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Mycobacterium gordonae, a low pathogenicity organism, is rarely implicated in skin and soft tissue infections. We present a 77-year-old returned diabetic traveler from rural Sudan with cutaneous M. gordonae infection. Several months of ciprofloxacin, rifampin and ethambutol led to resolution of his plaque, without signs of recurrence at 6-month follow-up.

Background: Non-tuberculous mycobacteria (NTM) are emerging as important pathogens. Their treatment also differs from that of Mycobacterium tuberculosis. In India, any datum on them is scarce as species identification and drug susceptibility are not performed in most laboratories. Susceptibility also differs from one geographic area to another, and in our country, there are no data even to guide the clinicians to start treatment empirically.Methodology: The present study endeavours to generate drug susceptibility data on NTM isolated from sputum samples collected and stored from 6445 symptomatics for pulmonary tuberculosis during a prevalence survey and from specimens received from the hospital. Isolates were not necessarily associated with the disease. Species were identified and antibiotic susceptibility was performed using micro-broth dilution technique as per the standard Clinical and Laboratory Standards Institute guidelines. Results: A total of 65 NTM with 11 species were identified, of which 27 belonged to Mycobacterium fortuitum complex, 14 Mycobacterium gordonae, 9 Mycobacterium avium, 7 Mycobacterium flavescens, 4 Mycobacterium scrofulaceum and one each of others. Sensitivity to amikacin for M. fortuitum was 95.22% (20 out of 21), followed by ciprofloxacin (76.19%) and clarithromycin (71.42%). All the 9 M. avium isolates, 11 of M. gordonae (78.57%), 5 of M. flavescens and 2 of M. scrofulaceum were sensitive to clarithromycin. All NTM were resistant to first-line antitubercular drugs except 8, which were sensitive to streptomycin. Conclusions: Drug sensitivity of NTM varies from species to species. While amikacin was the best for rapidly growing mycobacteria, clarithromycin was the most active drug against M. avium and other slow growers.

Objectives A simple, rapid, and new diagnostic test for mycobacteria, named Q Gene Mycobacteria, has been developed. It is based on multiplex PCR using primers harbouring DNA tags combined with a dipstick nucleic acid chromatography method, which does not require the denaturation of PCR products for hybridization and can identify five species of mycobacteria including Mycobacterium tuberculosis complex (MTC), Mycobacterium avium , Mycobacterium intracellulare , Mycobacterium kansasii , and Mycobacterium gordonae . This study aimed to evaluate Q Gene Mycobacteria for the accurate identification of these five species. Methods A total of 340 mycobacterial strains/isolates were tested, of which 159 were type strains (four MTC and 155 non-tuberculosis mycobacteria (NTM) including four subspecies) and 181 were clinical isolates (18 M. tuberculosis , two Mycobacterium bovis Bacillus Calmette et Guerin (BCG), and 161 NTM comprising 16 species) collected from eight laboratories and hospitals in Japan. Species identification of NTM isolates was performed using the DNA-DNA hybridization method and/or direct sequencing of 16S rRNA, hsp65 , and rpoB genes. Q Gene Mycobacteria was compared with above conventional methods for identifying the five species. Results Q Gene Mycobacteria showed excellent concordance for species identification, specifically 99.4% (158/159) for type strains and 99.4% (180/181) for clinical isolates. The two strains that were misidentified as M. gordonae were Mycobacterium paragordonae . As they are genetically close and there is few case reports of M. paragordonae , it might not be a serious critical issue to distinguish M. paragordonae from M. gordonae . Conclusions Q Gene Mycobacteria was able to identify frequently isolated mycobacterial species accurately and easily. Therefore, Q Gene Mycobacteria could be a useful tool for the identification of specific mycobacteria in clinical laboratories.

Based on microbiological and histopathological examinations and DNA sequencing, several outbreaks of mycobacteriosis in the reared sturgeons, including Chinese sturgeon (Acipenser sinensis Gray) and Amur sturgeon (Acipenser schrencki), were identified during 2009 to 2010. Forty-nine isolates of non-tuberculous mycobacteria(NTM)were isolated from 19 diseased sturgeons. In total, seven species of Mycobacterium were identified, namely, Mycobacterium chelonae, Mycobacterium marinum, Mycobacterium gordonae, Mycobacterium fortuitum, Mycobacterium szulgai, Mycobacterium arupense and Mycobacterium porcinum. Among them, M. marinum was found to be more prevalent (89.5%) compared with the other mycobacterial species. When two molecular biological methods, PCR-DGGE (denaturing gradient gel electrophoresis) analysis and rpoB gene library sequencing, were used to analyse the mycobacterial DNAs extracted from the diseased fish tissues, mixed infections of two or three mycobacterial species were found being the predominant infection form (94.7%) in sturgeon mycobacteriosis. M. marinum was the only one species that caused sturgeon mycobacteriosis alone. Virulence assay showed that M. marinum possessed stronger pathogenicity to zebrafish killing 100% of fish in 28days at 10(3) cfu/fish than the other species. These results suggested that M. marinum is the major pathogenic bacteria in sturgeon mycobacteriosis. To the best of our knowledge, this study is the first report on mycobacteriosis in farmed Chinese and Amur sturgeons as well as the first isolation of M. porcinum and M. arupense from fish.

PCR targeting the 16S–23S rRNA gene internally transcribed spacer (ITS) region has been proposed as a rapid and reliable method for the detection of Mycobacterium species in human clinical specimens. Because of variation in ITS sequences amongst Mycobacterium species, a single PCR amplification can be used to differentiate slowly growing and rapidly growing species within this genus. In the present study, analysis by ITS-PCR and ITS-restriction fragment length polymorphism (RFLP) was found to be a useful, simple and rapid method compared to current molecular and phenotypic techniques. The ITS was amplified from 13 reference strains and 59 fish isolated mycobacteria using a set of published PCR primers. After PCR, the banding patterns generated allowed slowly growing mycobacteria to be differentiated from all other rapidly growing species, with the exception of Mycobacterium conceptionense . Hae III was selected as one of two restriction enzymes that, together with the knowledge about amplicon sizes, would produce an acceptable level of discrimination in the resulting RLFP patterns, especially in the rapidly growing group of mycobacteria. After digestion with Sau 96I, the amplified products of most isolates of Mycobacterium fortuitum , including subtypes II and V and those 2 isolates with new patterns (220, 100 bp), presented identical or very similar patterns as obtained by Hae III digestion. All isolates of Mycobacterium marinum , Mycobacterium chelonae and Mycobacterium gordonae , whose PCR products were not digested with Hae III, produced two well-defined fragments with the Sau 96I restriction enzyme.

Methods: Case description of a 34 year old woman with chronic granulomatous anterior uveitis following implantation of a phakic anterior chamber intraocular lens (IOL). Aqueous humor was processed for bacterial, fungal culture, and PCR analysis.Results: PCR for 16S ribosomal RNA universal primers was positive and Southern-blot for Propionibacterium spp was negative. Panfungal PCR was negative. Growth of a pigmented yellow colony was detected on Lowenstein-Jensen medium. The isolate was subsequently identified as Mycobacterium gordonae. The patient was treated for atypical mycobacteria as well as an intracameral injection of moxifloxacin, with resolution of inflammation and improvement in visual acuity.Conclusions: This is the first reported case of Mycobacterium gordonae chronic postoperative endophthalmitis, and of chronic endophthalmitis following phakic IOL surgery. PCR and cultures were invaluable in making the diagnosis.

The main cause of pulmonary tuberculosis (TB) is infection with Mycobacterium tuberculosis (MTB). We aimed to evaluate the contribution of nontuberculous mycobacteria (NTM) to pulmonary disease in patients from the state of Rondônia using respiratory samples and epidemiological data from TB cases. Mycobacterium isolates were identified using a combination of conventional tests, polymerase chain reaction-based restriction enzyme analysis of hsp65 gene and hsp65 gene sequencing. Among the 1,812 cases suspected of having pulmonary TB, 444 yielded bacterial cultures, including 369 cases positive for MTB and 75 cases positive for NTM. Within the latter group, 14 species were identified as Mycobacterium abscessus, Mycobacterium avium, Mycobacterium fortuitum, Mycobacterium intracellulare, Mycobacterium gilvum, Mycobacterium gordonae, Mycobacterium asiaticum, Mycobacterium tusciae, Mycobacterium porcinum, Mycobacterium novocastrense, Mycobacterium simiae, Mycobacterium szulgai, Mycobacterium phlei and Mycobacterium holsaticum and 13 isolates could not be identified at the species level. The majority of NTM cases were observed in Porto Velho and the relative frequency of NTM compared with MTB was highest in Ji-Paraná. In approximately half of the TB subjects with NTM, a second sample containing NTM was obtained, confirming this as the disease-causing agent. The most frequently observed NTM species were M. abscessus and M. avium and because the former species is resistant to many antibiotics and displays unsatisfactory cure rates, the implementation of rapid identification of mycobacterium species is of considerable importance.

The incidence of lung and other diseases due to nontuberculous mycobacteria (NTM) is increasing. NTM sources include potable water, especially in households where NTM populate pipes, taps, and showerheads. NTM share habitats with free-living amoebae (FLA) and can grow in FLA as parasites or as endosymbionts. FLA containing NTM may form cysts that protect mycobacteria from disinfectants and antibiotics. We first assessed the presence of FLA and NTM in water and biofilm samples collected from a hospital, confirming the high prevalence of NTM and FLA in potable water systems, particularly in biofilms. Acanthamoeba spp. (genotype T4) were mainly recovered (8/17), followed by Hartmannella vermiformis (7/17) as well as one isolate closely related to the genus Flamella and one isolate only distantly related to previously described species. Concerning mycobacteria, Mycobacterium gordonae was the most frequently found isolate (9/17), followed by Mycobacterium peregrinum (4/17), Mycobacterium chelonae (2/17), Mycobacterium mucogenicum (1/17), and Mycobacterium avium (1/17). The propensity of Mycobacterium avium hospital isolate H87 and M. avium collection strain 104 to survive and replicate within various FLA was also evaluated, demonstrating survival of both strains in all amoebal species tested but high replication rates only in Acanthamoeba lenticulata . As A. lenticulata was frequently recovered from environmental samples, including drinking water samples, these results could have important consequences for the ecology of M. avium in drinking water networks and the epidemiology of disease due to this species.

Significance and Impact of the Study: Air sampling, combined with laboratory testing, has been pro- posed as a method to detect the presence of TB-infected badgers within their sett without the need to trap them. This work demonstrates that using a combination of a commercially available air sampling device and subsequent culture or real-time PCR, it was not possible to detect M. bovis in infected bad- gers or their environment. Abstract Environmental air sampling was evaluated as a method to detect the presence of M. bovis in the vicinity of infected badgers and their setts. Airborne particles were collected on gelatine filters using a commercially available air sampling instrument and tested for the presence of M. bovis using bacteriological culture and real-time PCR. The sensitivity of bacteriological culture was broadly similar to that of real-time PCR when testing samples artificially spiked with M. bovis. Sampling was undertaken from directly under the muzzles of badgers which had been experimentally infected with M. bovis (37 samples), within enclosures housing the experimentally infected animals (50 samples), and in the vicinity of setts with resident infected wild badgers (52 samples). The methods employed did not detect M. bovis from either infected badgers or artificial or natural setts known to contain infected animals. However, samples taken at four of the six natural setts were positive for Mycobacterium gordonae.

Interleukin-17 (IL-17) is produced by a subset of CD4(+) T helper (Th) lymphocytes known as Th17 cells. In humans, IL-1β, enhanced by IL-6 and IL-23 is crucial for differentiation of these cells. IL-17 evokes inflammation and is involved in host defence against microorganisms, although little is known about its role in diseases caused by non-tuberculous mycobacteria. The genus Mycobacterium contains both obligate and opportunistic pathogens as well as saprophytes, and the mycobacterial cell envelope is unique in its abundance of lipids. Here we investigated IL-17 and IL-23 production in human PBMC in response to intact UV-inactivated mycobacteria and mycobacterial surface lipids from two opportunistic (Mycobacterium avium and Mycobacterium abscessus) and one generally non-pathogenic (Mycobacterium gordonae) species. Representative Gram-positive (Enterococcus faecalis, Streptococcus mitis) and Gram-negative (Escherichia coli) bacteria were included as controls. Intact mycobacteria induced production of large amounts of IL-17, while IL-17 responses to control bacteria were negligible. Purified CD4(+) T cells, but not CD4-depleted cell fractions, produced this IL-17. Isolated mycobacterial surface lipids induced IL-17, but not IL-23 production. The ability of the non-tuberculous mycobacteria to induce IL-17 production in CD4(+) T cells was the same regardless of the pathogenic potential of the particular mycobacterial species.

症例は88歳女性。既往歴として平成9年に肺MAC 症の治療歴がある。平成21年4月咳嗽と血痰で当科を受診。胸部Ｘ線・CT で両肺に散布影と左舌区に多発する空洞病変を認めた。各種喀痰抗酸菌検査によりMycobacterium（以下M.）gordonaeが同定され、肺M. gordonae感染症の診断で抗菌薬3剤による治療を行ない改善した。M. gordonaeは希少な抗酸菌であり、抗酸菌の中でもヒトへの起病性が極めて低い菌として知られている。我々が検索し得た限りでは、肺MAC 症の治療後に発症した肺M. gordonae感染症の報告例はなかったことから、本例は非常に貴重な症例であると考える。

Here, we report the first draft genome sequence of the clinically relevant species Mycobacterium gordonae . The clinical isolate Mycobacterium gordonae 14-8773 was obtained from the sputum of a patient with mycobacteriosis.

Fish are commonly infected with non-tuberculous mycobacteria (NTM), which should be regarded as potential pathogens when handling aquarium fish and equipment. This study examined 107 aquarium fish from pet shops. Cultivation of the fish samples using different selective media was conducted for identification of NTM. Isolates were identified using the GenoType Mycobacterium common mycobacteria and additional species assays, sequencing of the 16S rRNA and rpoB genes, and real-time PCR assay for identification of Mycobacterium (M.) marinum. Among the investigated fish, 79.4% (85/107) were positive for mycobacteria, with 8.2% (7 of 85) having two mycobacterial species present. Among the positive fish, the common pathogens M. marinum, Mycobacterium fortuitum (M. fortuitum group) and Mycobacterium chelonae were identified in approx. 90% of fish and other NTM species in 10%, including Mycobacterium peregrinum/septicum, Mycobacterium gordonae, Mycobacterium arupense, Mycobacterium kansasii, Mycobacterium ulcerans and Mycobacterium setense. The well-known human pathogen M. marinum was present in 10.6% of the positive fish (9 of 85). The species of mycobacteria identified in the study are not only recognized as aquarium fish pathogens, but can also cause pathology in humans. Microbiological and clinical communities should therefore be sensitized to the role of NTM in infections associated with exposure to aquarium fish.

The genus Mycobacterium contains more than 150 species. Non-tuberculosis mycobacteria (NTM) often cause extrapulmonary and pulmonary disease. Mycobacteria detection at species level is necessary and provides useful information on epidemiology and facilitates successful treatment of patients. This retrospective study aimed to determine the incidence of the NTM isolates and Mycobacterium tuberculosis (Mtb) in clinical specimens collected from Iranian patients during February 2011–December 2013, by PCR–restriction fragment length polymorphism analysis (PRA) of the hsp65 gene. We applied conventional biochemical test and hsp65–PRA identification assay to identify species of mycobacteria in specimens from patients suspected of having mycobacterial isolates. This method was a sensitive, specific and effective assay for detecting mycobacterial species and had a 100% sensitivity and specificity for Mtb and Mycobacterium avium complex (MAC) species. Using PRA for 380 mycobacterial selected isolates, including 317 Mtb, four Mycobacterium bovis and of the 59 clinical isolates, the most commonly identified organism was Mycobacterium kansasii (35.6%), followed by Mycobacterium simiae (16.9%), Mycobacterium gordonae (16.9%), Mycobacterium fortuitum (5.1%), Mycobacterium intracellulare (5.1%), Mycobacterium avium (5.1%), Mycobacterium scrofulaceum (3.4%), Mycobacterium gastri (3.4%), Mycobacterium flavescens (3.4%), Mycobacterium chelonae (3.4%) and Mycobacterium nonchromogenicum (1.7%). PRA method, in comparison with classical methods, is rapid, useful and sensitive for the phylogenetic analysis and species detection of mycobacterial strains. Mycobacterium kansasii is the most common cause of infection by NTM in patients with non-HIV and HIV which demonstrated a high outbreak and diversity of NTM strains in our laboratory. Keywords: Non-tuberculosis mycobacteria, Heat shock protein 65, PCR–RFLP

Non-tuberculous mycobacteria (NTM) are increasingly important opportunistic pathogens responsible for a variety of clinical diseases. The aim of this study was to evaluate a novel technique, real-time PCR coupled with high-resolution melting analysis (real-time PCR-HRMA), for NTM identification. Two pairs of unique primers targeted to the 16S rRNA gene and the 16S-23S internal transcribed spacer region were selected for further evaluation. A total of 149 mycobacterial clinical isolates were subjected to analysis using the real-time PCR-HRMA system. Overall, 134 NTM identified by the 16S rRNA full-gene sequencing method were categorized into four major groups: Mycobacterium avium complex, Mycobacterium chelonae group, Mycobacterium gordonae and Mycobacterium fortuitum group. Of the 134 prevalent mycobacterial isolates, 101 mycobacteria (75.4 %) could be identified correctly by the real-time PCR-HRMA system. The individual sensitivities for the M. avium complex, M. chelonae group, M. gordonae and M. fortuitum groups were 90.9, 89.1, 100 and 36.8 %, respectively. The specificity of identifying these groups varied from 96.4 to 100 %. When identification failed, mostly it was attributable to various species in the M. fortuitum group. The real-time PCR-HRMA system is therefore a rapid and sensitive method for identifying prevalent NTM in a clinical laboratory.

Background The incidence of infection with non-tuberculous mycobacteria (NTM) after lung transplant is insufficiently defined. Data on the impact of NTM infection on lung transplant survival are conflicting. Methods To quantify the incidence and outcomes of colonization and disease with NTM in patients after lung transplantation, the medical records, chest imaging, and microbiology data of 237 consecutive lung transplant recipients between 1990 and 2005 were reviewed. American Thoracic Society (ATS)/Infectious Diseases Society of America and Centers for Disease Control criteria were used to define pulmonary NTM disease and NTM surgical-site infections (SSI), respectively. Incidence rates for NTM colonization and disease were calculated. Comparisons of median survival were done using the log-rank test. Results NTM were isolated from 53 of 237 patients (22.4%) after lung transplantation over a median of 25.2 months of follow-up. The incidence rate of NTM isolation was 9.0/100 person-years (95% confidence interval [CI), 6.8-11.8), and the incidence rate of NTM disease was 1.1/100 person-years (95% CI 0.49-2.2). The most common NTM isolated was Mycobacterium avium complex (69.8%), followed by Mycobacterium abscessus (9.4%), and Mycobacterium gordonae (7.5%). Among these 53 patients, only 2 patients met ATS criteria for pulmonary disease and received treatment for M. avium. One patient had recurrent colonization after treatment, the other one was cured. Four of the 53 patients developed SSI, 3 caused by M. abscessus and 1 caused by Mycobacterium chelonae. Three of these patients had persistent infection requiring chronic suppressive therapy and one died from progressive disseminated disease. A total of 47 (89%) patients who met microbiologic but not radiographic criteria for pulmonary infection were not treated and were found to have only transient colonization. Median survival after transplantation was not different between patients with transient colonization who did not receive treatment and those who never had NTM isolated. Conclusion Episodic isolation of NTM from lung transplant recipients is common. Most isolates occur among asymptomatic patients and are transient. Rapidly growing NTM can cause significant SSI, which may be difficult to cure. NTM disease rate is higher among lung transplant recipients than in the general population. In this cohort, NTM isolation was not associated with increased post-transplantation mortality.

Buruli ulcer disease, caused by infection with Mycobacterium ulcerans, is prevalent in central and western Sub-Saharan Africa and in southeastern Australia, and it occurs sporadically in other tropical and subtropical locales [1–3]. This report addresses a case of possible Buruli ulcer disease in an adolescent girl in Honduras. The subject was a 16-year-old female living in a rural coastal village in Honduras (near La Ceiba). She developed 2 ulcerative lesions on her leg that grew from small pinpoint lesions, to abscesses, to large undermined ulcers over a period of several weeks (see Figure ​Figure1).1). The lesions were painless. Local topical remedies were ineffective. She had no systemic symptoms, and she did not develop other lesions. There was no history of trauma. Insect and mosquito bites are a part of everyday life in her village, and she had not traveled elsewhere. There were no contacts with similar lesions. She was discovered during a medical mission trip to Honduras. Scrapings and small biopsies were performed and transported in sealed, insulated sterile tubes (with a minuscule amount of sterile saline) for processing in the United States. Treatment with topical antifungals and oral itraconazole was of no benefit. She was later treated with oral rifampin and clarithromycin [4, 5]. When other members of the mission team returned to the area several months later, the young woman could not be found. After return to the United States, the scrapings and biopsies were processed in the clinical microbiology laboratory of Parkland Health and Hospital System in Dallas, Texas. They were cultured in the MGIT (Becton-Dickenson) system as well as on Lowenstein-Jensen medium. Both scrapings and tissue biopsies were stained with acid-fast stains. The stains revealed clumps of acid-fast bacilli, both intra- and extracellular in location. Cultures performed at 30°C, and held for 8 weeks, grew scantily and were presumptively identified as M ulcerans by phenotypic methods. There was growth at 30°C, but not at higher temperatures, nor at 25°C. Small colonies showed rough morphology on Lowenstein-Jensen medium, they were nonpigmented, and niacin and nitrate reduction tests were negative [6]. AccuProbes for Mycobacterium tuberculosis, Mycobacterium avium, Mycobacterium gordonae, and Mycobacterium kansasii complexes were negative. Funding limitations precluded molecular characterization. We believe that this represents a case of Buruli ulcer disease acquired in coastal Honduras. No reports of this condition in Honduras were found in the PubMed, MeSH, or Cochrane Reviews databases [7, 8]. Thus, this is the first known report of Buruli ulcer disease acquired in Honduras. Figure 1.

Diseases that are caused by non-tuberculous mycobacteria (NTM) continue to pose difficult clinical problems, and the epidemiological aspect of NTM-caused diseases is of great importance. In the case of Mycobacterium gordonae there is no adequate genotyping scheme. Here we present a potential rapid and reproducible genetic assay that uses trinucleotide repeat sequence-based PCR (TRS-PCR) for genotyping M. gordonae. The proposed method constitutes a useful single-primer PCR screen for genotyping this species. Among 10 TRS-containing primers, after applying (CAC)₄-based PCR to 36 strains of M. gordonae, we found a discriminatory index of 0.975. The accuracy of this analysis was supported by a reasonable reproducibility of 92%. These results were compared with the Enterobacterial Repetitive Intergenic Consensus Sequences (ERIC)-PCR typing scheme which had lower discriminatory index of 0.93 and its reproducibility was only 86.3%.

Aims: To accelerate the identification and differentiation of clinically relevant nontuberculous mycobacteria (NTM) with two sets of multiplex PCR (mPCR) targeting the 16S–23S rRNA internal transcribed spacer (ITS) region for timely patient management. Methods and Results: Two mPCR assays were developed: Slow-Growers (SG) mPCR was used for the detection of slow-growing mycobacteria, which included Mycobacterium avium complex, Mycobacterium kansasii, Mycobacterium gordonae and Mycobacterium xenopi whilst the other mPCR assay labelled as Fast-Growers (FG) mPCR was used for the detection of Mycobacterium fortuitum complex, Mycobacterium abscessus and Mycobacterium chelonae. In these assays, a common forward primer based on a conserved section of the 16S rRNA region was used in conjunction with species-specific reverse primers. The mPCRs were tested against 247 clinical mycobacterial isolates and demonstrated 100% specificity and sensitivity. Identification of the mycobacterial species was also validated by DNA sequencing of the 16S–23S ITS region and when further confirmation was needed, hsp65 sequencing was performed. Conclusions: The mPCR assays could be a potentially useful diagnostic tool for the rapid and accurate identification of clinically relevant NTM. Significance and Impact of the Study: In this study, we looked at the frequency of hospital isolated NTM over the last 5 years (2005–2010), and an mPCR targeting the ITS region was developed for NTM species that appeared to be more prevalent in the context of Singapore.

A 69-year-old female complained of persistent abdominal pain, and annular ulcers and ulcer scars were detected endoscopically in the cecum. Pathological findings included caseous granulomas with some Langhans giant cells, and Ziehl-Neelsen staining was negative. Mycobacterium gordonae (M. gordonae) was identified by the DNA-DNA hybridization method and culture (Ogawa medium) of biopsy samples from ulcerous cecal lesions. After 6 months of antibiotic therapy, ulcerous cecal lesions were healed, and no acid-fast bacteria were detected by culture of biopsy samples from scar tissue. We believe this is the first report of M. gordonae infection in the alimentary tract.

Mycobacterium gordonae is a least pathogenic mycobacterium and a rare cause of infectionin human beings [1]. We report a case of 59 year old male with fever, cough, SOB and radiographic features of persistent left upper lobe consolidation/atelectasis with left hilar and mediastinal lymphadenopathy. Mycobacterium gordonae was isolated by DNA probe on bronchoalveolar lavage. He was observed without any active treatment for mycobacteria. He healed with minimal scarring of the lung. Mycobacterium gordonae can cause clinically significant pulmonary infection but the chances of it being just a contaminant should also be considered [1,2,6].

Summary Background The significance of nontuberculous mycobacteria (NTM) isolated from a patient during therapy for pulmonary tuberculosis (TB) is uncertain. We investigated the frequency and clinical significance of NTM isolated from patients receiving anti-TB treatment. Methods We conducted a retrospective cohort study of all patients with culture-confirmed pulmonary TB, and identified patients with respiratory cultures positive for NTM during therapy for pulmonary TB. Results From January 2003 to December 2005, 958 patients were diagnosed with culture-confirmed pulmonary TB. NTM were isolated from 113 specimens in 68 (7.1%) patients during anti-TB treatment. The most frequently isolated NTM species were Mycobacterium abscessus ( n =35, 31%), Mycobacterium fortuitum ( n =17, 15%), Mycobacterium avium complex ( n =9, 8%), and Mycobacterium gordonae ( n =9, 8%). Forty-eight (71%) patients had only one positive culture, while 20 (29%) had two or more positive cultures for NTM. Only two (3%) patients who had two or more positive culture after anti-TB treatment showed the same NTM species, which were M. abscessus . Conclusion The isolation of NTM in patients with pulmonary TB is not uncommon during anti-TB treatment. However, this is likely the result of colonization, a transient infection, or specimen contamination. The co-existence of pulmonary TB and NTM lung disease may be rare but should be considered in patients with relatively virulent NTM species such as M. abscessus .

Aims: To facilitate efficient identification of commonly encountered mycobacteria species (Mycobacterium tuberculosis, Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium fortuitum complex, Mycobacterium chelonae/abscessus, Mycobacterium kansasii, Mycobacterium gordonae) in high throughput laboratories, a 16s rDNA sequence based real-time PCR assay was developed and evaluated. Methods and Results: Oligonucleotide primers and hybridization probes were designed based on sequence differences of the mycobacterial 16S rDNA gene. This assay was evaluated with 1649 suspected non-tuberculosis mycobacterial isolates. Apart from 3 out of 40 M. avium isolates that showed false signal with M. intracellulare specific probe, 100% specificity was obtained for all tested probes. Assay sensitivity varied from 88·9 to 100% depending on species. Average cost for obtaining a definite identification was only USD 1·1 with an average turn around time of less than 3 days. Conclusions: A rapid, simple and inexpensive real-time PCR assay was developed for the identification of common encountered mycobacteria in a high throughput laboratory setting. Significance and Impact of the Study: With this assay, more than 80% of the clinically isolated nontuberculous mycobacteria could be identified in a highly cost effective manner. This helped to save resources for other laboratory activities especially in high throughput mycobacterial laboratories.

Milk is an important nutritional source to man and water buffalo raw milk is used to produce mozzarella cheese. Products from unpasteurized milk have been associated with certain infectious diseases and can carry pathogenic mycobacteria. Nontuberculous mycobacteria (NTM) are emerging pathogens causing opportunistic infections in humans and animals. The objectives of this study were to demonstrate the presence of mycobacteria in water buffaloes' milk and to determine their role as possible sources of NTM infections. In this study, raw milk samples from dairy water buffaloes (Bubalus bubalis) (N = 23) were decontaminated by Petroff method and inoculated on to Löwenstein-Jensen and Stonebrink medium. After confirming positive colonies for acid fast bacilli (AFB) by Ziehl-Neelsen technique, the isolated mycobacteria were identified by PCR-Restriction Enzyme Analysis (PRA) and mycolic acids analysis by thin-layer chromatography (TLC). Mycobacterium simiae (2 isolates), Mycobacterium kansasii (2 isolates), Mycobacterium flavescens (2 isolates), Mycobacterium gordonae (3 isolates) and Mycobacterium lentiflavum (1 isolate) were identified by these techniques. The isolation of opportunistic pathogens such as M. kansasii, M. simiae and M. lentiflavum from raw milk represent a risk for the consumers of mozzarella cheese made by this milk.

Mycobacterium gordonae is an occasional human pathogen associated with cutaneous infections and nodular granulomatous skin lesions. A case of cutaneous nodular infection caused by M. gordonae in a colony of African clawed frogs ( Xenopus tropicalis) is described and confirms this organism to be an opportunistic frog pathogen.

Background: Whole blood interferon-γ assay QuantiFERON-TB2G (QFT-2G), which is a new specific method for diagnosing tuberculosis (TB), has been developed and used in the clinical field. The aim of the present study was to assess the usefulness of QFT-2G as an indicator, both for diagnosing childhood TB and for assessing therapeutic effectiveness. Methods: The subjects were 61 children introduced to the TB outpatient department for the first time between June 2004 and March 2006. QFT-2G, the tuberculin test and chest computed tomography (CT) were performed for all patients. Results: Ten patients having typical characteristics of primary tuberculosis (PTB) on chest CT, and diagnosed as having tubercle bacillus infections, all had positive reaction on QFT-2G. Of seven patients who had no abnormalities on diagnostic imaging but who reacted positively on QFT-2G, one developed TB later, and no TB was detected over the period of observation in 44 patients with negative QFT-2G at their first consultation. Moreover, four patients with non-tuberculous acid-fast bacilli in which Mycobacterium avium or Mycobacterium gordonae was detected had negative reaction on QFT-2G. In addition, all 10 patients with positive reactions on QFT-2G in whom the subsequent course of the disease was observed had decrease on QFT after treatment. Conclusions: QFT-2G is a powerful tool with a wide application both in diagnosis and in assessment of treatment effectiveness in PTB.

An antigenic heptasaccharide motif of the cell-wall glycolipid of Mycobacterium gordonae strain 989 has been synthesized in a linear fashion by using a general glycosylation condition and minimum number of protecting group manipulation. All suitably protected monosaccharide intermediates were prepared from commercially available reducing sugars following some novel methodologies recently developed in our laboratory. Most of the synthetic intermediates were obtained as solid compounds in excellent yields.

Background Atypical mycobacteria can cause systemic infections in patients with certain types of immunodeficiency. Methods Clinical samples were decontaminated and cultured to assess the presence of mycobacterial species. Gene sequencing was performed to reveal interferon-gamma receptor 1 (IFN-gamma R1) deficiency. Results The index patient received a diagnosis of dominant IFN-gamma R1 deficiency during treatment for a serious infection due to atypical mycobacteria. She belongs to a Norwegian multiplex family comprising 3 generations and 5 patients with dominant IFN-gamma R1 deficiency. Four of these patients have been treated with tuberculostatics because of extensive infection due to atypical mycobacteria, such as Mycobacterium avium-intracellulare, Mycobacterium scrofulaceum, Mycobacterium bovis (bacille Calmette-Guerin), Mycobacterium bohemicum, and Mycobacterium gordonae. Two of the patients have also received subcutaneous injections of IFN-gamma. One family member with the deficiency has not received treatment and is still healthy at 13 years of age. Conclusions Serious infection due to atypical mycobacteria should initiate a search for primary immunodeficiencies, particularly IFN-gamma R1 deficiency. Treatment with IFN-gamma should be started when serious infection due to atypical mycobacteria is verified and dominant partial IFN-gamma R1 deficiency is suspected.

Nontuberculous mycobacteria (NTM) are ubiquitous organisms with nearly 100 different species found in soil and water. The fatty-acid and wax-rich impermeable cell wall of the mycobacteria allow for adherence to solid substrates such as pipes and leaves, allowing the organism to persist despite treatment with common disinfectants. Mycobacteria can cause infection in both humans and animals. It is difficult to assess the incidence or prevalence of NTM disease due to multiple factors. Nontuberculous mycobacteria infection may be difficult to differentiate from colonization, and when NTM infection is diagnosed, it is not a reportable disease. Furthermore, some species such as Mycobacterium gordonae may be a contaminant. Nontuberculous mycobacteria infection is not a communicable disease, although health care-associated outbreaks have been reported, associated with a single facility or procedure. While the nontuberculous infection may affect other organs, the most common site is the lung, and the most common species is Mycobacterium avium complex, commonly referred to as MAC infection. An increasing occurrence of MAC has been reported, especially in certain populations such as middle-aged or elderly thin women, patients with chronic lung disease, human immunodeficiency virus infection, and cystic fibrosis. An association of NTM infection with gastroesophageal reflux disease has also been noted. The clinical presentation often includes chronic productive cough. Other less common symptoms include dyspnea and hemoptysis. With increased use of computed tomography and high-resolution computed tomography, patterns of MAC pulmonary infection have been described. Recently, the American Thoracic Society has outlined guidelines for the diagnosis and management of NTM infection. Treatment of NTM infection requires at least 3 effective drugs for a minimum of 12 months after sputum conversion to negative cultures. Surgical therapy may be considered for localized disease which has failed medical management. In this article, the clinical presentation, radiographic features, diagnostic evaluation, and management are discussed.

We describe a patient with Mycobacterium marinum tenosynovitis associated with a fish spine injury acquired at a gambling fishing pond in southern Taiwan and identify the source of the infection. M. marinum was isolated from fishing ponds and underground water and wastewater at the site. The isolates shared the same pulsed-field gel electrophoresis pattern as the patient. M. marinum was not detected in 54 samples obtained from 27 fish. Mycobacterium gordonae was isolated from 24 samples collected from the fish. Mycobacterium abscessus was isolated from 3 fish samples (Lateolabrax japonicus 1 and Sciaenops ocellatus 2). M. abscessus and M. gordonae were isolated from all water samples. This investigation provides strong evidence that the predisposing factor for the M. marinum infection was with a fish spine injury acquired at a gambling fishing pond. The source of the infection was the contaminated pond water.

Rationale: The prevalence of pulmonary non-tuberculous mycobacterial (NTM) infection is reportedly increasing. Objectives: To study the “isolation prevalence” of pulmonary NTM in Ontario, Canada, between 1997 and 2003, and the frequency of pulmonary NTM “disease.” Methods: In a retrospective cohort, we studied “isolation prevalence” by reviewing all positive NTM culture results from the Tuberculosis and Mycobacteriology Laboratory, Ministry of Health and Long-Term Care, in Toronto, from 1997 to 2003. This laboratory identifies at least 90% of NTM isolates in Ontario, Canada. Prevalence was compared between years using with a negative binomial model. To study the frequency of “disease” (defined by American Thoracic Society criteria), we reviewed records of patients who had an isolate in 2003 and had been assessed at our hospital. Measurements and Main Results: There were 22,247 pulmonary isolates from 10,231 patients. The “isolation prevalence” of all species (excluding Mycobacterium gordonae) was 9.1/100,000 in 1997 and rose to 14.1/100,000 by 2003 (p

While nontuberculous mycobacterial peritonitis is uncommon among peritoneal dialysis (PD) patients, these infections have serious consequences. They present a significant diagnostic and therapeutic challenge for clinicians. Diagnosis can be delayed due to the slow growth rate of some mycobacterial species. These organisms can also be overlooked when adequate culture media are not used in the microbiological evaluation process. The choice of antimicrobial therapy depends upon isolation and speciation of the infecting Mycobacterium species, and prompt catheter removal is essential. Because serious intra-abdominal complications may follow infection, identifying patient risk factors for nontuberculous mycobacterial peritonitis and initiating prompt diagnosis and treatment are essential. We report three cases of peritonitis associated with Mycobacterium chelonae and Mycobacterium gordonae, each with a unique presentation, and discuss the appropriate diagnosis and treatment strategies for the management of PD-associated mycobacterial infections.

Over the last years, the clinical importance of mycobacteria has been raised. In this regard, it is important their identification in order to establish either the clinical significance or the appropriate therapy of the disease. Biochemical tests are usually time consuming until the report of results, that is why more rapid techniques are needed. As an alternative identification method, we have used a commercially available system for microbial identification based on whole cellular fatty acids analysis using gas-chromatography (GC). Sixty-eight strains of Mycobacterium tuberculosis, Mycobacterium gordonae, Mycobacterium xenopi, Mycobacterium kansasii, Mycobacterium fortuitum, and Mycobacterium avium-intracellulare were clearly identified by their unique fatty acid profile using the Sherlock Microbial Identification System (MIS). The results were in agreement with those obtained with traditional methods. This method is highly automated, rapid, easy to perform with a sample preparation for lipid analysis which is neither time consuming nor requiring a particular expertise. On this basis the MIS-GC method for the identification of some clinically important mycobacteria appears to be suitable for routine clinical use.

Background Nontuberculous mycobacteria are well known to be a cause of hot tub lung, however, to our knowledge, there exists no case report of humidifier lung induced by mycobacteria. Case presentation A case of a nonimmunocompromised female patient with Mycobacterium gordonae-induced humidifier lung is described. She spontaneously recovered after discontinuing ultrasonic humidifier use. When subjected to a provocation test, she demonstrated acute respiratory distress with signs and symptoms, consistent with hypersensitivity pneumonitis. Before and after the provocation test, water in the humidifier reservoir revealed only Mycobacterium gordonae by the microbiologic analyses. Conclusion To our knowledge, this is the first report of humidifier lung induced by nontuberculous mycobacteria. Although nontuberculous mycobacteria are well-known to be agents of hot tub lung or metal working fluid lung, physicians should also consider the pathogen as a cause of hypersensitivity lung reaction associated with humidifier use.

In comparison to the conventional real-time polymerase chain reaction method (PCR method) or the DNA-DNA hybridization method (DDH method), the utility of NTM identification by the matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) method has seldom been reported. In this study, 75 clinical NTM isolates from our hospital between April 2013 and July 2014 were identified and analyzed using PCR, DDH, and MALDI-TOF MS methods, and the results for the MALDI-TOF MS method were compared with the others. Identification at the species level was in agreement for 71 (94.5%) of the 75 isolates. For further details, identification was possible for 23 (95.8%) of 24 Mycobacterium avium, 11 (100%) of 11 Mycobacterium intracellulare, and 1 (50%) of 2 isolates mixed with M. avium and M. intracellulare. Mycobacterium ksansasii, Mycobacterium abscessus, Mycobacterium fortuitum, Mycobacterium gordonae, and Mycobacterium chelonae identified by DDH method were same result by MALDI-TOF MS. Additionally, Mycobacterium mucogenicum, which could not be identified by the DDH method, was identified by the MALDI-TOF MS method. However, two isolates identified as Mycobacterium terrae by DDH method could not be identified by the MALDI-TOF MS method and were determined to be Mycobacterium arupense by 16S ribosomal RNA (rRNA) sequence analysis. The present findings show that, for rare bacterial species, identification is sometimes not possible, but, in most cases, the results of identification by the MALDI-TOF MS method have a high concordance rate with the results of the PCR and DDH methods.

Nontuberculous mycobacteria (NTMs) are environmental microorganisms that can cause infections in humans, primarily in the lung and soft tissue. The prevalence of NTM-associated diseases is increasing in the United States. Exposure to NTMs occurs primarily through human interactions with water (especially aerosolized). Potable water from sites across the U.S. was collected to investigate the presence of NTM. Water from 68 taps was sampled 4 times over the course of 2 years. In total, 272 water samples were examined for NTM using a membrane filtration, culture method. Identification of NTM isolates was accomplished by polymerase chain reaction (PCR) amplification of the 16S rRNA and hsp65 genes. NTMs were detected in 78% of the water samples. The NTM species detected most frequently were: Mycobacterium mucogenicum (52%), Mycobacterium avium (30%), and Mycobacterium gordonae (25%). Of the taps that were repeatedly positive for NTMs, the species M. avium, M. mucogenicum, and Mycobacterium abscessus were found to persist most frequently. This study also observed statistically significant higher levels of NTM in chloraminated water than in chlorinated water.

Aim of this study was to investigate a pseudo-outbreak of Mycobacterium gordonae analyzing isolates detected from clinical and environmental samples. Mycobacterium gordonae was detected in 7 out of 497 broncho-alveolar lavage (BAL) samples after bronchoscopy procedure in patients admitted to a teaching hospital between January and April 2013. During this pseudo-outbreak clinical, epidemiological, environmental and molecular investigations were performed. None of the patients met the criteria for non-tuberculous mycobacterial (NTM) lung disease and were treated for M. gordonae lung disease. Environmental investigation revealed M. gordonae in 3 samples: in tap water and in the water supply channel of the washer disinfector. All the isolates were subjected to genotyping by pulsed-field gel electrophoresis (PFGE). The PFGE revealed that only patients' isolates presented the same band pattern but no correlation with the environmental strain was detected. Surveillance of the outbreak and the strict adherence to the reprocessing procedure and its supplies resulted afterwards in no detection of M. gordonae in clinical respiratory samples. Clinical surveillance of patients was crucial to establish the start of NTM treatment. Regular screening of tap water and endoscopic equipment should be adopted to compare the clinical strains with the environmental ones when an outbreak occurs.

Made available in DSpace on 2015-10-21T20:56:59Z (GMT). No. of bitstreams: 0 Previous issue date: 2015-03-24. Added 1 bitstream(s) on 2015-10-22T09:52:42Z : No. of bitstreams: 1 WOS000353889600144.pdf: 503172 bytes, checksum: c34e46d02eedd63a2e54d831b00aa1d2 (MD5) Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) Pro-Reitoria de Pesquisa/UNESP (ProPe) Several differences concerning bacterial species, opportunistic pathogens, elements of the resistome as well as variations concerning the CFU/mL counts were identified in some of the five most marketed bottled mineral water from Araraquara city, Sao Paulo, Brazil. Two out of five brands tested were confirmed as potential source of opportunistic pathogens, including Mycobacterium gordonae, Ralstonia picketti and Burkholderia cepacia complex (Bcc). A total of one hundred and six isolates were recovered from four of these bottled mineral water brands. Betaproteobacteria was predominant followed by Alphaproteobacteria, Gammaproteobacteria and Firmicutes. Ninety percent of the bacteria isolated demonstrated resistance to seventeen of the nineteen antimicrobials tested. These antimicrobials included eight different classes, including 3rd and 4th generation cephalosporins, carbapenems and fluoroquinolones. Multidrug resistant bacteria were detected for fifty-nine percent of isolates in three water brands at counts up to 103 CFU/ml. Of major concern, the two bottled mineral water harboring opportunistic pathogens were also source of elements of the resistome that could be directly transferred to humans. All these differences found among brands highlight the need for continuous bacteriological surveillance of bottled mineral water. Univ Estadual Paulista, Sch Pharmaceut Sci, Araraquara, SP, Brazil Univ Buenos Aires, Fac Med, Dept Microbiol, Buenos Aires, DF, Argentina Fiocruz MS, ILMD, Manaus, Amazonas, Brazil Univ Estadual Paulista, Faculdade de Ciências Farmacêuticas de Araraquara, Departamento de Ciências Biológicas, Araraquara, SP, Brazil CAPES: 18207/12-0 ProPe: 05/2012

Item does not contain fulltext A case of disseminated nontuberculous mycobacteria(l) (NTM) infection in a patient with positive neutralizing anti-interferon-gamma (IFN-gamma) autoantibodies involving bone, bronchus, systemic lymph nodes, and skin is reported. The causative NTMs were two different strains: Mycobacterium gordonae, which rarely causes true disease, and Mycobacterium mantenii, which is extremely rare. Anti-mycobacterial treatment successfully ameliorated all disseminated lesions. Although the concentration of anti-IFN-gamma autoantibodies increased during the pre-treatment period, it gradually decreased after anti-mycobacterial treatment was started.

Nontuberculous mycobacteria (NTM) are opportunistic pathogens found in natural and human-engineered waters, including drinking water distribution systems. In 2009, a relative increase in the isolation of M. gordonae from pulmonary samples originated from General Hospital Zabok was noted by the National Mycobacteria Reference Laboratory, Zagreb. Retrospective epidemiological survey was conducted to asses possible contamination of patient samples with NTM. Total of 150 individual cases with pulmonary NTM isolates were identified in the period 2007-2012, and M. gordonae was the most frequently isolated species (n=135 ; 90%). Epidemiological service performed inspection and confirmed a contamination of cold tap water with M. gordonae. None of the 150 cases met the ATS/IDSA criteria for pulmonary NTM disease, and concomitant M. tuberculosis infection was confirmed in only 6 (4%) patients. Still, due to low awareness of NTM infections, in a significant proportion of patients (n=64 ; 42.6%), tuberculosis was diagnosed and anti- tuberculosis treatment was initiated. In addition to considerable costs due to patient isolation and treatment, unnecessary physical exams, radiological and laboratory testing were performed on patients’ contacts. Our work points out the importance of faster NTM identification in patient samples as well as the continuing education of hospital personnel.

The identification of mycobacteria, other than the Mycobacterium tuberculosis complex, is a challenge for most routine clinical microbiology laboratories. Laboratories may use nucleic acid probe identification of Mycobacterium avium-M. intracellulare, Mycobacterium gordonae, and Mycobacterium kansasii, if financial resources are available. Currently, 109 species of mycobacteria have been described using nucleic acid sequencing. Phenotypic characters are generally incomplete in the literature, and for this reason, it is recommended that routine clinical microbiology laboratories refer cultures to state health laboratories or other reference laboratories for identification.

Mycobacterium gordonae（M. gordonae）による肺感染症に対し，手術を行った一例を報告する．症例は70歳，男性．原発性胆汁性肝硬変にてプレドニゾロン5mg/日の投与と糖尿病に対する内服治療を受けていた．2004年2月，右胸痛が出現したため近医を受診し，胸部X線写真で異常陰影を指摘されて当院へ紹介となった．胸部X線写真とCTで，右肺上葉に空洞性病変が認められた．気管支鏡下での気管支肺胞洗浄液から抗酸菌が検出され，DNA-DNA hybridizationによりM. gordonaeが証明された．外来で，投薬せずに経過観察されていたが，2005年3月から時々血痰が認められ，さらに空洞周囲に散布陰影が現れたためINH，RFP，EBによる化学療法を行い，同年10月，右肺上葉切除術を施行した．

L'eau des aquariums est source de Mycobactéries atypiques qui peuvent être pathogènes pour l'homme et les poissons.Une étude a été réalisée à l'aquarium du Musée de Zoologie de Nancy. La recherche de Mycobactéries a été effectuée dans 40 aquariums équipés de lampes germicides à UV : 11 bassins étaient alimentés en eau douce et 29 en eau de mer. Deux aquariums non équipés de système de désinfection ont également été analysés, les propriétaires de ces derniers ayant présenté un granulome cutané à M. marinum. L'action des UV sur M. marinum en suspension dans l'eau a été testée expérimentalement dans des bassins d'eau douce peuplés de Cichlidés.Pour chaque aquarium, un échantillon de 250 ml a été prélevé. Les cultures après décontamination au lauryl sulfate de soude ont été réalisées sur milieu de Loewenstein.Les résultats indiquent que, quel que soit l'aquarium, la présence de mycobactéries est constante. L'isolement des mycobactéries peut être gêné par la présence d'une flore bactérienne ou fongique importante (≥ 103 U.F.C./ml).Les espèces les plus fréquemment isolées sont M. gordonae et M. fortuitum ; M. kansasii et M. marinum ont rarement été isolées (6140 aquariums). Différents facteurs peuvent intervenir sur la sélection des espèces. La salinité de l'eau limite le développement de certaines espèces, alors qu'elle permet la croissance de M. fortuitum. Dans notre étude, la température de l'eau n'a pas été un facteur sélectif. L'utilisation de lampes UV limite le nombre de Mycobactéries. Dans les bassins expérimentaux, les radiations UV se sont révélées très actives sur M. marinum en présence ou en l'absence de poissons. A l'inverse de M. gordonae, M. fortuitum est rarement isolée en présence d'UV. Un nombre important de poissons par aquarium augmente la flore bactérienne et mycobactérienne.La prévention des infections à Mycobactéries atypiques chez l'homme comme chez les poissons devrait pouvoir être assurée par des mesures d'hygiène élémentaire., Water is a natural habitat of mycobacteria. In aquaria 3 species of atypical mycobacteria are frequently present : M. marinera, M. kansasitand M. fortuitum. They are potential pathogen for fishes and men. Tuberculosis has been recognized as the cause of mortality in marine and fresh water fishes. Clinical signs of fish tuberculosis are variable : ascites, skin ulcerations, skeletal deformities. The human infection is cutaneous granuloma occuring after in jury in aquaria.In the aquaria, of which two patients with cutaneous lesions due to M. marinum were analysed, UV lamps were not used. Many factors have an influence on the number of mycobacterial organisms in aquaria waters : number of fishes per tank, decontamination system, salinity, temperature.To determine the consequence of each factor, a study has been conducted at the “Aquarium du Musée de Zoologie de Nancy”. Research of mycobacteria was carried out in the water of 40 tanks : 11 were supplied with fresh water and 29 with salt water. Each tank was equipped with germicide UV (λ : 253,7 nm) : the intensity was 15 watts for aquaria smaller than 1 000 liters and 36 watts for aquaria larger than 5 000 liters. The effectiveness of UV radiation against M. marinum was tested in 3 experimental fresh water tanks of 280 liters. The first part of the experiment was tested without fish. Tank n° 1 was a control, lamp was switched on during the complete study, M. marinum was not added. In tank n° 2 (with UV) and n° 3 (without UV), 2 ml of M. marinum (of suspension 107 CF/ml) was added. Samples of water were analysed every two weeks. After six weeks tanks n° 2 and n° 3, were prepared for the next study : UV lamps were switched on in n° 2 and switched off in n° 3 both of which were contaminated by M. marinum. After 4 weeks 27 fishes, Cichlids, were introduced in the three aquaria. The day after, M. marinum was added to tanks n° 2 and n° 3. Every week water analysis was done, as well as an identification and quantification of all species of mycobacteria.From each tank 250 ml of water were collected. The water was passed through a 0,2 mµ membrane. The filters were introduced in distilled water and decontaminated by lauryl sulfate. The culture of mycobacteria was grown with Loewenstein medium at 30 and 37 °C. Each colony type was identified by cultural and biochemical characteristics.This study shows the richness in aquaria of mycobacteria; whatever the tanks, mycobacteria presence was constant. In non-treated home aquaria, the presence of mycobacteria was very important, 4 to 6 species per tank, (but in this case M. marinum was not found). In aquaria with UV lamps, the number of species per tank was lower (1 to 3).The growth of mycobacleria could be prevented when the samples were contaminated by fungi and bacteria. However, inability to recover mycobacteria from water occurred only when a massive over-growth by non-mycobacterial contaminant was present (103 CFU/ml). This was the case of non-treated tanks, belonging to patients who developed a chronic granuloma on their hands, M. marinum was not isolated in these aquaria. The evaluation of slowly growing mycobacteria could be altered by the important development of fast growing mycobacteria on the same culture tube. Among isolated species, M. fortuitum and M. gordonae saprophytic strains were frequent; M. kansasii and M. marinum involved in human cutaneous granuloma were unusual, as were the non-pigmented strains of groupe III of Runyon ; M. avium was not isolated.During this study, we observed a relationship between the mycobacteria presence and the cleanness of tank and the fishes population. A great number of fishes per tank was a factor which increased the bacterial and mycobacterial contamination. During this experiment fishes didn't present tubercular-lesions but when a dead fish was examinated, the culture from post-modem samples revealed the presence of M. marinum. The microbiological examination of skip and viscera was negative.The comparison of results in non-treated home tanks and UV treated tanks of the Museum indicates the role of water treatment by UV lamps on the number of isolated mycobacteria.The germicide UV camps are frequently used for the decontamination of tanks. The efficiency is good for bacteria, but unknown for mycobacteria. This study shows that UV radiation decreased the mycobacterial contamination. The species of mycobacteria differ in their sensitivity to UV radiation. In experimental tanks, the results showed the great susceptibility of M. marinum to UV lamps such they were used in aquaria. Presence of fish dues not change the results. If the addition of M. marinum and the lighting of lamps were simultaneous, M. marinum was not isolated in water. If the contamination by M. marinum preceded the lighting of UV lamps, must of bacteria was eliminated in one week and the totality in 4 weeks. For the other species, we observed that the mycobacterial sensitivity to UV light decreases in the following order : quickly growing mycobacteria, photochromogen and scotochromogen strains. During our experimental study, M. gordonae was isolated more frequently when UV lamps were switched on. The results obtained in the 40 tanks with UV lamps allowed the evaluation of the influence of salinity and temperature of water on mycobacterial survival and the selection of species. We did not observed a difference in the concentration of mycobacteria in two types of aquaria, fresh water and salt water. Na Cl is known as an inhibitor of the mycobacterial growth. The sensitivity of strains differs. The salinity of water appears to be a selection factor. M. forfuitum was isolated more frequently in salt water.M. marinum was isolated only in salt water and M. kansasii in fresh water. These results are surprising, as these strains have about the same metabolism. The temperature of water can also be a selection factor for mycobacteria, but in our study the temperature was similar in each aquarium (25°-26°). In this study, we did not observe thermophile strains such as M. avium.Aquarists must be informed of the aquarium contamination by atypical mycobacteria and their role in the evolution of skin lesions after injury of hands and arms. The use of germicide UV lamps improves the bacteriological quality of water.

Diagnosis of infections caused by mycobacteria, especially nontuberculous mycobacteria still represents a difficult task both in microbiology and pathology. The aim of this study was to determine the frequency of mycobacterial DNA detectable by PCR in formalin-fixed paraffin-embedded tissues showing suspicious granulomatous lesions. A total of 190 archival specimens were analyzed, using a nested PCR protocol, which amplifies a fragment of the mycobacterial 65-kDa heat-shock protein gene. Restriction fragment-length polymorphisms and sequencing were utilized to further analyze the obtained PCR products. Corresponding microbiological culture results were available for 41 cases. We detected mycobacterial DNA in 119 cases (63%), of which 71 (60%) were positive for Mycobacterium tuberculosis complex DNA and 41 (34%) for DNA of nontuberculous mycobacteria. Seven cases (6%) could not be subtyped for technical reasons. The largest group of nontuberculous mycobacteria comprised 29 cases (25% of the 119 positive cases), which were assigned to Mycobacterium fortuitum complex. Mycobacterium avium-intracellulare complex was detected in eight (7%) cases, Mycobacterium gordonae in three (2.5%) and Mycobacterium rhodesiae in a single case (0.8%). All cases of Mycobacterium tuberculosis were unequivocally identified by restriction fragment-length polymorphism analysis. In contrast, sequencing provided a gain of information over restriction fragment-length polymorphism analysis in 37% of the nontuberculous mycobacteria cases (15 of 41). Alignment studies on DNA of nontuberculous mycobacteria showed frequent sequence variations, supporting the existence of sequevars. Comparison of molecular data to available results of microbiological culture assays showed a good concordance of 83%. In conclusion, amplification and sequencing of the mycobacterial 65-kDa heat-shock protein gene is an excellent tool for species identification of mycobacteria, especially nontuberculous mycobacteria, in formalin-fixed paraffin-embedded tissues.

The mutant prevention concentration (MPC) of fluoroquinolones, linezolid, rifabutin, rifampicin and macrolides was determined in 16 nontuberculous Mycobacterium species in order to compare the incidence of resistant mutant occurrence for different antibiotics. Linezolid showed good activity against Mycobacterium gordonae and Mycobacterium kansasii. Rifabutin had a lower MPC values than rifampicin. The fluoroquinolones were more active against M. gordonae and Mycobacterium chelonae. Clarithromycin had better MPC values than azithromycin but its intracellular concentration was high. Mutant prevention concentration may help to establish a better clinico-microbiological correlation, the sub-populations of drug-resistant micro-organisms may be detected by that method. It is still important to administer correct drug doses, to use combined therapies and to find new drugs that are useful in the treatment of these infections.