Background: Archaea are Extrtermophile microorganisms and for several decades it has been believed that they are only found in harsh environments, such as volcanoes, deep oceans and salt lakes. However, at present time, their existence in human and mammal’s intestine has been proved. The most important Archaea in human intestine is Methanobrevibacter smithii, which has a major role is some gastrointestinal disorders, as well as obesity. Therefore, Methanogens isolation and detection has such a crucial clinical importance. In this study, we isolated this microorganism for the first time using local technique.Materials and methods: In this study, Archaea DNA was extracted from healthy subject’s stool samples, considering the specific criteria for choosing the healthy group. PCR reaction was performed to amplify the RPOB. Enzyme digestion was operated using restriction enzyme to confirm the RPOB gene. The PCR product was then cloned in E.coli (DH5α) host and sequencing process was performed.Results: Of 20 stool samples, the RPOB gene was confirmed in 18 samples (90%) and also the AVAII enzyme digestion results proved the gene identity. Sequencing results in NCBI site proved that isolated microorganisms were Methanobrevibacter smithii.Conclusion: This study revealed that by considering the microorganisms’ variety in intestine, the precise gene detection methods for selecting the specific microbiota, in order to prevent existing similarities between homolog microbiota is vital in microbiota isolation.

Background: Rifampicin resistant tuberculosis is a serious problem faced by tuberculosis control in China, and rapid detection of rifampicin resistance is urgently needed. Objectives: This study aimed to describe the molecular characteristics and frequency of RNA polymerase ,subunit (RPOB) gene mutations in rifampicin-resistant tuberculosis (RR-TB) in the Anqing area. Methods: The RPOB gene fragment was amplified by polymerase chain reaction (PCR), and all isolates were sequenced for mutations in the RPOB gene. The mutations were obtained by comparing the sequencing results with the MUBII database. In addition, logistic regression was used to analyze the relationship between RPOB mutations and rifampicin (RIF) resistance. Results: There were 152 males and 42 females in this study, and the mean age was 56. 60 ±,17. 91 years. Mutations in the RPOB gene were a risk factor for rifampicin resistance (,= 5. 271, P < 0. 001 OR = 195. 192). Among the 19 RR-TB strains, 16 (84. 21%) had mutations in the ropB gene, and three (1. 71%) of 175 rifampicin-sensitive strains were mutated. The mutation sites of five strains (31. 58%) were at the codon 526 and five strains (31. 58%) at the codon 531. However, there were two strains at the codon 513 and two strains at the codon 533 (15. 79%), and two strains (10. 53%) were double mutations. Conclusions: The mutation characteristics of the RPOB gene in the Anqing area are complex, and RPOB mutation detection can be used as an indicator to screen drug resistance of RIF.

Background and objective: Nontuberculous mycobacteria (NTM) are a large family of acid-fast bacteria that are widely distributed in the environment. Although most NTM species are saprophytic, about 30% of these species are associated with human diseases. NTM and Mycobacterium tuberculosis complex have common microbiological characteristics, such as induce similar immune response and same clinical manifestations. However, disease caused by NTM is a diagnostic challenge for physicians, because it cannot readily be distinguished from tuberculosis on the basis of clinical history, tuberculin skin test results, radiological patterns, and initial laboratory reports. Therefore, accurate detection of NTM is necessary due to different therapeutic strategies and should be considered seriously. Materials and methods: In this study, 120 water samples were collected for isolation of NTM species, and cultured in a Lowenstein-Jensen medium after decontamination and processing. Then, accuracy and sensitivity of the three conserved genes (16S rRNA, RPOB and hsp65), were evaluated in identification of NTM species isolated. Result: In this study, 87 NTM colonies were isolated from water samples. The results of the sequence analysis of the three conserved genes showed that the hsp65 gene compared to other two genes has a high degree of accuracy and sensitivity. Conclusion: Unlike hsp65 gene, partial sequence analysis of the 16S rRNA and RPOB genes does not seem to be sufficient for recognizing NTM species.

Aim and Background: The aim of this study was to investigate the frequency, location and type of RPOB mutations in Mycobacterium tuberculosis isolated from patients in Tehran City.Materials and Methods: 65 sputums were collected from suspected tuberculosis patients, 35 Rif-r isolates were identified as Mycobacterium tuberculosis. PCR Amplification and DNA sequencing methods were performed.411 bp fragments of RPOB gene were sequenced and mutations in 81 bp regions were analyzed.Results: 38 mutations were identified in 14 RIF-r MBT (70%). Missense mutations produced 38 types of amino acid substitutions. In 6 RIF-r MBT isolates (30%) no mutations were found in the core region of the RPOB gene. Most frequent mutations detected from Tehranian strains were in codons 531 and 515. Two alleles in codon 531 and one allele in all codons 526, 515, 510, 566, 490 and 476 were found. In the 6 isolates were identified 2 mutation in different codons and 8 strains harboured single mutations in codons.Conclusion: In this study, has been investigated the significance of mutations in the RPOB gene, its correlation with genotype and phenotype agents and high level of resistance to rifampicin in 14 isolates of M. tuberculosis collected from patients with active pulmonary tuberculosis from different geographic regions of Tehran.

Background: Prevention and treatment of drug-resistant clones is important in guiding TB control strategies. The simultaneous rapid detection of the type of mutation conferring resistance and the genotype reflect the extent of drug resistant TB transmission within the communities.Mutations conferring resistance to rifampin in rifampin-resistant clinical Mycobacterium tuberculosis isolates occur mostly in the 81 bp rifampin-resistance-determining region (RRDR) of the RPOB gene.Materials and Methods: Spoligotyping, IS6110- restriction fragment length polymorphism (RFLP) typing and sequencing of the RPOB gene were performed for 30 rifampin resistant M. tuberculosis isolates from patients referred to "Iranian National TB Laboratory" from 2006 to 2007.Results: Mutations in the RRDR of the RPOB gene were identified in 96.6% of rifampin-resistant isolates. The spoligotyping analysis identified one (3.3%) East African-Indian (EAI) family, 7 (23.3%) Haarlem family, 9 (30.0%) Beijing family and 12 (40.0%) Central Asia (CAS) family isolates. Sixty- six percent of CAS isolates carried a mutation in codon 516, 37% of Beijing isolates carried a mutation in codon 531 and 33% of Haarlem isolates carried a mutation in codon 526.Conclusion: Overall, there appeared to be a correlation between the genotype and specific mutations conferring resistance to rifampin in the Beijing and Haarlem families.