h Fluoroquinolones (FQs) are broad-spectrum antibiotics recommended for the treatment of multidrug-resistant tuberculosis (MDR-TB) patients. FQ resistance, caused by mutations in the gyrA and gyrB genes of Mycobacterium tuberculosis, is increasingly reported worldwide; however, information on mutations occurring in strains from the Indian subcontinent is scarce. Hence, in this study, we aimed to characterize mutations in the gyrA and gyrB genes of acid-fast bacillus (AFB) smear-positive sediments or of M. tuberculosis isolates from AFB smear-negative samples from patients in India suspected of having MDR-TB. A total of 152 samples from patients suspected of having MDR-TB were included in the study. One hundred forty-six strains detected in these samples were characterized by sequencing of the gyrA and gyrB genes. The extracted DNA was subjected to successive amplifications using a nested PCR protocol, followed by sequencing. A total of 27 mutations were observed in the gyrA genes of 25 strains, while no mutations were observed in the gyrB genes. The most common mutations occurred at amino acid position 94 (13/27 [48.1%]); of these, the D94G mutation was the most prevalent. The gyrA mutations were significantly associated with patients with rifampin (RIF)-resistant TB. Heterozygosity was seen in 4/27 (14.8%) mutations, suggesting the occurrence of mixed populations with different antimicrobial susceptibilities. A high rate of FQ-resistant mutations (17.1%) was obtained among the isolates of TB patients suspected of having MDR-TB. These observations emphasize the need for accurate and rapid molecular tests for the detection of FQ-resistant mutations at the time of MDR-TB diagnosis.T reatment of multidrug-resistant tuberculosis (MDR-TB) patients, with strains resistant to rifampin (RIF) and isoniazid (INH), is further complicated by the presence of fluoroquinolone (FQ) resistance, due to prolonged, limited, and expensive treatment options (1, 2). A recent meta-analysis of the responses to treatment of 6,724 MDR-TB patients from 26 centers revealed that the frequency of treatment success was 64%, while that for patients with MDR-TB plus FQ resistance was only 48% (3). This clearly indicates the need for routine molecular screening for FQ resistance-associated molecular markers so that the treatment of such patients can be optimized without delay. Many such patients develop extensively drug resistant tuberculosis (XDR-TB), defined as MDR-TB plus resistance to any one FQ and any of the aminoglycosides/cyclic peptides (A-CP) (4). This poses an even more serious threat to TB management, since only 40% of XDR-TB patients have successful treatment outcomes (2). Previous studies have suggested that 5.4% (95% confidence interval [95% CI], 3.4 to 7.5%) of MDR-TB cases may actually be XDR-TB (2).FQs are oral antibacterial agents that have activity against Mycobacterium tuberculosis (5, 6). Hence, FQs are recommended for the treatment of MDR-TB patients and patients with intolerance to RIF (2). FQs are also being evaluated in ...

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A novel slowly growing, non-chromogenic species of the class Actinobacteria was isolated from a human respiratory sample in Nebraska, USA, in 2012. Analysis of the internal transcribed spacer sequence supported placement into the genus Mycobacterium with high sequence similarity to a previously undescribed strain isolated from a patient respiratory sample from Oregon, USA, held in a collection in Colorado, USA, in 2000. The two isolates were subjected to phenotypic testing and whole genome sequencing and found to be indistinguishable. The bacteria were acid-fast stain-positive, rod-shaped and exhibited growth after 7-10 days on solid media at temperatures ranging from 25 to 42°C. Colonies were non-pigmented, rough and slightly raised. Analyses of matrix-assisted laser desorption ionization time-of-flight profiles showed no matches against a reference library of 130 mycobacterial species. Full-length 16S rRNA gene sequences were identical for the two isolates, the average nucleotide identity (ANI) between their genomes was 99.7 % and phylogenetic comparisons classified the novel mycobacteria as the basal most species in the slowly growing Mycobacterium clade. Mycobacterium avium is the most closely related species based on rpoB gene sequence similarity (92 %), but the ANI between the genomes was 81.5 %, below the suggested cut-off for differentiating two species (95 %). Mycolic acid profiles were more similar to M. avium than to Mycobacterium simiae or Mycobacterium abscessus. The phenotypic and genomic data support the conclusion that the two related isolates represent a novel Mycobacterium species for which the name Mycobacterium talmoniae sp. nov. is proposed. The type strain is NE-TNMC-100812T (=ATCC BAA-2683T=DSM 46873T).

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An adult female captive-born Atlantic guitarfish ( Rhinobatos lentiginosus) was found acutely moribund on exhibit and died soon after presentation. Abnormalities on autopsy were focal cutaneous erythema on the tail, a small liver, many variably sized friable ovarian follicles, and coelomic effusion. Histologic examination revealed systemic bacterial embolization, and yolk coelomitis with minimal associated inflammation and some mineralization. Bacterial culture of blood and coelomic effusion grew a rapidly growing Mycobacterium species that was further identified as Mycobacterium chelonae by PCR amplification and sequencing of the RNA polymerase subunit beta ( rpoB) gene of isolated genomic DNA. Concurrent reproductive disease may have caused immunosuppression, thus predisposing to the mycobacterial infection. At another institution, an adult male wild-caught Atlantic guitarfish was found dead on exhibit with no premonitory signs. Abnormalities on autopsy were a thin body condition, small liver, and coelomic effusion. Histologic examination revealed acute mycobacterial septicemia. M. chelonae was also identified in this fish by PCR amplification and sequencing. Mycobacteriosis has rarely been reported in captive elasmobranchs. Guitarfish may have greater susceptibility to mycobacteriosis than other elasmobranchs, and acute and chronic manifestations of the disease may exist in this species.