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We evaluated PyroMark Q24 (QIAGEN) pyrosequencing as a method for the identification of mycobacteria, with potential application in clinical practice. Sequence data from the hypervariable region A of the 16S rRNA gene (43 and 35bp sequences) were obtained using PyroMark Q24, and a similarity search was performed automatically with PyroMark IdentiFire software. Of the 148 mycobacterial type strains tested, 138 (93.2%) were accurately identified to single or clade species level, including complex level. From the remaining 10 strains, 3 (Mycobacterium gilvum, Mycobacterium goodi, and Mycobacterium thermoresistible) showed poor sequencing quality of homopolymers. For 6 other strains (Mycobacterium cosmeticum, Mycobacterium flavescens, Mycobacterium pallens, Mycobacterium hodleri, Mycobacterium xenopi, and Mycobacterium crocinum), the sequences were unreadable from the middle, and Sanger sequencing indicated biallelic site. Finally, a 40bp sequence for Mycobacterium gordonae could not be obtained despite repeated attempts. PyroMark Q24 provided accurate identification of multiple mycobacterial strains isolated from common clinical settings, but additional gene sequencing is required to distinguish species identified as a group or complex.

Nontuberculous mycobacteria (NTM) can carry two or more 16S rRNA gene copies that are, in some instances, non-identical. In this study, we used a combined cloning and sequencing approach to analyze 16S rRNA gene sequences of six NTM species, Mycobacterium cosmeticum, M. pallens, M. hodleri, M. crocinum, M. flavescens, and M. xenopi. Our approach facilitated the identification of two distinct gene copies in each species. The two M. cosmeticum genes had a single nucleotide difference, whereas two nucleotide polymorphisms were identified in M. hodleri, M. flavescens, and M. xenopi. M. pallens had a difference in four nucleotides and M. crocinum ? in 23 nucleotides. Thus, we showed that the six NTM species possess at least two non-identical 16S rRNA gene copies. The full-length sequences of the intraspecies 16S rRNA variants will facilitate NTM identification and sequence analysis of specimens or other samples., Journal of Microbiological Methods, 155, pp.34-36; 2018

Objective/Background: Mycobacterium cosmeticum, first described in 2004, was recovered from a patient undergoing a cosmetic procedure. Subsequently, this species was associated with an outbreak in a nail salon. In all cases, the isolates were susceptible to all antibiotics tested. Recently, however, we recovered a strain of M. cosmeticum from the Chesapeake Bay, resistant to 11 of 14 antimicrobials. The objective of this work was to present our findings on the resistance and susceptibility of this isolate to various antibiotics. Materials and Methods: Surface water samples were collected from 10 sites in the Chesapeake Bay and upper tributaries to assess microbial diversity and antibiotic resistance. Site selection was based on proximity to agricultural runoff, industrial contaminants, and sewage effluents. Samples were processed and recovered organisms were identified and subjected to antimicrobial-susceptibility testing. Results: One nontuberculous species, identified as M. cosmeticum, was recovered from Sandy Point State Park. Resistance was detected to several antibiotics: doxycycline (16 μg/mL), tigecycline (≥4 μg/mL), clarithromycin (8 μg/mL), trimethoprim/sulfamethoxazole (≥8/152 μg/mL), imipenem (32 μg/mL), cefoxitin (32 μg/mL), ethionamide (≥20 μg/mL), and streptomycin (16 μg/mL). Of the 14 antibiotics tested, only the fluoroquinolones, linezolid, and amikacin demonstrated potent activity with susceptible minimum inhibitory concentrations. Conclusion: The antimicrobial resistance identified in this M. cosmeticum isolates from the Chesapeake Bay raises some important concerns: (a) why is the susceptibility pattern in this isolate so different from the previously published reports, (b) how did resistance emerge in this isolate, (c) is there a source of environmental exposure to antibiotics, (d) is it a human isolate transferred to the watershed, or (e) is it the result of lateral gene transfer with other resistant organisms in the Bay?

Mycobacterium cosmeticum is a nontuberculous Mycobacterium recovered from different water sources including household potable water and water collected at nail salon. Individual cases of this bacterium have been reported to be associated with gastrointestinal tract infections. Here we present the first whole-genome study and comparative analysis of two new clinically-derived Mycobacterium sp. UM_RHS (referred as UM_RHS after this) and Mycobacterium sp. UM_NYF (referred as UM_NYF after this) isolated from patients in Indonesia and Malaysia respectively to have a better understanding of the biological characteristic of these isolates. Both strains are likely Mycobacterium cosmeticum as supported by the evidence from molecular phylogenetic, comparative genomic and Average Nucleotide Identity (ANI) analyses. We found the presence of a considerably large number of putative virulence genes in the genomes of UM_RHS and UM_NYF. Interestingly, we also found a horizontally transferred genomic island carrying a putative dsz operon proposing that they may have potential to perform biodesulfization of dibenzothiophene (DBT) that may be effective in cost reduction and air pollution during fuel combustion. This comparative study may provide new insights into M. cosmeticum and serve as an important reference for future functional studies of this bacterial species.

A 69-year-old man renal transplant recipient for 4 years, presented with 4-day history of cough and dyspnoea. He was diagnosed with community-acquired pneumonia and treated accordingly. He deteriorated requiring intensive care unit admission and intubation. Mycobacterial culture from bronchoalveolar lavage grew colonies within 7 days of incubation while Mycobacterium tuberculosis PCR was negative. The antibiotic regimen was adjusted to cover for rapidly growing mycobacteria with imipenem, amikacin and clarithromycin. The final culture reported Mycobacterium cosmeticum. He improved on the antibiotic regimen given which the organism turned to be sensitive to. We reported the second case with M. cosmeticum that fulfilled the diagnostic criteria for non-tuberculous mycobacterial lung infection. Improvement of patient’s lung infection on appropriate antibiotics points to a causal relationship.

Multicentric Castleman's Disease (MCD) is a rare and poorly understood disorder that straddles the intersections of hematology, oncology, rheumatology, and virology and can be fatal if improperly treated. The etiology of HHV8 negative disease is unknown and only isolated cases of associations with infectious agents have been reported in literature. However the co-occurrence of Mycobacterium cosmeticum with Castleman’s disease has never been previously reported in literature. Hence we would like to report a case of Multicentric Castleman’s disease with a coexisting M. cosmeticum infection.

As few data are available in the Pacific countries and territories of the Oceania region regarding nontuberculous mycobacteria, we retrospectively identified 87 such isolates from French Polynesia from 2008 to 2013 by hybridization using DNA-strip, matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) and partialrpoBgene sequencing. PartialrpoBgene sequencing classified 42/87 (48.3%) isolates in theMycobacterium fortuitumcomplex, 28 (32.2%) in theMycobacterium abscessuscomplex, 8 (9.2%) in theMycobacterium mucogenicumcomplex, and 5 (5.7%) in theMycobacterium aviumcomplex. Two isolates were identified asMycobacterium acapulcensisandMycobacteriumcosmeticumby partial 16S rRNA gene sequencing. One isolate, unidentified by MALDI-TOF MS and yielding less than 92% and 96% sequence similarity withrpoBandhsp65reference sequences, respectively, was regarded as a potentially new species. Samples from three patients exhibiting ≥2Mycobacterium porcinumisolates and from one patient with emphysema and a lung abscess exhibiting 2Mycobacterium senegalenseisolates fulfilled the American Thoracic Society microbiological criteria for nontuberculous mycobacterial lung infection. Remote geographic areas, such as French Polynesia, are potential sources for the discovery of new mycobacterial species.

Four isolates of a rapidly growingMycobacteriumspecies had a mycolic acid pattern similar to that ofMycobacterium smegmatis, as determined by HPLC analyses. Three of the isolates were from footbath drains and a sink at a nail salon located in Atlanta, GA, USA; the fourth was obtained from a granulomatous subdermal lesion of a female patient in Venezuela who was undergoing mesotherapy. By random amplified polymorphic DNA electrophoresis and PFGE of large restriction fragments, the three isolates from the nail salon were shown to be the same strain but different from the strain from the patient in Venezuela. Polymorphisms in regions of therpoB,hsp65and 16S rRNA genes that were shown to be useful for species identification matched for the two strains but were different from those of otherMycobacteriumspecies. The 16S rRNA gene sequence placed the strains in a taxonomic group along withMycobacterium frederiksbergense,Mycobacterium hodleri,Mycobacterium diernhoferiandMycobacterium neoaurum. The strains produced a pale-yellow pigment when grown in the dark at the optimal temperature of 35 °C. Biochemical testing showed that the strains were positive for iron uptake, nitrate reduction and utilization ofd-mannitol,d-xylose, iso-myo-inositol,l-arabinose, citrate andd-trehalose. The strains were negative ford-sorbitol utilization and production of niacin and 3-day arylsulfatase, although arylsulfatase activity was observed after 14 days. The isolates grew on MacConkey agar without crystal violet but not on media containing 5 % (w/v) NaCl or at 45 °C. They were susceptible to ciprofloxacin, amikacin, tobramycin, cefoxitin, clarithromycin, doxycycline, sulfamethoxazole and imipenem. The nameMycobacterium cosmeticumsp. nov. is proposed for this novel species; two strains, LTA-388T(=ATCC BAA-878T=CIP 108170T) (the type strain) and 2003-11-06 (=ATCC BAA-879=CIP 108169) have been designated, respectively, for the strains of the patient in Venezuela and from the nail salon in Atlanta, GA, USA.

Case: A 67-year-old immunocompetent man presented with a periprosthetic infection 4 months after a left total knee arthroplasty. Repeated aspirations failed to show any organism growth. After irrigation and debridement with polyethylene exchange, there was minimal clinical improvement. Subsequently, the patient developed an arthrocutaneous fistula, which prompted a 2-stage revision arthroplasty. Mycobacterium cosmeticum grew on the culture specimens taken during this procedure, and the patient was treated with 6 months of antibiotics. Conclusion: To our knowledge, this is the first reported case of an M. cosmeticum prosthesis-related infection. The patient recovered well following a 2-stage revision arthroplasty and treatment with a Mycobacterium species-specific antibiotic.

We describe the identification and characterization of a novel nontuberculous mycobacterium (NTM), isolated from an ear swab of an adult male patient with chronic otitis externa. Genetically, the bacterium is most closely related toMycobacterium cosmeticum; however, growth and biochemical features indicate that it is distinctly different. Here, we highlight for the first time an unusual NTM that is a probable cause of ear infection.

We announce the draft genome sequence of Mycobacterium cosmeticum strain DSM 44829, a nontuberculous species responsible for opportunistic infection. The genome described here is composed of 6,462,090 bp, with a G+C content of 68.24%. It contains 6,281 protein-coding genes and 75 predicted RNA genes.

To the Editor: Mycobacterium cosmeticum is a rapidly growing nontuberculous mycobacteria species that was first described in November 2004. The first strains were obtained from cultures of a sink drain in a nail salon in Atlanta, Georgia, USA, and from a granulomatous lesion of a female mesotherapy patient in Venezuela (1). Among 3 additional isolates of M. cosmeticum obtained from July 2003 through November 2004, one was obtained from a 77-year-old man who was admitted to Ohio hospital A on September 22, 2004, with fever, exacerbation of chronic obstructive pulmonary disease, and urosepsis. Underlying medical conditions included diabetes, discitis, hyperlipidemia, coronary artery disease, and coal worker’s pneumoconiosis. He had received intravenous antimicrobial agents (rifampin and daptomycin) through a Groshong catheter that had been inserted to treat discitis. A routine blood culture was performed according to standard methods (2), and the catheter was removed. A diagnosis of catheter-associated bacteremia (CAB) was made, but the patient’s overall condition improved without antibacterial drug therapy, and he was discharged 4 days after admission. The culture specimen yielded only mycobacteria and was sent on to ARUP Laboratories, where it was identified as M. cosmeticum by 16S rDNA sequence analysis. The isolate was then sent to the Centers for Disease Control and Prevention (CDC) Mycobacteriology Laboratory Branch (Atlanta, GA, USA) and designated OH1. A 43-year-old woman with a diagnosis of non-Hodgkin lymphoma, who had received regular central venous catheterizations, was admitted to Ohio hospital B on August 20, 2004. A left subclavian catheter was inserted, and a routine blood specimen for culture was subsequently obtained on the day of admission. Before admission, the woman had been receiving acyclovir and cefepime. She received chemotherapeutic agent injections, platelet infusion, and an autologous stem cell transplant 6 days after admission. The blood culture was positive only for rapidly growing mycobacteria, and the final diagnosis was CAB. However, no symptoms of infection were observed, and no antimycobacterial drug therapy was administered. She was discharged without complications after the transplant was received and the catheter removed. The bacterial isolate was forwarded to CDC’s Special Bacteriology Reference Laboratory, where it was identified as M. cosmeticum by 16S rDNA sequence analysis, sent on to the CDC Mycobacteriology Laboratory Branch, and designated OH2. A 36-year-old man with AIDS was admitted to hospital C in Caracas, Venezuela, in June 2003 with dyspnea, fever, and expectoration. A sputum sample was positive by acid-fast bacillus smear and culture, yielding both M. cosmeticum (designated VZ1) and M. scrofulaceum on Middlebrook 7H10 agar (Remel Co., Lenexa, KS, USA). At the time the sputum was obtained, the patient was receiving only trimethoprim-sulfamethoxazole, but he experienced respiratory arrest and died ≈6 weeks later. The 3 isolates were confirmed to be M. cosmeticum by high-performance liquid chromatography mycolate analyses and by PCR restriction analysis of a 440-bp segment of hsp65 (1). The relationship of these isolates to the only documented strains of M. cosmeticum was evaluated by analysis of large restriction fragments with pulsed-field gel electrophoresis (1) and by repetitive element PCR (3). Banding patterns for isolates OH1 and OH2 were different from one another as well as from isolate VZ1 and the 2 control strains. Typing patterns for isolate VZ1, however, matched the control strain from Venezuela (ATCC BAA-878T), which indicates that these 2 isolates are likely a common strain (Figure). Figure Repetitive element (Rep)–PCR (A) and pulsed-field gel electrophoresis (PFGE) (B) patterns of Mycobacterium cosmeticum isolates from 2 patients in Ohio and 1 patient in Venezuela. Rep-PCR was performed by using BOXA1R primer (3), and PFGE was performed ... Of the >125 recognized Mycobacterium species, ≈50 are etiologic agents of human disease (4). The type strain of M. cosmeticum (ATCC BAA-878T) was associated with a soft-tissue infection in which the source was postulated to be environmental contamination of an unknown substance administered to the patient by injection as part of a weight loss regimen. This strain and isolate VZ1 were isolated from clinics in Caracas, Venezuela; both were found to be a common strain, but no other factors suggest that these represent an epidemic cluster. Although the Venezuelan patient from whom isolate VZ1 was obtained exhibited symptoms consistent with mycobacterial pulmonary disease, M. cosmeticum involvement cannot be proven because an additional Mycobacterium species, M. scrofulaceum, was isolated from the patient’s sputum. Because each of these organisms is found in the aqueous environment, they may represent colonization or may have been transiently present in the patient. Nonetheless, additional nontuberculous mycobacteria species have been reported to cause pulmonary disease, and the involvement of M. cosmeticum in this case cannot be excluded. Successful treatment of CAB infections caused by rapidly growing mycobacteria has most often been achieved by removing the catheter with or without the use of antimicrobial drug therapy (4). Criteria to support a true bloodstream infection were met by one of the patients in Ohio. These criteria include the absence of a source for bacteremia alternative to M. cosmeticum OH1 and the resolution of the febrile syndrome after removal of the device. The second patient had no symptoms when the blood culture was obtained; thus, the clinical significance of M. cosmeticum in this case is unclear. When all identified strains of M. cosmeticum are considered, this species is clearly present in diverse geographic regions and in healthcare institutions. These findings suggest that it may be widely distributed in the environment and should be regarded, along with other rapidly growing mycobacteria species, as a potential pathogen.

Mycobacterium cosmeticum is a rapidly growing mycobacterium that was recently identified. However, its role in human infection has not been well described. We present a case of an extremely premature neonate born with bacteraemia due to M. cosmeticum that resolved without sequelae. The infection likely occurred during a prolonged intrauterine transfusion procedure.

To characterize sulfate-producing microorganisms from the deteriorated sandstones of Angkor monuments in Cambodia, strains of Mycobacterium spp. were isolated from most probable number-positive cultures. All five strains isolated were able to use both elemental sulfur (S(0)) for chemolithoautotrophic growth and organic substances for chemoorganoheterotrophic growth. Results of phylogenetic and phenotypic analyses indicated that all five isolates were rapid growers of the genus Mycobacterium and were most similar to Mycobacterium cosmeticum and Mycobacterium pallens. Chemolithoautotrophic growth was further examined in the representative strain THI503. When grown in mineral salts medium, strain THI503 oxidized S(0) to thiosulfate and sulfate; oxidation was accompanied by a decrease in the pH of the medium from 4.7 to 3.6. The link between sulfur oxidation and energy metabolism was confirmed by an increase in ATP. Fluorescence microscopy of DAPI-stained cells revealed that strain THI503 adheres to and proliferates on the surface of sulfur particles. The flexible metabolic ability of facultative chemolithoautotrophs enables their survival in nutrient-limited sandstone environments.

Despite an increase in the prevalence of non-tuberculous mycobacterium (NTM), the diagnosis remains difficult and its treatment controversial. Prolonged duration of treatment is frequently needed and eradication is often unlikely.