Genomic epidemiology of a national outbreak of post-surgical Mycobacterium abscessus wound infections in Brazil.

An epidemic of post-surgical wound infections, caused by a non-tuberculous mycobacterium, has been on-going in Brazil. It has been unclear whether one or multiple lineages are responsible and whether their wide geographical distribution across Brazil is due to spread from a single point source or is the result of human-mediated transmission. 188 isolates, collected from nine Brazilian states, were whole genome sequenced and analysed using phylogenetic and comparative genomic approaches. The isolates from Brazil formed a single clade, which was estimated to have emerged in 2003. We observed temporal and geographic structure within the lineage that enabled us to infer the movement of sub-lineages across Brazil. The genome size of the Brazilian lineage was reduced relative to most strains in the three subspecies of Mycobacterium abscessus and contained a novel plasmid, pMAB02, in addition to the previously described pMAB01 plasmid. One lineage, which emerged just prior to the initial outbreak, is responsible for the epidemic of post-surgical wound infections in Brazil. Phylogenetic analysis indicates that multiple transmission events led to its spread. The presence of a novel plasmid and the reduced genome size suggest that the lineage has undergone adaptation to the surgical niche.

Multilocus sequence typing scheme versus pulsed-field gel electrophoresis for typing Mycobacterium abscessus isolates.

Outbreaks of infections by rapidly growing mycobacteria following invasive procedures, such as ophthalmological, laparoscopic, arthroscopic, plastic, and cardiac surgeries, mesotherapy, and vaccination, have been detected in Brazil since 1998. Members of the Mycobacterium chelonae-Mycobacterium abscessus group have caused most of these outbreaks. As part of an epidemiological investigation, the isolates were typed by pulsed-field gel electrophoresis (PFGE). In this project, we performed a large-scale comparison of PFGE profiles with the results of a recently developed multilocus sequence typing (MLST) scheme for M. abscessus. Ninety-three isolates were analyzed, with 40 M. abscessus subsp. abscessus isolates, 47 M. abscessus subsp. bolletii isolates, and six isolates with no assigned subspecies. Forty-five isolates were obtained during five outbreaks, and 48 were sporadic isolates that were not associated with outbreaks. For MLST, seven housekeeping genes (argH, cya, glpK, gnd, murC, pta, and purH) were sequenced, and each isolate was assigned a sequence type (ST) from the combination of obtained alleles. The PFGE patterns of DraI-digested DNA were compared with the MLST results. All isolates were analyzable by both methods. Isolates from monoclonal outbreaks showed unique STs and indistinguishable or very similar PFGE patterns. Thirty-three STs and 49 unique PFGE patterns were identified among the 93 isolates. The Simpson's index of diversity values for MLST and PFGE were 0.69 and 0.93, respectively, for M. abscessus subsp. abscessus and 0.96 and 0.97, respectively, for M. abscessus subsp. bolletii. In conclusion, the MLST scheme showed 100% typeability and grouped monoclonal outbreak isolates in agreement with PFGE, but it was less discriminative than PFGE for M. abscessus.

Characterization of non-tuberculosis mycobacteria by neutron radiography.

The genus Mycobacterium shares many characteristics with Corynebacterium and Actinomyces genera, among which the genomic guanine plus cytosine content and the production of long branched-chain fatty acids, known as mycolic acids are enhanced. Growth rate and optimal temperature of mycobacteria are variable. The genus comprises more than 140 known species; however Mycobacterium fortuitum, a fast growing nontuberculous mycobacterium, is clinically significant, because it has been associated to several lesions following surgery procedures such as liposuction, silicone breast and pacemaker implants, exposure to prosthetic materials besides sporadic lesions in the skin, soft tissues and rarely lungs. The objective of the present study is to reduce the time necessary for M. fortuitum characterization based on its morphology and the use of the neutron radiography technique substituting the classical biochemical assays. We also aim to confirm the utility of dendrimers as boron carriers. The samples were sterilized through conventional protocols using 10% formaldehyde. In the incubation process, two solutions with different molar ratios (10:1 and 20:1) of sodium borate and PAMAM G4 dendrimer and also pure sodium borate were used. After doping and sterilization procedures, the samples were deposited on CR-39 sheets, irradiated with a 4.6×10(5) n/cm(2)s thermal neutron flux for 30 min, from the J-9 irradiation channel of the Argonauta IEN/CNEN reactor. The images registered in the CR-39 were visualized in a Nikon E400 optical transmission microscope and captured by a Nikon Coolpix 995 digital camera. Developing the nuclear tracks registered in the CR-39 allowed a 1000× enlargement of mycobacterium images, facilitating their characterization, the use of more sophisticated equipment not being necessary. The use of neutron radiography technique reduced the time necessary for characterization. Doping with PAMAM dendrimer improved the visualization of NTM in neutron radiography images.

Correlation between resistance to pyrazinamide and resistance to other antituberculosis drugs in Mycobacterium tuberculosis strains isolated at a referral hospital.

The correlation between resistance to pyrazinamide (PZA) and resistance to other first-line antituberculosis drugs was investigated in 395 Mycobacterium tuberculosis strains isolated from clinical specimens, representing 14% of the overall number of M. tuberculosis isolates obtained between 2003 and 2008 at the laboratory of a referral university hospital for tuberculosis. A high correlation was found between resistance to PZA and multidrug resistance, as well as between PZA resistance and resistance to rifampin, isoniazid, and ethambutol (p < 0.01 for all). These results highlight the importance of performing PZA susceptibility testing prior to the prescription of this drug in order to treat drug-resistant and multidrug-resistant tuberculosis.