelonae at a hospital in New Jersey. The isolates were later identified as M. abscessus. Several bacterial species, including M. abscessus, were cultured from an opened multidose supplement vial (BBL Septi-Chek AFB Supplement) that had been used for mycobacterial blood cultures. The M. abscessus isolates from case patients and the supplement vial had identical multilocus enzyme electrophoresis and antimicrobial susceptibility patterns. Finding a contaminated vial of supplement, together with the lack of a distinct syndrome in case patients, was consistent with a pseudo-outbreak.     

Clinicopathologic implications of MDM2, p53 and K-ras gene alterations in osteosarcomas: MDM2 amplification and p53 mutations found in progressive tumors

The role of rapidly growing mycobacteria in the pathogenesis of pulmonary disease is being increasingly recognized; however, the clinical significance of these mycobacteria in patients with underlying malignancy has not been well studied. Over a 6-year period, 37 cancer patients with rapidly growing mycobacteria isolated from respiratory specimens were identified at our center. Mycobacterium chelonae group was isolated in 24 cases and Mycobacterium fortuitum in 13 cases. Of the 24 cases with cultures yielding Mycobacterium chelonae group, eight met the study criteria for infection and were determined to be clinically significant, whereas only one of the Mycobacterium fortuitum isolates was determined to represent infection. An average of two antimicrobial agents were used for treatment, most commonly clarithromycin, ciprofloxacin, and trimethoprim/sulfamethoxazole. Although the isolation of rapidly growing mycobacteria represents colonization in most cases, these bacteria, especially the Mycobacterium chelonae group, may cause pulmonary disease in cancer patients. The clinical and radiological findings are usually non-specific in this population, and patients with respiratory cultures yielding rapidly growing mycobacteria should be assessed carefully to distinguish infection from colonization. Effective therapy can be provided with oral regimens that include at least two antibiotics to which the organism is susceptible.     

Bacteriology of mycobacteria: taxonomic and morphological characteristics

Bacteriological characteristics of organisms belonging to Genus Mycobacterium which involves more than 60 species are described. Mycobacterial organisms can be divided into the following groups having differential characteristics, on the basis of the results of biological, biochemical, and genetic investigations, including lipid analysis, DNA probe test, and comparative 16S ribosomal RNA sequencing. First, Mycobacterium tuberculosis complex (M. tuberculosis, M. bovis, M. africanum, etc.). Second, cultivable but slowly growing nontuberculous mycobacteria, including photochromogens (Runyon Group I) such as M. kansasii, M. marinum, M. simiae, M. intermedium, and M. asiaticum, scotochromogens (Runyon Group II) such as M. scrofulaceum, M. szulgai, M. injectum, M. lentiflavum, and M. gordonae, nonphotochromogenens (Runyon Group III) such as M. avium, M. intracellulare, M. xenopi, M. malmoense, M. genavense, M. celatum, and M. gastri. Third, cultivable rapidly growing nontuberculous mycobacteria (Runyon Group IV) including M. fortuitum, M. chenolae, M. abscessus, M. phlei, and M. smegmatis. Fourth, noncultivable mycobacteria including M. leprae. About 30 species of Mycobacterium cause pulmonary, dermal, lymphatic, and disseminated infections in human beings. This paper mainly deals with the taxonomic, morphological, and other biological characteristics of these mycobacterial organisms.     

Identification of mycobacteria infecting fish to the species level using polymerase chain reaction and restriction enzyme analysis

An assay is described utilizing PCR technology for a rapid diagnostic test to identify fish infection with Mycobacterium marinum, M. fortuitum and M. chelonae. A 924 bp DNA fragment from a highly conserved area of the mycobacterial 16S rRNA gene was amplified using mycobacteria genus-specific primers and digested with restriction enzymes (BanI and ApaI). This examination yielded unique restriction patterns for each mycobacterial specie enabling identification of mycobacteria infecting fish to the species level. The protocol can be applied to purified DNA, a simple colony preparation or infected fish tissue. This protocol can be completed in 1-2 days.     

Rapid development of resistance to clarithromycin following monotherapy for disseminated Mycobacterium chelonae infection in a heart transplant patient

Mycobacterium chelonae (formerly known as M. chelonae subspecies chelonae) is a rapidly growing mycobacterium that can cause disseminated infections, especially in immunocompromised hosts. The bacterium is typically resistant to antimicrobial agents; less than 20% of M. chelonae isolates are susceptible to trimethoprim-sulfamethoxazole, doxycycline, erythromycin, or ciprofloxacin. Findings in a recent study suggested that clarithromycin may be the drug of choice for the treatment of cutaneous (disseminated) disease due to M. chelonae. We describe a 60-year-old heart transplant patient with disseminated M. chelonae infection for whom monotherapy with clarithromycin failed because of the rapid development of resistance to the drug.     

Mycobacterium novocastrense sp. nov., a rapidly growing photochromogenic mycobacterium

A strain isolated from a biopsy sample taken from a slowly spreading skin granulation on a child's hand was found to have properties consistent with its classification in the genus Mycobacterium. An almost complete gene sequence of the 16S rRNA of the strain was determined following the cloning and sequencing of the amplified gene. The sequence was aligned with those available for mycobacteria, and phylogenetic trees were inferred with four tree-making algorithms. The organism, which formed a distinct phyletic line within the evolutionary radiation occupied by rapidly growing mycobacteria, was readily distinguished from members of validly described species of rapidly growing mycobacteria on the basis of its mycolic acid pattern and a number of other phenotypic features, notably its ability to form yellow pigmented colonies when incubated in the light. The name proposed for this new species is Mycobacterium novocastrense. The type strain is DSM 44203.     

Adaptive antioxidant response of manganese-superoxide dismutase following repetitive UVA irradiation

To explore a simple, rapid, and inexpensive way to identify Mycobacterium tuberculosis complex culture, dot blot hybridization using IS6110 as the marker was performed against 2788 known clinical isolates of mycobacteria including M. tuberculosis (n = 721), M. kansasii (177), M. marinum (10), M. avium complex (700), M. terrae complex (203), M. fortuitum (476), M. chelonae (439), and other nonpigmented Runyon's Group IV mycobacteria (62). We found that the sensitivity and specificity of the test were 94.3% and 100%, respectively. When this method was evaluated in a laboratory blind study of 1253 initially unknown clinical isolates, its sensitivity and specificity were 91.2% and 100%, respectively. Because this identification test is technically simple, rapid, and can be done in batches, together with its high sensitivity and specificity, it is a cost-effective method for routine identification of M. tuberculosis complex in laboratories of areas with a high incidence of tuberculosis.     

Megaesophagus and pneumonia associated with Mycobacterium chelonei. A case report and a literature review

The case of a 37-year-old woman who developed a subacute, bilateral, noncavitary pneumonia 5 years after a colon interposition esophagoplasty is presented. Mycobacterium chelonei, subspecies abscessus, was assigned a pathogenic role based on the findings of (1) a clinical and roentgenographic picture consistent with tuberculosis, (2) sputum smears showing acid-fast bacilli, (3) repeated sputum cultures yielding heavy growths of Mycobacterium chelonei, subspecies abscessus, and (4) a 12-mm by 12 mm-skin test response to homologous antigen (purified protein derivative-CL) with no response to an equivalent dose of purified protein derivative-S. The patient recovered fully without significant antituberculous chemotherapy. A survey of the literature revealed 11 similar case reports featuring a documented association between megaesophagus and pulmonary infection with rapidly growing mycobacteria.     

Role of embB in natural and acquired resistance to ethambutol in mycobacteria

The mycobacterial embCAB operon encodes arabinosyl transferases, putative targets of the antimycobacterial agent ethambutol (EMB). Mutations in embB lead to resistance to EMB in Mycobacterium tuberculosis. The basis for natural, intrinsic resistance to EMB in nontuberculous mycobacteria (NTM) is not known; neither is the practical implication of resistance to EMB in the absence of embB mutations in M. tuberculosis well understood. The conserved embB resistance-determining region (ERDR) of a collection of 13 strains of NTM and 12 EMB-resistant strains of M. tuberculosis was investigated. Genotypes were correlated with drug susceptibility phenotypes. High-level natural resistance to EMB (MIC, . or =64 microg/ml) was associated with a variant amino acid motif in the ERDR of M. abscessus, M. chelonae, and M. leprae. Transfer of the M. abscessus emb allele to M. smegmatis resulted in a 500-fold increase in the MICs. In M. tuberculosis, embB mutations were associated with MICs of > or =20 microg/ml while resistance not associated with an ERDR mutation generally resulted in MICs of < or =10 microg/ml. These data further support the notion that the emb region determines intrinsic and acquired resistance to EMB and might help in the reassessment of the current recommendations for the screening and treatment of infections with EMB-resistant M. tuberculosis and NTM.     

A single 16S ribosomal RNA substitution is responsible for resistance to amikacin and other 2-deoxystreptamine aminoglycosides in Mycobacterium abscessus and Mycobacterium chelonae

Twenty-six clinical isolates of Mycobacterium abscessus resistant to amikacin were identified. Most isolates were from patients with posttympanostomy tube placement otitis media or patients with cystic fibrosis who had received aminoglycoside therapy. Isolates were highly resistant (MICs > 1024 microg/mL) to amikacin, kanamycin, gentamicin, tobramycin, and neomycin (all 2-deoxystreptamine aminoglycosides) but not to streptomycin. Sequencing of their 16S ribosomal (r) RNA revealed that 16 (94%) of 17 had an A-->G mutation at position 1408. In vitro-selected amikacin-resistant mutants of M. abscessus and Mycobacterium chelonae had the same resistance phenotype, and 15 mutants all had the same A-->G substitution at position 1408. Introducing an rRNA operon from Mycobacterium smegmatis with a mutated A-->G at this position into a single functional allelic rRNA mutant of M. smegmatis produced the same aminoglycoside resistance phenotype. These studies demonstrate this 16S rRNA mutation is responsible for amikacin resistance in M. abscessus, which has only one copy of the rRNA operon.     

Effect of decreased glucose concentration on cerebrovascular tone in vitro

A single Polymerase Chain Reaction (PCR) within the rpoV gene was developed to rapidly distinguish mycobacteria isolated from clinical specimens. The species identifications of Mycobacterium avium complex (MAC) and Mycobacterium tuberculosis were congruent with standard typing techniques. The analysis was targeted toward the identification of species-specific markers for the clinically relevant M. tuberculosis and M. avium. In addition, HaeIII digestion of the amplification products yielded isolates-specific bands.     

Epidemiology of disease caused by nontuberculous mycobacteria

The clinically important nontuberculous mycobacteria include M. kansasii, M. genavense, M. marinum, M. simiae, M. scrofulaceum and M. szulgai, M. avium, M. haemophilum, M. intracellulare, M. malmoense, M. ulcerans, and M. xenopi, M. abscessus, M. chelonae, M. fortuitum, and (rarely) M. smegmatis. Four clinical syndromes account for nearly all cases: pulmonary disease, lymphadenitis, skin or soft tissue disease, and disseminated disease in AIDS. M. avium and M. intracellulare (known together as M. avium complex), are the most common causes of pulmonary disease, lymphadenitis, and disseminated disease. All four clinical syndromes seem to be increasing in frequency, particularly in immunosuppressed hosts. Nontuberculous mycobacteria are acquired from the environment, but specific reservoirs of these organisms leading to human disease have not been defined.     

Evaluation of mycobacteria growth indicator tube for recovery and drug susceptibility testing of Mycobacterium tuberculosis isolates from respiratory specimens

The new BBL mycobacteria growth indicator tube (MGIT) was evaluated for its ability to detect mycobacteria directly from patient specimens and to determine the drug susceptibility of Mycobacterium tuberculosis isolates. A total of 85 respiratory specimens were tested. Specimens were digested, concentrated, examined microscopically for acid-fast bacilli, and inoculated into MGITs and onto Lowenstein-Jensen slants by standard procedures. The tubes were incubated at 37 degrees C and were examined daily for fluorescence to 365-nm UV light. All 25 specimens smear positive for acid-fast bacilli were tested for drug susceptibility in MGITs containing 1.0 mu g of rifampin per ml, 0.1 mu g of isoniazid per ml, 2.0 mu g of streptomycin per ml, and 2.0 mu g of ofloxacin per ml. These results were compared with those obtained by testing the same M. tuberculosis isolates by the indirect proportion method at drug concentrations of 4.0 mu g of rifampin per ml, 0.2 mu g of isoniazid per ml, 2.0 mu g of ethambutol per ml. 4.0 mu g of streptomycin per ml, and 2.0 mu g of ofloxacin per ml. No significant difference in the sensitivity of detection of M. tuberculosis isolates was found between the two methods. However, the time to detection was significantly shorter in MGITs. Drug susceptibility test results for M. tuberculosis isolates by the two methods demonstrated an excellent correlation. The mean time to reporting of drug susceptibility results was 5 days for MGITs versus 16 days for Lowenstein-Jensen slants. The results of this preliminary study indicate that the MGIT system appears to have potential for routine use in mycobacteriology for both the detection and the drug susceptibility testing of M. tuberculosis isolates. However, it is important to emphasize that simple nonautomated equipment should be developed to improve the accuracy of fluorescence detection.     

Purification and in vitro reconstitution of the essential protein components of an aromatic polyketide synthase

The in vitro activities of seven quinolones and the sequences of the quinolone resistance-determining regions (QRDR) in the A and B subunits of DNA gyrase were determined for 14 mycobacterial species. On the basis of quinolone activity, quinolones were arranged from that with the greatest to that with the least activity as follows: sparfloxacin, levofloxacin, ciprofloxacin, ofloxacin, pefloxacin, flumequine, and nalidixic acid. Based on MICs, the species could be organized into three groups: resistant (Mycobacterium avium, M. intracellulare, M. marinum, M. chelonae, M. abscessus [ofloxacin MICs, >/=8 microg/ml]), moderately susceptible (M. tuberculosis, M. bovis BCG, M. kansasii, M. leprae, M. fortuitum third biovariant, M. smegmatis [ofloxacin MICs, 0.5 to 1 microg/ml]), and susceptible (M. fortuitum, M. peregrinum, M. aurum [ofloxacin MICs, 相似文献   

Mechanisms involved in the intrinsic isoniazid resistance of Mycobacterium avium

Isoniazid (INH), which acts by inhibiting mycolic acid biosynthesis, is very potent against the tuberculous mycobacteria. It is about 100-fold less effective against Mycobacterium avium. This difference has often been attributed to a decreased permeability of the cell wall. We measured the rate of conversion of radiolabelled INH to 4-pyridylmethanol by whole cells and cell-free extracts and estimated the permeability barrier imposed by the cell wall to INH influx in Mycobacterium tuberculosis and M. avium. There was no significant difference in the relative permeability to INH between these two species. However, the total conversion rate in M. tuberculosis was found to be four times greater. Examination of in vitro-generated mutants revealed that the major resistance mechanism for both species is loss of the catalase-peroxidase KatG. Analysis of lipid and protein biosynthetic profiles demonstrated that the molecular target of activated INH was identical for both species. M. avium, however, formed colonies at INH concentrations inhibitory for mycolic acid biosynthesis. These mycolate-deficient M. avium exhibited altered colony morphologies, modified cell wall ultrastructure and were 10-fold more sensitive to treatment with hydrophobic antibiotics, such as rifampin. These findings may significantly impact the design of new therapeutic regimens for the treatment of infections with atypical mycobacteria.     

A novel pathogenic taxon of the Mycobacterium tuberculosis complex, Canetti: characterization of an exceptional isolate from Africa

In an attempt to characterize an unusual mycobacterial strain isolated from a 2-year-old Somali patient with lymphadenitis, we applied various molecular methods not previously used for the taxonomic classification of mycobacteria. This isolate, designated So93, did not differ from Mycobacterium tuberculosis in the biochemical tests and in its 16S rRNA sequence, but produced smooth and glossy colonies, which is highly exceptional for this species. This smooth phenotype was unstable and switched nonreversibly to a rough colony morphology with a low frequency. The two colony types were equally virulent for the guinea pig, exhibiting characteristic tuberculous disease. Both morphotypes had shorter generation times than the M. tuberculosis reference laboratory strain H37Rv and clinical isolates of M. tuberculosis and Mycobacterium bovis. Furthermore, the So93 isolate differed from all M. tuberculosis complex strains described thus far by having only a single copy of insertion sequence IS1081, an unusual composition of the direct repeat cluster, and a characteristic phenolic glycolipid and lipooligosaccharide. This glycolipid had previously been observed only in a smooth isolate of M. tuberculosis obtained in 1969 by Canetti in France. Analysis of the Canetti strain showed that it shared virtually all genetic properties characteristic of So93, distinguishing these two strains from the known M. tuberculosis complex taxa, M. tuberculosis, Mycobacterium africanum, M. bovis, and Mycobacterium microti. The natural reservoir, host range, and mode of transmission of the group of bacteria described in this paper are presently unknown. This study, partly based on not previously used molecular criteria, supports the idea that the established members within the M. tuberculosis complex and the newly described Canetti grouping should be regarded as a single species, which likely will be designated "M. tuberculosis".     

Failure to recognize rapidly growing mycobacteria in a proficiency testing sample without specific request--a wider diagnostic problem?

Fifty participants in the Swiss External Quality Control Program in Bacteriology/Mycology received a diagnostic sample containing Mycobacterium fortuitum. Only 31 used some sort of acid-fast stains, and 13 reported the diagnosis of M. fortuitum or rapidly growing mycobacteria. We conclude that the presence of 'rapid growers' in routine bacteriology samples is underestimated, and that acid-fast stains should be performed on suspicious Gram-positive rods.     

Mycobacterium fortuitum infection of the sternum. Review of the literature and case illustration

Sternal wound infection with atypical mycobacteria following open heart surgery is a rare occurrence. Previous reports have described infection by Mycobacterium fortuitum, an acid-fast bacillus and member of a larger family of rapidly growing mycobacteria. The source and mode of transmission have not been identified. Surgical debridement and the combination of aminoglycosides and quinolones have been shown to be effective methods of treatment. More recently, clarithromycin has been shown to be the drug of choice against rapidly growing mycobacteria. We describe a 49-year-old woman who underwent infundibular stenosis repair and in whom M fortuitum sternal osteomyelitis developed. Total sternectomy, muscle flap reconstruction, and antibiotic treatment successfully eradicated the infection.     

Evaluation of single-use masks and respirators for protection of health care workers against mycobacterial aerosols

BACKGROUND: The recent increase in multidrug-resistant tuberculosis has spawned a major controversy concerning the degree of respiratory protection needed by health care workers, particularly during sputum-inducing procedures. The objective of this study was to measure the filtration efficiencies of a single-use submicron surgical mask, two disposable dust/mist respirators, a dust/mist/fume respirator, and a high-efficiency particulate air respirator against aerosolized mycobacteria. Facial fit was not addressed. METHODS: In a specially designed enclosed test apparatus, an aerosol was generated with a Collison nebulizer from a known concentration of Mycobacteria chelonae, used as a surrogate for Mycobacterium tuberculosis. Aerosol concentrations were measured with Anderson samplers upstream and downstream of the test masks and respirators, which were heat sealed to a metal plate. RESULTS: Mean efficiencies ranged from approximately 97% for the surgical mask and a dust/mist respirator to more than 99.99% for the high-efficiency particulate air respirator. Measurements of filter efficiency with an Aerodynamic Particle Sizer for the M. chelonae aerosol and independent challenge tests with latex spheres correlated closely with measurements of M. Chelonae collection efficiency determined with Andersen samplers. CONCLUSIONS: Analysis of variance and Tukey's method for multiple comparisons indicated that the dust/mist/fume respirator and the HEPA respirator collected M. chelonae with significantly greater efficiency than did either the surgical mask or the dust/mist respirator. Even the least efficient mask tested, however, had a filter efficiency of more than 97% against particles averaging less than 1 micron in aerodynamic diameter.     

Identification and strain differentiation of Mycobacterium species on the basis of DNA 16S-23S spacer region polymorphism

Amplification of the region separating the genes coding for the two rRNA species 16S and 23S was performed to identify 56 mycobacterial strains, belonging to eleven species: Mycobacterium tuberculosis, M. avium, M. kansasii, M. gordonae, M. abscessus, M. fortuitum, M. xenopi, M. bovis, M. bovis/BCG, M. africanum and M. intracellulare. Reproducible amplification patterns were obtained with most species with the exception of M. kansasii which showed heterogeneity, confirming the existence of a genetically distinct subspecies within this species. In addition, we used the amplified products as target DNA for restriction endonuclease digestion and RAPD (randomly amplified polymorphic DNA) analysis to compare strains of M. abscessus, M. tuberculosis and M. avium. The discriminatory power of these two typing methods was higher than when whole genomic DNA is used as target. Our results demonstrate that the two-step approach to identification and typing on the basis of the hypervariability of 16S-23S spacer region is reliable, rapid and simple, and consequently could be an epidemiological tool in clinical laboratories.