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BackgroundTuberculosis (TB) transmission is influenced by patient-related risk, environment and bacteriological factors. We determined the risk factors associated with cluster size of IS6110 RFLP based genotypes of Mycobacterium tuberculosis (Mtb) isolates from Vitoria, Espirito Santo, Brazil.MethodsCross-sectional study of new TB cases identified in the metropolitan area of Vitoria, Brazil between 2000 and 2010. Mtb isolates were genotyped by the IS6110 RFLP, spoligotyping and RDRio. The isolates were classified according to genotype cluster sizes by three genotyping methods and associated patient epidemiologic characteristics. Regression Model was performed to identify factors associated with cluster size.ResultsAmong 959 Mtb isolates, 461 (48%) cases had an isolate that belonged to an RFLP cluster, and six clusters with ten or more isolates were identified. Of the isolates spoligotyped, 448 (52%) were classified as LAM and 412 (48%) as non-LAM. Our regression model found that 6-9 isolates/RFLP cluster were more likely belong to the LAM family, having the RDRio genotype and to be smear-positive (adjusted OR = 1.17, 95% CI 1.08-1.26; adjusted OR = 1.25, 95% CI 1.14-1.37; crude OR = 2.68, 95% IC 1.13-6.34; respectively) and living in a Serra city neighborhood decrease the risk of being in the 6-9 isolates/RFLP cluster (adjusted OR = 0.29, 95% CI, 0.10-0.84), than in the others groups. Individuals aged 21 to 30, 31 to 40 and > 50 years were less likely of belonging the 2-5 isolates/RFLP cluster than unique patterns compared to individuals

The present study aimed to determine the genetic diversity of isolates of Mycobacterium tuberculosis (Mtb) from presumed drug-resistant tuberculosis patients from several states of Brazil. The isolates had been submitted to conventional drug susceptibility testing for first- and second-line drugs. Multidrug-resistant (MDR-TB) (54.8%) was the most frequent phenotypic resistance profile, in addition to an important high frequency of pre-extensive resistance (p-XDR-TB) (9.2%). Using whole-genome sequencing (WGS), we characterized 298 Mtb isolates from Brazil. Besides the analysis of genotype distribution and possible correlations between molecular and clinical data, we determined the performance of an in-house WGS pipeline with other online pipelines for Mtb lineages and drug resistance profile definitions. Sub-lineage 4.3 (52%) was the most frequent genotype, and the genomic approach revealed a p-XDR-TB level of 22.5%. We detected twenty novel mutations in three resistance genes, and six of these were observed in eight phenotypically resistant isolates. A cluster analysis of 170 isolates showed that 43.5% of the TB patients belonged to 24 genomic clusters, suggesting considerable ongoing transmission of DR-TB, including two interstate transmissions. The in-house WGS pipeline showed the best overall performance in drug resistance prediction, presenting the best accuracy values for five of the nine drugs tested. Significant associations were observed between suffering from fatal disease and genotypic p-XDR-TB (p = 0.03) and either phenotypic (p = 0.006) or genotypic (p = 0.0007) ethambutol resistance. The use of WGS analysis improved our understanding of the population structure of MTBC in Brazil and the genetic and clinical data correlations and demonstrated its utility for surveillance efforts regarding the spread of DR-TB, hopefully helping to avoid the emergence of even more resistant strains and to reduce TB incidence and mortality rates. [ABSTRACT FROM AUTHOR]

Five mycobacterial isolates from sewage were classified as members of the genus Mycobacterium but presented inconclusive species assignments. Thus, the isolates (MYC017, MYC098, MYC101, MYC123 and MYC340) were analyzed by phenotypical, biochemical, matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF-MS) and genomic features to clarify their taxonomic position. Phenotypic analysis and biochemical tests did not distinguish these isolates from other non-pigmented mycobacteria. In contrast, MALDI-TOF MS analysis showed that isolates were not related to any previously described Mycobacterium species. Comparative genomic analysis showed values of ANI and dDDH between 81.59-85.56% and 24.4-28.8%, respectively, when compared to the genomes of species of this genus. In addition, two (MYC101 and MYC123) presented indistinguishable protein spectra from each other and values of ANI = 98.57% and dDDH = 97.3%, therefore being considered as belonging to the same species. Phylogenetic analysis grouped the five isolates within the Mycobacterium terrae complex (MTC) but in a specific subclade and separated from the species already described and supported by 100% bootstrap value, confirming that they are part of this complex but different from earlier described species. According to these data, we propose the description of four new species belonging to the Mycobacterium genus: (i) Mycobacterium defluvii sp. nov. strain MYC017T (= ATCC TSD-296T = JCM 35364T), (ii) Mycobacterium crassicus sp. nov. strain MYC098T (= ATCC TSD-297T = JCM 35365T), (iii) Mycobacterium zoologicum sp. nov. strain MYC101T (= ATCC TSD-298T = JCM 35366T) and MYC123 (= ATCC BAA-3216 = JCM 35367); and (iv) Mycobacterium nativiensis sp. nov. strain MYC340T (= ATCC TSD-299T = JCM 35368T). [ABSTRACT FROM AUTHOR]

Little is known about transmission and drug resistance of tuberculosis (TB) in Bauru, State of São Paulo. The objective of this study was to evaluate risk factors for transmission of Mycobacterium tuberculosis strains in this area. Strains were collected from patients attended at ambulatory services in the region and susceptibility towards the main first line antibiotics was determined and fingerprinting performed. A total of 57 strains were submitted to susceptibility testing: 23 (42.6%) were resistant to at least one drug while 3 (13%) were resistant against both rifampicin and isoniazide. Resistant strains had been isolated from patients that had not (n = 13) or had (n = 9) previously been submitted to anti-TB treatment, demonstrating a preoccupying high level of primary resistance in the context of the study. All strains were submitted to IS6110 restriction fragment length polymorphism (IS6110-RFLP) and double repetitive element PCR (DRE-PCR). Using IS6110-RFLP, 26.3% of the strains were clustered and one cluster of 3 patients included 2 HIV-infected individuals that had been hospitalized together during 16 days; clustering of strains of patients from the hospital was however not higher than that of patients attended at health posts. According to DRE-PCR, 55.3% belonged to a cluster, confirming the larger discriminatory power of IS6110-RFLP when compared to DRE-PCR, that should therefore be used as a screening procedure only. No clinical, epidemiological or microbiological characteristics were associated with clustering so risk factors for transmission of TB could not be defined in the present study.

Mycobacterium bovis is the causative agent of bovine tuberculosis (BTB), the pathogen responsible for serious economic impact on the livestock sector. In order to obtain data on isolated M. bovis strains and assist in the control and eradication program for BTB, a cross sectional descriptive molecular epidemiology study in the Brazilian Midwest was conducted. Through spoligotyping and 24-loci MIRU-VNTR methods, 37 clinical isolates of M. bovis circulating in the region were analyzed, 10 isolated from the state of Mato Grosso, 12 from the state of Mato Grosso do Sul and 15 from the state of Goiás. The spoligotyping analysis identified 10 distinct M. bovis profiles (SB0121 n = 14, SB0295 n = 6, SB0140 n = 6, SB0881 n = 3, SB1144 n = 2, SB1145 n = 2, SB0134 n = 1, SB1050 n = 1, SB1055 n = 1, SB1136 n = 1) grouped in six clusters and four orphan patterns. The MIRU-VNTR 24-loci grouped the same isolates in six clusters and 22 unique orphan patterns, showing higher discriminatory power than spoligotyping. When associating the results of both techniques, the isolates were grouped in five clusters and 24 unique M. bovis profiles. Among the 24-loci MIRU-VNTR evaluated, two, ETR-A and QUB 11b loci, showed high discriminatory ability (h = ≥ 0.50), while MIRU 16, MIRU 27, ETR-B, ETR-C, Mtub21 and QUB 26 loci showed moderate ability (h = 0.33 or h = 0.49) and were the most effective in evaluating the genotypic similarities among the clinical M. bovis isolate samples. Herein, the 29 patterns found amongst the 37 isolates of M. bovis circulating in the Brazilian Midwest can be due to the animal movement between regions, municipalities and farms, thus causing the spread of various M. bovis strains in herds from Midwest Brazil. [ABSTRACT FROM AUTHOR]

There is only scarce information available on genotypic diversity of the Mycobacterium tuberculosis complex (MTBC) clinical isolates circulating in the Northern part of Brazil, a relatively neglected region regarding research on tuberculosis. We therefore characterized 980 MTBC clinical isolates from the state of Pará, by spoligotyping and data was compared with patterns from around the world, besides analyzing drug susceptibility, and collecting sociodemographic data. We also performed 24 loci MIRU-VNTR typing to evaluate phylogenetic inferences among the East-African-Indian (EAI) lineage strains. The Geographic Information System analyses were performed to generate a descriptive visualization of MTBC strain distribution in the region. A total of 249 different spoligopatterns primarily belonging to evolutionary recent Euro-American lineages, as well as Central-Asian, Manu and ancestral EAI lineages, were identified, in addition to strains with reportedly unknown lineage signatures. The most frequent lineages were Latin American Mediterranean, T and Haarlem. Interestingly, EAI lineage strains were found in a significantly higher proportion in comparison with previous studies from South America. Regarding EAI lineage, the absence of spacers 4–9 and 23–24 co-related to 24 loci MIRU-VNTRs may suggest a close evolutionary relationship between such strains in Pará and those prevalent in Mozambique, which might have contributed to the genetic diversity of MTBC strains in this region. [ABSTRACT FROM AUTHOR]