ABSTRACT
This study evaluated the biodiversity of 28 clinical and 24 environmental Mycobacterium isolates from Rio de Janeiro, Brazil, by using hsp65 sequences, with the aim of contributing to a better understanding of the genetic diversity and usefulness of this marker. An extensive phylogenetic analysis was performed. The nucleotide diversity was similar between clinical (0.06508) and environmental (0.06221) isolates.
Subject(s)
Bacterial Proteins/genetics , Biodiversity , Chaperonins/genetics , Environmental Microbiology , Mycobacterium/classification , Mycobacterium/isolation & purification , Tuberculosis/microbiology , Animals , Brazil/epidemiology , Cattle , Chaperonin 60 , DNA, Bacterial/chemistry , DNA, Bacterial/genetics , Humans , Molecular Sequence Data , Mycobacterium/genetics , Phylogeny , Sequence Analysis, DNA , Swine , Tuberculosis/epidemiologyABSTRACT
We developed a Reverse Hybridization Assay (RHA) slightly modified from the Rifoligotyping assay and analyzed the presence of mutations in a specific region of rpoB gene in 157 isolates (90 rifampin-resistant and 67 rifampin sensitive) of Mycobacterium tuberculosis from patients attended in South and Southeast region of Brazil. Comparing to standardized drug susceptibility testing results, the sensitivity and specificity of the RHA was respectively 93% (95% IC: 86.6%-97.2%) and 100% respectively. Additionally, a high agreement (kappa coefficient 95%) between the RHA assay and sequencing was obtained. Among the 90 rifampicin-resistant isolates, RHA identified point mutations in the following codons: 42 isolates (46.6%) in 531; 29 isolates (32.2%) in 526, 6 isolates (6.7%) in 516, 3 isolates (3.3%) in 522, 2 isolates (2.2%) in 515, 514, 513 and 1 isolate (1.1%) in 511, 524 and 525. Mutations in different codons were simultaneously identified in 8 isolates (8.9%). The RHA used in the present study had a high accuracy and can be rapidly performed. However, more reproducible hybridization conditions should be looked for to increase reliability of mutant probe interpretation.
Subject(s)
Antitubercular Agents/pharmacology , Mycobacterium tuberculosis/drug effects , Nucleic Acid Hybridization/methods , Rifampin/pharmacology , DNA, Bacterial/chemistry , DNA, Bacterial/genetics , Drug Resistance, Bacterial , Genotype , Humans , Microbial Sensitivity Tests/methods , Mycobacterium tuberculosis/genetics , Mycobacterium tuberculosis/growth & development , Mycobacterium tuberculosis/isolation & purification , Point Mutation , Polymerase Chain Reaction , Sequence Analysis, DNA , Tuberculosis/microbiologyABSTRACT
The presence of mutations in specific regions of the katG, inhA, and ahpC genes was analyzed with 69 Mycobacterium tuberculosis isoniazid-resistant isolates from three Brazilian states. Point mutations in codon 315 of the katG gene were observed in 87.1, 60.9, and 60% of the isolates from Rio Grande do Sul, Rio de Janeiro, and São Paulo, respectively. Mutations in the inhA gene were identified only in one isolate from RJ State, and the ahpC promoter region revealed mutations in distinct positions in 12.9, 21.7, and 6.7% of the isolates from RS, RJ and SP, respectively.
