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Background: Multidrug-resistant tuberculosis (MDR-TB) is a serious threat to global tuberculosis (TB) control efforts. This study aims to investigate the trend of MDR-TB prevalence in Iran over 20 years. Methods: A systematic literature search was conducted in various databases, including PubMed, Embase, and Web of Science, from 1981 to 2023. Studies reporting the prevalence of MDR-TB in Iran were included in the meta-analysis. Statistical analysis was performed using Comprehensive Meta-Analysis software. Results: A total of 58 studies from different provinces of Iran were included in the meta-analysis. The majority of studies were from Tehran (n = 33), Kermanshah (n = 5), Mashhad (n = 4), and Tabriz (n = 4) provinces. Overall, 1885 cases of MDR-TB were reported in Iran during the study period. The highest number of MDR-TB cases was reported in 2000 (582 cases) and the lowest in 2001 (1 case). An increasing trend in MDR-TB prevalence was observed, particularly between 2018 and 2019. The pooled prevalence of MDR-TB in Iran was 12.31% (95% CI: 11.83–12.80) using the fixed-effects model and 20.21% (95% CI: 15.70–26.01) using the random-effects model. No evidence of publication bias was found. Conclusion: The results of this comprehensive meta-analysis highlight the increasing trend of MDR-TB in Iran over the past two decades. This underscores the urgent need for strengthening TB control strategies, including improved surveillance, case detection, treatment, and management of MDR-TB in the country. Developing diagnostic and treatment approaches for MDR-TB should be prioritized by Iranian medical universities and public health authorities.

Background: Pharmacogenetic research has led to significant progress in understanding how genetic factors influence drug response in tuberculosis (TB) treatment. One ongoing challenge is the variable occurrence of adverse drug reactions in some TB patients. Previous studies have indicated that genetic variations in the N-acetyltransferase 2 (NAT2) and solute carrier organic anion transporter family member 1B1 (SLCO1B1) genes can impact the blood concentrations of the first-line anti-TB drugs isoniazid (INH) and rifampicin (RIF), respectively. This study aimed to investigate the influence of pharmacogenetic markers in the NAT2 and SLCO1B1 genes on TB treatment outcomes using whole-exome sequencing (WES) analysis. Methods: DNA samples were collected from 30 healthy Iranian adults aged 18–40 years. The allelic frequencies of single-nucleotide polymorphisms (SNPs) in the NAT2 and SLCO1B1 genes were determined through WES. Results: Seven frequent SNPs were identified in the NAT2 gene (rs1041983, rs1801280, rs1799929, rs1799930, rs1208, rs1799931, rs2552), along with 16 frequent SNPs in the SLCO1B1 gene (rs2306283, rs11045818, rs11045819, rs4149056, rs4149057, rs2291075, rs201722521, rs11045852, rs11045854, rs756393362, rs11045859, rs74064211, rs201556175, rs34671512, rs71581985, rs4149085). Conclusion: Genetic variations in NAT2 and SLCO1B1 can affect the metabolism of INH and RIF, respectively. A better understanding of the pharmacogenetic profile in the study population may facilitate the design of more personalized and effective TB treatment strategies. Further research is needed to directly correlate these genetic markers with clinical outcomes in TB patients.

Background: The overexpression of efflux pumps (Eps) was reported to contribute to multidrug resistant tuberculosis (MDR-TB). Increases in Eps that expel structurally unrelated drugs contribute to reduced susceptibility by decreasing the intracellular concentration of antibiotics. In the present study, an association of mycobacterial membrane protein (MmpS5-MmpL5) Ep and its gene regulator (Rv0678) was investigated in MDR-tuberculosis isolates. Methods: MTB strains were isolated from patients at two different intervals, i.e., once when they had persistent symptoms despite 3–15 ≥ months of treatment and once when they had started new combination therapy ≥2–3 months. Sputum specimens were subjected to Xpert MTB/rifampicin test and then further susceptibility testing using proportional method and multiplex polymerase chain reaction (PCR) were performed on them. The isolates were characterized using both 16S-23S RNA and hsp65 genes spacer (PCR-restriction fragment length polymorphism). Whole-genome sequencing (WGS) was investigated on two isolates from culture-positive specimen per patient. The protein structure was simulated using the SWISS-MODEL. The input format used for this web server was FASTA (amino acid sequence). Protein structure was also analysis using Ramachandran plot. Results: WGS documented deletion, insertion, and substitution in transmembrane transport protein MmpL5 (Rv0676) of Eps. Majority of the studied isolates (n = 12; 92.3%) showed a unique deletion mutation at three positions: (a) from amino acid number 771 (isoleucine) to 776 (valine), (b) from amino acid number 785 (valine) to 793 (histidine), and (c) from amino acid number 798 (leucine) to 806 (glycine).” One isolate (7.6%) had no deletion mutation. In all isolates (n = 13; 100%), a large insertion mutation consisting of 94 amino acid was observed “from amino acid number 846 (isoleucine) to amino acid number 939 (leucine)”. Thirty-eight substitutions in Rv0676 were detected, of which 92.3% were identical in the studied isolates. WGS of mycobacterial membrane proteins (MmpS5; Rv0677) and its gene regulator (Rv0678) documented no deletion, insertion, and substitution. No differences were observed between MmpS5-MmpL5 and its gene regulator in isolates that were collected at different intervals. Conclusions: Significant genetic mutation like insertion, deletion, and substitution within transmembrane transport protein MmpL5 (Rv0676) can change the functional balance of Eps and cause a reduction in drug susceptibility. This is the first report documenting a unique amino acid mutation (insertion and deletion ≥4–94) in Rv0676 among drug-resistant MTB. We suggest the changes in Mmpl5 (Rv0676) might occurred due to in-vivo sub-therapeutic drug stress within the host cell. Changes in MmpL5 are stable and detected through subsequent culture-positive specimens.

Background: Proline-glutamate (PE)/proline-PE (PPE) proteins play an important role in the development of mycobacterial pathogenicity by modulating the host immune system. In the present investigation, the structural changes in PE-polymorphic GC-rich sequences (PGRS) family protein Wag22 (Rv1759c), PE_PGRS31 (Rv1768), PE_PGRS32 (Rv1803), and PE_PGRS33 gene (Rv1818c) were compared and analyzed in exponential state and under in vitro model of latency in same clinical isolates of Mycobacterium tuberculosis (MTB). Methods: MTB strains were isolated from clinically and laboratory-confirmed cases of tuberculosis (TB). The TB isolates were subjected to the Xpert MTB/rifampin test and then, further susceptibility testing using proportional methods was performed on them. The isolates were characterized using both 16S–23S RNA and hsp65 genes spacer polymerase chain reaction-restriction fragment length polymorphism. Selected isolates studied at two experimental set–up at exponential phase OD 600 = 0.05 (5 cfu/mL × 106 cfu/mL) and under zero oxygen and nutrition for 26 months to selected isolates studied at two experimental setup in exponential phase OD600 = 0.05 (5 cfu/mL × 106 cfu/mL) and under zero oxygen and nutrition after 26 months. Whole-genome sequencing was performed on studied isolates and the protein structures were analyzed using a bioinformatics web server. Results: No deletion, insertion, or substation occurred in susceptible, mono-drug and multidrug resistant-TB isolates were observed at PE-PGRS family protein Wag22 (Rv1759c) and PE_PGRS31 (Rv1768) at exponential phase. Although, a large deletion (at Rv1759c; Rv1768) was observed in extensively drug-resistant (XDR) and totally drug-resistant (TDR) TB isolates at the exponential phase. All studied TDR-TB isolates had a common deletion position from amino acid 1 (methionine) to amino acid 83 (glycine) and from amino acid 725 (proline) to amino acid 914 (threonine) at PE-PGRS family protein Wag22 (Rv1759c). At PE_PGRS32 (Rv1803), deletion occurred from amino acid 1 (methionine) to amino acid 212 (glycine) in latent TDR-TB bacilli. No changes in Rv1803 were observed in other studied isolates. In contrast, 66.6% of studied isolates had either insertion, deletion, substitution, or combination of changes at PE_PGRS33 (Rv1818c). However, the majority of changes at Rv1818c occurred in drug-resistant isolates. We also documented the region of deletion and insertion at PE_PGRS33 (Rv1818c) is different in active and latent TDR-TB isolates. Conclusions: Changes in these PE-PGRS family protein was associated with drug susceptibility patterns of individual isolates. Our result showed a total frameshift mutation of protein that had a different length in comparison to the original protein. These changes might disturb the interactions between XDR and TDR-TB isolates and immune responses, which needs further investigation.

Background: Recent pandemic of coronavirus SARS-CoV-2 (COVID-19) caused limitations in the country's strategies to fight against mycobacterial infections. The aim of this study was to compare the suspected tuberculosis (TB) pulmonary patients before and during the COVID-19 pandemic (January 2018–December 2021) who were referred to the National Reference TB Laboratory (NRL TB), Tehran, Iran. The mycobacterial isolated strains were identified and compared with previous data. Methods: A total of 16,899 clinical samples collected from 7041 suspected pulmonary TB patients were received from 2018 to 2021. Primary isolation of Mycobacterium isolates was done on Löwenstein–Jensen medium. Then, the DNA was extracted from acid-fast bacillus culture-positive samples and identification was performed by IS6110, Hsp65, and 16S-23S rRNA genes using polymerase chain reaction (PCR), PCR-restriction fragment length polymorphism, and nested PCR methods. Results: A total of 11679 specimens (69.1%) from 4866 suspected TB patients were collected in 2018–2019 and 5220 specimens (30.8%; from 2175 patients) in 2020–2021. Out of 11679 specimens, 2046 samples that belong to 852 patients were infected with Mycobacterium tuberculosis, and the remaining were non-TB Mycobacterium (NTM) species (n = 244) isolated from 102 patients. The cultures for 12894 specimens were either negative (76.3%) or contaminated (845/16899; 5%). A comparison of the total number of patients who were referred for diagnosis and treatment (954/666 patients, P > 0.05) showed a 30.1% reduction during the COVID-19 pandemic. Although, with these low number of patients, the significant increases of NTM species (P < 0.05) among suspected TB pulmonary patients were observed. Besides, new species of NTM, for example, Mycobacterium peregrinum and Mycobacterium montefiorense, were detected. For the past 20 years, these two species were not reported from pulmonary patients in Iran. Conclusions: During the pandemic of COVID-19, the TB diagnosis network became irregular, as a consequence, many patients could not reach the treatment center, and this could increase the circulation of mycobacterial diseases (TB and NTM). The study shows the emergence of new opportunistic NTM species also.

Background: The aim of the present study was to investigate the susceptibility of purified protein derivative (PPD) plus health-care workers to SARS-CoV-2 (COVID-19). For this reason, single-nucleotide polymorphism (SNP) of interferon-gamma (IFN-γ) gene at position +2109 and IFN-γ receptor 1 (R1) at position −56 was assessed in PPD plus group before and after COVID-19 infection (2017–2018; 2020–2021). Methods: The selected study cases (n = 100) that were working in tuberculosis (TB) unite (5–10 years) with PPD positivity >15 mm (16–20 mm) were included in this investigation. Sampling was done twice, once before and after the COVID-19 pandemic. Group A contains 50 samples collected from the GenBank TB laboratory that belong to TB staff before the pandemic (2017–2018). The other sample (Group B; 2021) was collected from the same unite during the COVID-19 pandemic. The SNP in the IFN-γ gene (+2109; 670 bp) and IFN-γ R 1 (−56; 366 bp) was performed using a specific primer and the polymerase chain reaction products were digested using restriction enzyme Fau I and Bts I, respectively. Statistical analyses were used to obtain the frequency of alleles among all studied cases. The confidence intervals (CIs) and t-test were calculated using the SPSS and GraphPad Prism software. Results: In overall, the most frequent genotype in Group A was AA (41/50; 82%) and Group B was 76% (38/50) in position + 2109 (odds ratio [OR] = 0.69, 95% CI, 0.26–1.83, and P = 0.46). Although in position −56, the most frequent genotype in Group A was TT (35/50; 70%) which significantly was than Group B TT (15/50; 30%) (OR = 0.184, 95% CI, 0.78–0.43, and P = 0.00). The frequency of allele A was more in both groups at position + 2109 (OR = 0.815, 95% CI, 0.23–2.86, and P = 0.75), whereas the dominate allele at position −56 was T in Group A (OR = 1.37, 95% CI, 0.62–3.02, and P = 0.42). Conclusion: No significant differences were observed in + 2109 in genotype among Group A and B. The main differences were seen in IFN-γ R1 at position (−56) between Group A and B. Hence, the IFN-γ R1 may play important role in COVID-19 infection. However, more study is needed to clear the IFN-γ correlation to COVID-19 infection.

Background: Pili producing genes in different life cycles of Mycobacterium tuberculosis (M. tuberculosis) were assessed. M. tuberculosis has two life cycles: dormant and active states. We aimed to assess the pili producing genes such as curli pili of M. tuberculosis (mtp) encoded by the mtp gene (Rv3312A) and fimbrial low-molecular-weight protein encoded by flp gene (Rv3656c) which were compared and analyzed. Methods: Two hundred M. tuberculosis isolates were investigated both at active and dormant states for production and expression of pili. The dormant M. tuberculosis was achieved by incubation in a sealed tube (modified Wayne method). The susceptibility of M. tuberculosis was evaluated on genes, rpob, inh, katg, and gyra by using multiplex polymerase chain reaction (PCR) and single-strand conformational polymorphism methods. The PCR–restriction fragment length polymorphism was used to express pili genes mtp and flp and then the PCR products was digested using restriction enzyme Fnu4HI, XmaI, and MspJI and AciI, TagII, and HaeII, respectively. The transmission electron microscopy was also used to detect pili in different isolates. The result was compared and analyzed using H37RV as a gold standard. Results: The mtp and flp PCR products were 263 and 122 bp in the studied strains irrespective of M. tuberculosis different life cycles, respectively. The PCR products were analyzed on 8% Polyacrylamide gel electrophoresis (PAGE), and in the 180/200 (20%), producing five fragments of 25,40,45,63,90 bp with the Fun4HI and two fragments of 126,138 bp with the XmaI and uncut with the MspJI for mtp gen were obtained at the dormant and active states of M. tuberculosis (P < 0.05). Similarly in flp gene producing three fragments of 22,35,65 bp with AciI and two fragments of 35.87 bp with TagII and two fragments of 38.84 bp with HaeII were obtained (P < 0.05). In contrast to genotyping analysis, the electron microscopy examination showed protruding of pili from M. tuberculosis, especially in dormant mycobacterium (15/100; 15%), that was multidrug resistance and extensive drug resistance isolates (P > 0.05). Conclusion: Pili were shown by electron microscopy, although at the gene expression, the insignificant difference was observed at the dormant strains in comparison to active states. Therefore, we may conclude that other genes might be involved in pili production of M. tuberculosis that needs further investigation. Although, the resistance phenomena might influence the pili producing gene expression that showed in our results.

Mycobacterium tuberculosis has infected more than two billion individuals worldwide, of whom 5%–10% have clinically active disease and 90%–95% remain in the latent stage with a reservoir of viable bacteria in the macrophages for extended periods of time. The tubercle bacilli at this stage are usually called dormant, non-viable, and/or non-culturable microorganisms. The patients with latent bacilli will not have clinical pictures and are not infectious. The infections in about 2%–23% of the patients with latent status become reactivated for various reasons such as cancer, human immunodeficiency virus infection, diabetes, and/or aging. Many studies have examined the mechanisms involved in the latent state of Mycobacterium and showed that latency modified the expression of many genes. Therefore, several mechanisms will change in this bacterium. Hence, this study aimed to briefly examine the genes involved in the latent state as well as the changes that are caused by Mycobacterium tuberculosis. The study also evaluated the relationship between the functions of these genes.

Mycobacterium tuberculosis (Mtb) and nontuberculous mycobacteria (NTM) remain the leading causes of lung disease and mortality worldwide. Interferon-gamma (IFN-γ) and its receptor (IFN-γR) play a key role in mediating immunity against Mtb and NTM. This study was conducted as a systematic review; all information was collected from databases such as: PubMed, Scopus, Medline, SID, and medical databases. Finally, all the collected data were reviewed, and all content was categorized briefly. There is growing evidence that IFN-γ plays an important role in host defense against these two intracellular pathogens by activating macrophages. In addition, IFN-γ has been shown to be an integral part of various antibacterial methods such as granuloma formation and phagosome-lysosome fusion, both of which lead to the death of intracellular Mycobacterium. As a result, its absence is associated with overgrowth of intracellular pathogens and disease caused by Mtb or Mycobacterium nontuberculosis. We also look at the role of IFN-γR in Mtb or NTM because IFN-γ acts through IFN-γR. Finally, we introduce new approaches to the treatment of M. tuberculosis complex (MTC) and NTM disease, such as cell and gene-based therapies that work by modulating IFN-γ and IFN-γR.

Objective: To investigate the prevalence of non-tuberculosis mycobacteria (NTM) among the samples deposited from the National Tuberculosis Reference Laboratory of Iran between 2011 and 2018. Methods: The study evaluated the prevalence of NTM among specimens from patients with pulmonary tuberculosis symptoms (n=15 771) deposited at the National Tuberculosis Reference Laboratory of Iran from 2011 to 2018. Detection of Mycobacterium (M.) tuberculosis was based on presence of a 190-bp amplicon from IS6110 insertion sequence using Tb1 and Tb2 primers, and amplicon-negative specimens were tested for NTM and M. tuberculosis (refractory to IS6110 amplification) using restriction fragment length polymorphism PCR of hsp65 amplicon fragment. Results: A total of 7 307 (46.33%) M. tuberculosis and 658 (4.17%) NTM specimens were found, the latter mainly comprising M. abscessus (10.18%), M. avium (2.28%), M. chelonae (8.97%), M. intracellulare (10.49%), M. kansasii (4.71%), and M. simiae (56.08%). Conclusions: As treatment for NTM differs from that for M. tuberculosis, accurate detection of Mycobacterium sp. is of public health significance.

Severe acute respiratory syndrome-coronavirus-2 (SARS-CoV-2) causes the COVID-19 infectious disease. Despite the high level of quarantine control in many well-developed countries, the disease has spread to more than 113 countries all over the world, creating a global pandemic. This emerging situation raises many questions. Can the infection be just through human-to-human transmission or are other sources, for example, animal to human or other environmental sources, also involved? We reviewed the previous literature and assessed the potential risk of transmitting the coronavirus from pets to humans. The pets (dogs and cats), especially dogs, have a habit of licking themselves in the buttocks, or smelling others in the buttocks. In such a way, they may get the virus into their respiratory or digestive tract. In return, once they lick people in the face, they may transmit the diseases. In this study, we not only reviewed the literature on pets and their relationship to the coronavirus but we also tested 22 stray dogs and cats, collected from nearby areas that were used for hospitalization of COVID-19 patients. Although no direct connection was found between these pets and the hospital, we found that 14 (63.6%) out of the 22 pets were positive for COVID-19 with the molecular test. Among them, seven (31.8%) had coughing and sneezing symptoms. The studied cases were stray pets, but the question that remains to be clarified is whether home pets are engaged in a chain of transmission or not? This needs further investigation.

Tuberculosis (TB), especially in developing countries, is a major threat to human health. The pathogenesis of TB remains poorly understood, and

In December 2019, there was an outbreak of pneumonia with an unknown cause in Wuhan, Hubei province, China. Within less than 3–4 months, the virus has spread to more than 200 countries. The virus rapidly transmitted from different places of the world with various cultural traditions. Thereby, the question is remained to be resolved about the main intermediate carrier of diseases in different animals. Studied showed that severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was originated from horseshoe bats, but its intermediate hosts are still largely unknown. Therefore, the possibility of horseshoe bat infection by ticks is high during the flight between plants or when resting in these places. Female ticks can either feed on horseshoe bat blood for 5–7 days before shedding or can form large aggregations by laying several thousand eggs in the right places or on them. Once they feed on horseshoe bat blood containing SARS-CoV-2, they can transfer the infection to the next generations without the need to reuse infections from the reservoir host. With all the potential features that this creature has, it is mandatory to investigate the presence or absences of SARS-CoV-2, especially when we know that ticks co-exist with that horseshoe bats. These factors highlight the importance of studying the viral epidemiology in the tick population. Thereby, based on the wide distribution of ticks and their co-existence with horseshoe bat, we suggest further studies on ticks that might act as one of the main intermediate hosts in different animals.

Coronavirus (COVID-19), a type of RNA virus, has a positive, single, sense stranded RNA. Studies of coronaviruses have shown high mutations in the virus's RNA. The more the virus infects people, the more the genome will mutate. In addition, evolutionary and genetic studies have the potential to recombine and easily jump from one host to another host. This means flexibility to adapt to new conditions and proliferation in new host cells. Examination of the hosts of this virus shows their extent. Studies show that the virus closely resembles hosts such as bats and pangolins. Genomic studies on severe acute respiratory syndrome coronavirus (SARS-CoV)-2 and bat coronavirus (RaTG13) showed 96.2% genetic similarity although there are differences in the nucleotides of different hosts. The importance of examining mutated areas in coronaviruses for diagnosis and treatment is necessary because molecular methods such as RT-PCR, although having a specific probe, may not cause a connection between the primer and the specific probe and cause a false-negative response be reported. This issue has led various countries to sequence SARS-CoV-2 for their own countries, eventually leading to the identification of specific drugs and vaccines for their country.

Background: Medical geographic information systems (Medical GIS) has a pronounced capacity to help us understand infectious disease epidemiology, and the transmission of pathogens in a defined geographical area over time. This could not only help us increase the understanding of public health, but also be a most useful base for medical intervention as improved infection control where it is most needed. In this study, we applied GIS to characterize Tuberculosis (TB) epidemiology in Iran. TB, caused by Mycobacterium tuberculosis, is a severe bacterial infection that can be transmitted among humans and more than 10 million people develop active TB every year and with a mortality of 1.4 million. Methods: In this cross-sectional study carried out on 3710 tuberculosis patients who had been diagnosed in 2013 to 2017 at the National Tuberculosis Reference Laboratory, we aimed to explore the geographic and gender patterns of tuberculosis in Iran using GIS. Results: Results showed that the number of patients with tuberculosis was highest in Tehran city, despite the fact that it is the richest and most developed city of Iran and that the tuberculosis dispersion reflected by GIS revealed a significant geographical heterogeneity. Conclusion: These results could be useful to establish and implement new guidelines for effective control strategies in geographic areas with the highest prevalence of tuberculosis.

Tuberculosis is a highly infectious illness that has been considered a worldwide health danger. Existence inside macrophages is a main characteristic of Mycobacterium tuberculosis virulence and essential to make a perdurable infection in the human body. Recently, a pivotal cytotoxicity determinant of M. tuberculosis in macrophages was discovered and named channel protein with necrosis-inducing toxin (CpnT). CpnT includes an N-terminal channel domain and a C-terminal domain (CTD). The CTD is a secreted toxin and can induce a necrotizing process in the macrophages. The CTD has strong nicotinamide adenine dinucleotide (NAD+)-glycohydrolase activity that empties NAD+ reservoirs of cells, resulting in human cell necrosis. In this study, the structural and functional properties of the CTD were reviewed. Besides, to predict local similarity between the CTD and other protein sequences and infer the functional and evolutionary relationships, the Basic Local Alignment Search Tool was used. Several protein sequences of the Mycobacterium showed >50% similarity to the CTD, indicating species specificity of the CTD. However, some prokaryotic and eukaryotic sequences showed 20%–45% similarity to the CTD, indicating that the CTD belongs to an uncharacterized protein family including nonbacterial proteins.

Bacteria show a plenty of cellular shapes and can alter their forms. The bacterial cell shape is functionally important. Bacteria have a number of options to select their shapes in order to uptake more nutrient, motility, attachment to surface, symmetrical division of chromosomal elements, and localization of complex secretion apparatuses. Some factors including peptidoglycan and cytoskeleton-like proteins can regulate and keep the bacterial shape. In the case of Mycobacterium tuberculosis, the reported morphological variation in the pathogen are classified into two categories; those which frequently seen at exponential phase of growth that is rod, V, Y-shape, branched, or buds, and those that are seen occasionally under stress or environmental conditions which are round, oval, ultra-virus, spore-like, and cell wall defiant or L-forms. Growth conditions and age of the cells can influence on the shape and size of the pathogen in a range from coccobacilli to long rods. Under unsuitable conditions including starvation or oxygen deprivation, tubercle bacillus assumed a swollen shape without making the vacuolar or globoid bodies. The physical circumstances and nutritional feature will control the temporary lifestyle of the pathogen.

Background : Daily intake of air is 30 Ib as compared with 2-Ib of food and 4½ of water. Thereby, breathing-contaminated air is as harmful as drinking or eating contaminated water or food, respectively. Recent research has highlighted the extent of outdoor air pollution in large cities and warranted high-quality studies to clarify the magnitude of the problem. Here, we evaluated the possible association between tuberculosis (TB) development and exposure to outdoor air pollution in a metropolitan city of Tehran. Methods: Extraction and analysis of relevant data. Investigation performed on TB patients (n = 1167) that were residing in Tehran for the past 10 years. The average concentration of sulfur dioxide (SO2), nitrogen dioxide, carbon monoxide (CO), and particles with an aerodynamic diameter of ≤ 2.5μm (PM2.5) and ≤ 10.0 (PM10) was measured from Tehran Air Quality Control Corporation (TAQCC). Patient and ecological informations were analyzed using geographical information system. Results: Based on TAQCC, Tehran had an average of 180–250 polluted days per year for the last 10 years. The high incidence of pulmonary TB (18 to 31/100,000) was detected in populations which were exposed to high concentration of CO (2.7 to 5.2 parts per million, 95% confidence interval [CI]; 1.10 to 1.90) and PM2.5(35 to 42μg/m3; 95% CI 1.03 to 1.80). The level of SO2,NO, and PM10was also high but not significantly related to TB (P > 0.05). Conclusion: The long-term exposure to PM2.5 and CO was positively associated with TB development.

Abstract Background Tuberculosis (TB) remains a significant global health concern and its diagnosis is challenging due to the limitations in the specificity and sensitivity of the current diagnostic tests. Exosomes are bioactive 30–100 nm vesicles produced by most cell types and are found in almost all human body fluids. Exosomal microRNAs (miRNAs) can transfer biological information between cells and tissues and may act as potential biomarkers in many diseases. In this pilot study, we assessed the miRNA profile of exosomes released from human monocyte-derived macrophages upon infection with Mycobacterium bovis Bacillus Calmette–Guerin (BCG). Methods Human monocytes were obtained from the peripheral blood of three healthy subjects and driven to a monocyte-derived macrophage (MDM) phenotype using standard protocols. MDMs were infected with BCG or left uninfected as control. 72 h post-infection, exosomes were collected from the cell culture medium, RNA was isolated and RNA-seq performed. The raw reads were filtered to eliminate adaptor and primer sequences and the sequences were run against the mature human miRNA sequences available in miRBase. MicroRNAs were identified using an E value

Cystic fibrosis (CF) has been generally well defined throughout the world although its prevalence is very difficult to ascertain for a number of reasons, including the fact that the medical/scientific literature and patient registries vary in quality in different countries. In the present review literature (1938–2017), we found out the higher frequency of registered CF patients in the European Union. This could be due to strong clinical awareness and greater health facilities. In contrast, we found a very irregular report from Asian and African countries. In these countries, the national CF registration system is mostly lacking or it is individualized-based research. In this situation, the estimation of various risk factors such as ethnicity and/or races is remained to be resolved. Overall, this review outlines the urgent need for revitalization of national and global CF registration, worldwide.

Background: By invention of the atomic force microscope (AFM) in 1986, the imaging of surfaces objects at nanometer-scale resolutions becomes possible. Although, in the beginning, AFM was applied almost exclusively to characterize the surfaces of nonbiological materials, at present the application of the AFM to biological and biomedical research has increased exponentially. Methods: In this study, we tried to investigate and visualize Mycobacterium tuberculosis under AFM. The transformation of bacterial shape and the formation of “hard” shell in resistant or dormant conditions were characterized and identified. Results: Application of AFM for the study of antibiotic-resistant forms of M. tuberculosis revealed the presence of round-shaped bacteria along with conventional rod-shaped ones. There has also been concluded changing of the surface charge of the cell membrane for mutated forms since round-shaped bacteria fixed on the charged surface less strongly and can be moved along the surface by a microscope tip with easily. Conclusions: In brief, this article highlights the optimal operation modes and base principal to study dangerous bacilli under AFM.

In this brief review, the geographic information systems (GISs) and its beneficial influence in the field of health were discussed and highlighted. Basically, GIS is a computerized system that can significantly add to assess and monitor the effects of environmental factors (namely, population, air pollution, location, climatic changes) on public health problems. The main benefits of GIS in health services are mapping and/or visualizing of disease distribution, which will ultimately improves our understanding of disease diversities and their spatial patterns. Therefore, GIS technology is capable of providing the intra- and inter-logical connection between health, social services, and natural environment.

Objective/Background: Mycobacterium tuberculosis (MTB) causes active tuberculosis (TB) in only a small percentage of infected people. In most cases, the infection is clinically latent, where bacilli can persist in human hosts for years without causing disease. Surprisingly, the biology of such persister cells is largely unknown. This study describes the isolation, identification, and whole-genome sequencing (WGS) of latent TB bacilli after 782 days (26 months) of latency (the ability of MTB bacilli to lie persistent). Methods: The in vitro double-stress model of latency (oxygen and nutrition) was designed for MTB culture. After 26 months of latency, MTB cells that persisted were isolated and investigated under light and atomic force microscopy. Spoligotyping and WGS were performed to verify the identity of the strain. Results: We established a culture medium in which MTB bacilli arrest their growth, reduce their size (0.3–0.1 μm), lose their acid fastness (85–90%) and change their shape. Spoligopatterns of latent cells were identical to original H37Rv, with differences observed at spacers two and 14. WGS revealed only a few genetic changes relative to the already published H37Rv reference genome. Among these was a large 2064-bp insertion (RvD6), which was originally detected in both H37Ra and CDC1551, but not H37Rv. Conclusion: Here, we show cell-wall free cells of MTB bacilli in their latent state, and the biological adaptation of these cells was more phenotypic in nature than genomic. These cell-wall free cells represent a good model for understanding the nature of TB latency.

Objective/background: Detection of mutations in the quinolone resistance-determining region (QRDR) of the gyrA gene could determine resistance to fluoroquinolone antituberculosis drugs. The aim of this study was to detect mutations in QRDRs. Methods: From 184 clinical isolates of Mycobacterium tuberculosis, ofloxacin resistance was proven in 42 isolates using the proportion method. The molecular basis of resistance to ofloxacin were investigated by the determination of mutations in the QRDR region of the gyrA gene. Extracted DNA fragments of 194bp from the gyrA gene were amplified and an automatic DNA sequencer was used for the sequencing process. Results: Molecular genetic analysis of 42 resistant M. tuberculosis strains demonstrated that they belong to Principal Genetic Group (PGG) 1 in 19 cases (45.2±10.9%), to PGG2 in 15 cases (35.7±10.5%), and to PGG3 in eight cases (19.0±8.4%). Isolates from PGG1 were dominant among resistant isolates (p

Nontuberculous mycobacteria (NTM) are a diverse group of bacterial species that are distributed in the environment. Many of these environmental bacteria can cause disease in humans. The identification of NTM in environmental sources is important for both clinical and epidemiological purposes. In this study, the distribution of NTM species from environmental and clinical samples in the Middle East was reviewed. In order to provide an overview of NTM, as well as recent epidemiological trends, all studies addressing NTM in the Middle East from 1984 to 2014 were reviewed. A total of 96 articles were found, in which 1751 NTM strains were isolated and 1084 of which were obtained from clinical samples, 619 from environmental samples and 48 were cited by case reports. Mycobacterium fortuitum was the most common rapid growing mycobacteria (RGM) isolated from both clinical (269 out of 447 RGM; 60.1%) and environmental (135 out of 289 RGM; 46.7%) samples. Mycobacterium avium complex (MAC) was the most common slow growing mycobacteria (SGM) isolated from clinical samples (140 out of 637 SGM; 21.9%). An increasing trend in NTM isolation from the Middle East was noted over the last 5 years. This review demonstrates the increasing concern regarding NTM disease in the Middle East, emphasizing the need for regional collaboration and coordination in order to respond appropriately.

Summary: The objectives of this study were to determine drug resistance pattern in new and previously treated tuberculosis (TB) patients, to assess function of TB control program, and to characterize multidrug resistant TB (MDR-TB) by molecular fingerprinting methods. Anti-micorbial susceptibility testing (AST) to the first line anti-TB drugs was performed on Löwenstein–Jensen (middlebrook 7H10) medium according to the proportion method. Molecular fingerprinting of all MDR strains was performed by spoligotyping and MIRU-VNTR. Mycobacterium tuberculosis strains were isolated from 53 Iraqi patients with pulmonary TB. Thirty eight patients (71.7%) tested cases, and 15 (28.3%) were previously treated. Four of the 38 new cases (10.5%) had resistant, of which 3 (7.9%) were MDR. Eight (53.3%) of the 15 previously treated patients had resistant strains, of which 7 (46.7%) were MDR. Spoligotyping of MDR strains showed CAS family (40%) as the predominant genotype. Using MIRU-VNTR typing, all isolates had a unique profile. MDR-TB prevalence is higher among previously treated patients than among the new cases. The many drug resistant strains, in absence of evidence of recent transmission and in combination with the many previously treated cases, highlight the need for an improved control program, coupled with a need to improve detection rate and early diagnosis of MDR-TB. Keywords: Tuberculosis, Multidrug resistant, Iraq, Drug susceptibility testing, Molecular genotyping

We report a case of pulmonary infection caused by Mycobacterium branderi, a slow growing non-tuberculosis mycobacteria, in a patient with acute myelogenous leukemia. The pulmonary disease was treated successfully with the combination of Ciprofloxacin, Doxycycline and Clarithromycin. M. branderi may be considered as an opportunistic pathogen, especially among immunologically compromised patients.

Objectives/background: The region of differences (RDs) polymorphisms is a potential molecular epidemiology method to distinguish origins of Mycobacterium tuberculosis. To date, 68 RDs have been identified in M. tuberculosis. This study was designed to determine the frequency of RD deletions in M. tuberculosis strains that were isolated from patients with pulmonary tuberculosis who were referred to the National Research Institute of Tuberculosis and Lung Disease for diagnosis and treatment. Therefore, highly polymorphic regions (RD1, RD150, and RD181) among M. tuberculosis strains isolates were investigated. Methods: A total of 250 M. tuberculosis isolates were identified by conventional and molecular methods. Subsequently, spoligotyping and RD typing (RD1, RD150 and RD181) were performed to genotype these strains. Results: The most frequent spoligotype belonged to Haarlem (n = 85, 34.0%) followed by CAS (n = 54, 21.6%), T1 (n = 27, 10.8%), and Beijing (n = 28, 11.2%) lineages. Deletion in RD181 was identified only among the Beijing lineage ([Figure 1]).{Figure 1} Conclusion: As we found a deletion in RD181 in the Beijing strains only, we propose to use this marker as an identification tool for genotyping of the Beijing strain.

Objective/background: The risk of mortality and morbidity among tuberculosis (TB) and human immunodeficiency virus (HIV) coinfected patients is significantly higher than that of patients infected with TB alone. The aim of this study was to evaluate the survival of TB-HIV patients in a TB-referral center during a 10-year follow-up. Methods: All TB-HIV patients in our referral center were enrolled in the study from 2003 to 2014, and patients were divided into two groups: HIV-TB patients without a history of TB treatment (new cases of TB) and HIV-TB patients with a history of TB treatment. Both groups were treated based on World Health Organization TB-treatment guidelines, and multivariate analysis was performed to evaluate risk factors of all-cause mortality. Results: During the study, 22 HIV-TB patients with a history of TB treatment and 263 HIV-TB patients with newly diagnosed TB were included. Baseline demographic and clinical characteristics were similar, except that miliary TB (98% vs. 2%) and mortality (97% vs. 3%; p = 0.06) were more likely in HIV patients with newly diagnosed TB. During TB treatment and subsequent follow-up, two patients did not respond to treatment and 92 (32.3%) patients died, whereas the cure rate was 60%. Pneumothorax [hazard ratio (HR): 3.17], coinfection (herpes zoster, toxoplasmosis, cytomegalovirus infection, Pneumocystis jiroveci, candidiasis, and other opportunistic infection; HR: 1.75), CD4

Objective/background: The GenoType MTBDRsl test rapidly detects resistance to ethambutol, fluoroquinolones, second-line aminoglycosides (amikacin [AMK] and kanamycin [KAN]), and cyclic peptides (capreomycin [CAP]) in Mycobacterium tuberculosis. According to data from Global Drug Resistance Surveillance Report (2007), Armenia is counted as a high-burden country for multidrug-resistant tuberculosis (MDR-TB). The estimated burden of MDR-TB in 2012 was 9.4 (7–12) and 43 (38–49) among retreatment TB cases. A total of 92 laboratory confirmed cases were reported to the World Health Organization (57 new and 35 previously treated) out of 511 cases tested for MDR-TB. Methods: A set of 77 drug-resistant TB isolates during 2011 and 2012 period, being either acid-fast bacterium positive or negative but culture-positive resistant to isoniazid, rifampin, or both according to the GenoType MTBDR plus assay, were consecutively tested using GenoType MTBDRsl. rrs gene analysis and the results from GenoType MTBDRsl were compared with phenotypic drug resistance testing. The DNA preparation method was performed as recommended by the manufacturer (Genotype MTBDR plus version 1.0 and Genotype MTBDRsl version 2.0 Hain Lifescience Nehren, Germany). Results: Aminoglycosides are key drugs for the treatment of MDR-TB. A total of 77 drug-resistant TB and four extensively drug-resistant M. tuberculosis isolates from Armenian TB patients were analyzed to characterize mutations within rrs and to compare with phenotypic drug resistance testing. Simultaneously, the following were identified: 65 (84.41%) rrs wild type (WT), 1 (1.3%) rrs WT MUT1 and MUT2 (WT; A1401G and G1484T), 1 (1.3%) rrs WT1, MUT1 (A1401G), 9 (11.7%) rrs WT1, MUT1 (A1401G), and 1 (1.3%) rrs WT1, MUT1. Mutation at position 1401 in rrs leads to resistance to KAN (7/77 = 9%), AMK (9/77 = 11.68%), and CAP (5/77 = 6.49%). Eleven (14.28%) streptomycin-resistant strains had a rrs mutation. Conclusion: Isolates with rrs structural gene mutations were cross-resistant to streptomycin, KAN, CAP, and AMK. Detection of the A1401G mutation appeared to be 100% specific for the detection of resistance to KAN and AMK. Being the first assessment, these data establish the presence of phenotypic drug-resistant and extensively drug-resistant strains using molecular profiling and are helpful in understanding aminoglycoside resistance on a molecular level.

Objective/Background: Nontuberculosis mycobacteria (NTM) are a diverse group of microorganisms that cause a variety of diseases in humans including skin, respiratory, and gastrointestinal tract infection. Generally, NTM are classified into two categories: rapid (7 days). In this study, we aimed to investigate NTM frequency and prevalence in environmental samples. Additionally, we tried to identify various subtypes of isolated rapid growing mycobacteria (RGM). Methods: Through a prospective descriptive cross-sectional study, water and soil samples were gathered from four neighboring towns around Tehran, the capital of Iran, at different geographic directions. Every 100m2 of the studied areas gave one sample containing 6g of soil in 3–5 cm depth deposited in 50 mL sterile water as sampling media. After digestion and decontamination, DNA from culture-positive specimens (RGM) were extracted using phenol–chloroform methods. Then the molecular identification of species and subspecies were performed using 16s–23s rRNA and hsp65 gene. Results: In total, 341 RGM were found, out of which 322 (94.4%) were identified and 20 (5.8%) could not be identified. The most frequent RGM was, Mycobacterium fortuitum (72; 22%), Mycobacterium senegalense (58; 17.7%), Mycobacterium parafortuitum (44; 13.4%) and Mycobacterium conceptionense type 1 (24; 7.2%), and Mycobacterium cheloni type 1 (20; 6.0%). As shown in [Table 1], M. fortuitum had more subtypes (8), and the frequency of subtypes 1 (27.7%), 4 (16.6%), and 5 (13.8%) were higher. Among subtypes of M. senegalense, subtype 1 had a higher frequency (70.4%) in comparison to subtype 2 (29.5%). M. cheloni had just one subtype.{Table 1} Conclusion: Our results showed M. fortuitum as the most prominent strain isolated from environmental samples. The frequency was similar in different places, irrespective of climatic variations. Availability of various subtypes of M. fortuitum might indicate a large circulation of this RGM in soil and water of Iranian territory. This high prevalence of M. fortuitum might raise the risk infection, especially in children, immunocompromised patients, diabetics, and cancer cases.

Objective/Background: The susceptibility to tuberculosis (TB) depends upon different factors, and the risk of developing diseases after infection with Mycobacterium tuberculosis ranges from 5% to 10%. This suggests that besides the mycobacterial itself, the host genetic factors may determine the differences in host susceptibility to TB. Among the important risk factors, cytokines and especially tumor necrosis factor alpha (TNF-α) genes, are thought to be responsible for regulating the protective immune responses. The TNF-α gene that encodes the cytokines TNF-α is located within the class III region of the major histocompatibility complex (MHC). The TNF-α gene and its receptors have significant suppressive effects on bacterial growth into macrophages. Tumor necrosis factor receptor 1 (TNFR1) is more responsible when apoptosis is needed but tumor necrosis factor receptor 2 (TNFR2) is involved in cell survival. TNF-α conducts its replicative effects on immune cells via the latter. Up to now, several polymorphisms within the promoter region of the TNF-α gene have been shown to be associated with susceptibility or resistance to TB in different ethnic groups. By contrast, the correlation of TNF-α gene with their receptors such as TNFR2 in susceptibility to TB has not been resolved yet. In this study, we aimed to analyze the single nucleotide polymorphisms (SNPs) in the TNF-α gene at the –238, –308, –857, and –863 positions, and compare the susceptibility to TB with polymorphisms at TNFR2 (T587G). Methods: One hundred fifty-one tuberculosis patients (n = 151) and 83 control subjects (n = 83) were included in this study. Polymorphisms in the TNF promoter region, namely TNF (SNP), –238, –308, –857, and –863 were studied using polymerase chain reaction–restriction fragment length polymorphism (PCR–RFLP). For TNFR2 polymorphisms at 587 positions, the following primers were used to amplify a 242 base pair (bp) product: 5'-TTCTGGAGTTGGCTGCGTGT-3' and ACTCTCCTATCCTGCCTGCT-3'. PCR products of TNFR2 digested with 2U enzymes of NLA III. Results: The current study found a strong correlation between two polymorphisms in different loci of TNF-α gene including 857 C/C (85; 56.2%) and TNF 238 A/A 127 (84.1%). However, there were no associations between polymorphisms of the TNF-α gene and its receptor, that is, TNFR2. Conclusion: Concerning our current study, screening assessments for TNF-α-857 and A238GSNPs in Iran would be important in order to make future decisions for preventive treatments before getting the disease among individuals who are at high risk considering their genotyping.

Objective/background: Tuberculosis (TB) remains the leading cause of AIDS-related deaths among adults in countries with resource limitations. The emergence of the Xpert MTB/RIF rapid molecular assay and its subsequent World Health Organization endorsement in 2010 transformed the TB-diagnostic landscape. Xpert provided diagnostic accuracy that was far superior to that of the sputum-smear microscopy test previously used. The detection of mycobacterial lipoarabinomannan (LAM) antigen in urine has emerged as a potential point-of-care test for TB. LAM antigen is a lipopolysaccharide present in mycobacterial cell walls which is released from metabolically active or degenerating bacterial cells and appears to be present only in people with active TB. Urine-based testing has advantages over sputum-based testing because urine is easy to collect and store and lacks the infection control risks associated with sputum collection. A previously study reported that urinary-LAM testing is a rapid, low-cost, ante-mortem diagnosis for human immunodeficiency virus (HIV)-associated TB. The objective of this study was to investigate the levels of LAM in HIV patients referred to the Mashih Daneshvari Hospital Tehran, Iran. Methods: Urine from 31 HIV patients without TB, 33 HIV patients with pulmonary TB, and eight HIV patients with extrapulmonary TB was analyzed for LAM using enzyme-linked immunosorbent assay kits (Mybiosource, San Diego, CA, USA). Results: The plasma levels of LAM in pulmonary TB/HIV patients was 7.67 ± 2.3 ng/ml compared with 4.5 ± 1.6 ng/ml in extrapulmonary TB/HIV and 6.7 ± 1.2 ng/ml in HIV patients without TB. There was no significant difference in urine LAM levels between the three groups. Conclusion: Our results highlight the limitations of using urine LAM levels for differentiating HIV-associated TB patients in Iran.

Background: Iran has extended borders with high-TB burden countries (Afghanistan and Pakistan) and immigrations of these populations influences TB distribution in the region and threatens the control strategies. The aim of this study was to evaluate the extent of recent TB transmission among Iranian and Afghan cases. Materials and Methods: Spoligotyping and 15-locus variable number tandem repeat (VNTR) typing were applied to genotype 102 MTB isolates (2009 to 2010). Phylogenetic relationships were analysed by two methods: a cluster-graph method and a minimum spanning tree (MST) method. Furthermore, evaluation of recent TB transmission was assessed with three indices including, RTIn, RTIn-1 and TMI. Results: Using molecular typing, 35 different spoligotypes were detected among the studied isolates. Seventy seven cases (75.4%) were distributed into 10 clusters and the remaining 25 (24.5%) isolates had a unique pattern. The cluster sizes also ranged from 2 to 21 isolates. The most frequent spoligotype in our populations belong to Haarlem (n=30, 29.4%) followed by CAS (n= 29, 28.4%) and Beijing (n=16, 15.6%) lineages. The used indices give the following values: RTIn = 0.75, RTIn-1 = 0.65 and TMI = 0.24. Conclusion: The low rate of TB transmission in our findings (24%) showed that the mode of TB transmission in Iran is mostly associated with reactivation of a previous TB infection and that recently a transmitted disease has a minor role. However, the increasing incidence of the intra-community transmission in recent years highlights the need for establishing new strategies for control of TB.

The six major lineages of Mycobacterium tuberculosis [MTB] are found to be strongly associated with specific geographical outbreaks. But whether these bacterial lineages influence the host genetic polymorphism is uncertain. The present study was designed to evaluate the relevance of strain diversity and host genetic polymorphisms in susceptibility to pulmonary tuberculosis [PTB]. For this reason, single –nucleotide polymorphisms [SNPs] in interferon- γ [IFN-γ] receptor-1[G-611A], IFNG [G+ 2109A] and tumor necrosis factors [TNF-α] genes [at -238, 308,-857position] in patients [n=151] were analyzed and compared with controls [n=83]. The genetic diversity of M. tuberculosis isolates were performed using spacer oligonucleotide typing. Thereafter, the profile of IFN-γ and TNF-α allele frequency were investigated in each subtype of M .tuberculosis. The results showed C allele of TNF 857 and A allele of TNF 238 were more frequent in PTB cases [[TNF 857 C allele OR [CI95%] 0.6[0.4-0.9], p= 0.02] for TNF 238 A allele OR [CI95%] 5.5[3.4-9.0], p= 0.00]]. Similarly, G allele in IFNG+ 2109 A/G polymorphism were significantly more in patients than control subject[OR[CI95%] 0.3; p< 0.05]. The major identified clinical isolates of M. tuberculosis were EAI[42; 27.8% ], Haarlem[ 31; 20.5% ], CAS [ 23;15.2% ], Beijing[14; 9.2%], and T [11; 7.2% ] lineages. No correction was observed between strains diversity and frequency of SNPs in studied PTB cases. In conclusions, we exclude the possibility of genetic mutation in IFN-γ and TNF-α gene by different subtypes of M .tuberculosis. Although, our results supports a positive correlation between host SNPs and susceptibility to PTB.

While NTM infection is mainly acquired from environmental exposure, monitoring of environmental niches for NTM is not a routine practice. This study aimed to find the prevalence of environmental NTM in soil and water in four highly populated suburbs of Tehran, Iran.A total of 4014 samples from soil and water resources were collected and studied. Sediments of each treated sample were cultured in Lowenstein-Jensen medium and observed twice per week for growth rate, colony morphology, and pigmentation. Colonies were studied with phenotypic tests. Molecular analysis was performed on single colonies derived from subculture of original isolates. Environmental samples were compared with 34 NTM isolates from patients who were residents of the study locations.Out of 4014 samples, mycobacteria were isolated from 862 (21.4%) specimens; 536 (62.1%) belonged to slow growing mycobacteria (SGM) and 326 (37.8%) were rapid growing mycobacteria (RGM). The five most frequent NTM were M. farcinogens (105/862; 12.1%), M. fortuitum (72/862; 8.3%), M. senegalense (58/862; 6.7%), M. kansasii (54/862; 6.2%), and M. simiae (46/862; 5.3%). In total, 62.5% (539/862) of mycobacterial positive samples were isolated from water and only 37.4% (323/862) of them were isolated from soil samples (P

Background and Objectives: Molecular epidemiology tools are widely used in determining epidemiology of tuberculosis. Spoligotyping is a molecular epidemiology method that is used for characterization and typing of Mycobacterium tuberculosis complex strains. The method is based on polymorphism of the chromosomal DR locus consisting of identical 36-bp DRs alternating with 35-41 unique spacers. The objective of this study was to investigate the prevalence of M. tuberculosis spoligotypes in different provinces of Iran. Materials and Methods: 1242 M. tuberculosis strains were isolated from TB patients of Mycobacteriology Research center (MRC). DNA was extracted from patient’s clinical samples. PCR was performed by using of specific primers for DR region. The amplified DNA was hybridized to the spoligotyping Membrane. Hybridized DNA was detected with ECL detection kit and by exposing ECL Hyperfilm to the membrane. The obtained result was entered to a binary format and was analyzed using SpolDB4 database. Results: Spoligotyping resulted in 136 different patterns. Out of 1242 M. tuberculosis strains, 1165 strains (93.8%) were classified into 59 clusters and the remaining strains (6.2 %) were singleton. Conclusions: The results of present study showed that strains of CAS family were more prevalent than other strains in Iran. Other prevalent families were Haarlem, T and Beijing, respectively.

Background and Objectives: With increasing rate of immunodeficiency diseases in the world, opportunistic micro-organism such as Pneumocystis jirovecii (P. jirovecii) become more important. Little information is available on prevalence of this life- threatening microorganism in Iran. This study was designed to determine the colonization and the rate of active disease caused by P. jirovecii in two groups of Iranian immunosuppressed patients. Materials and Methods: Two hundred and fifty five pulmonary samples were collected from two groups of immunosuppress- ed patients to detect a 260bp fragment of mt LSU rRNA gene of P. jirovecii by nested PCR. The first group was COPD patients consumed oral, inhaled or injectable corticosteroid and the second group was patients with malignancies under chemotherapy. Both groups were referred to National Research Institute of Tuberculosis and Lung Disease and Imam Khomeini hospital because of pulmonary symptoms. All patients introduced to this project were confirmed HIV sera-negative by ELISA and western blot test. Results: The mean age of COPD patients was 66.5 ± 11 (41-88) years and all of them were men. The mean age of patients with malignancy (PMs) was 43 ± 11 (23-65) years and 51.6% were men. The P. jirovecii was colonized in 7 of 89 COPD patients (7.9%) and its DNA was isolated from 11 of 153 PMs (7.2%). The microorganism could cause active disease in 7 of 67 (10.5%) PMs who suffered from pneumonia. Conclusion: The study showed that P. jirovecii was one of colonizing agents in the COPD patients, but it could cause active disease in PMs. Generally, the microorganism can exist in the lung of non-HIV+ immunosuppressed patients. Therefore, it should be considered as a potential infective agent in non-HIV+ immunocompromised patients.

Even in the 21st century, we are losing the battle against eradication of tuberculosis (TB). In 2015, 9.6 million people were estimated to have fallen ill with TB, of which 1.5 million people died. This is the real situation despite the well-structured treatment programs and availability of effective treatment options since the 1950s. The high mortality rate has been associated with other risk factors, such as the HIV epidemic, underlying diseases, and decline of socioeconomic standards. Furthermore, the problem of drug resistance that was recognized in the early days of the chemotherapeutic era raises serious concerns. Although resistance to a single agent is the most common type, resistance to multiple agents is less frequent but of greater concern. The World Health Organization estimated approximately 5% of all new TB cases involved multidrug-resistant (MDR)-TB. The estimation for MDR-TB is 3.3% for new cases, and 20.5% for previously treated cases. Failure to identify and appropriately treat MDR-TB patients has led to more dangerous forms of resistant TB. Based on World Health Organization reports, 5% of global TB cases are now considered to be extensively drug resistant (XDR), defined as MDR with additional resistance to both fluoroquinolones and at least one second-line injectable drug. XDR-TB had been reported by 105 countries by 2015. An estimated 9.7% of people with MDR-TB have XDR-TB. More recently, another dangerous form of TB bacillus was identified, which was named totally drug resistant (TDR-TB) or extremely drug resistant TB. These strains were resistant to all first- and second-line anti-TB drugs. Collectively, it is accepted that 2% of MDR-TB strains turn to be TDR-TB. This number, however, may not reflect the real situation, as many laboratories in endemic TB countries do not have proper facilities and updated protocols to detect the XDR or TDR-TB strains. Nevertheless, existing data emphasize the need for additional control measures, such as new diagnostic methods, better drugs, and more effective vaccines to prevent the spread of these strains around the world.

Objective background: Mycobacterium simiae has been identified as the most prevalent slow growing mycobacteria that has been isolated among Iranian patients. Due to an importance of molecular approaches in the treatment of nontuberculosis mycobacteria, this study aimed to evaluate molecular drug susceptibility testing against the first-line (rifampin and isoniazid) and second-line (ciprofloxacin–amikacin and kanamycin) treatment in different subtypes of M. simiae. Methods: The study involved all patients presenting to our referral tuberculosis center from March 2014 to August 2016, with confirmation of M. simiae. For all sputum samples, after digestion and decontamination, followed by DNA extraction, polymerase chain reaction-restriction fragment length polymorphism was performed using the hsp65 gene. Furthermore, susceptibility to rifampin, isoniazid, ciprofloxacin, amikacin, and kanamycin were evaluated using rpoB, inhA, katG, gyrA, and rrs genes, respectively. Results: In total, 60 cases (58.33% men and 41.66% women) of M. simiae infections were identified and all were confirmed as subtype I. All the patients were resistant to the first line-tuberculosis drugs, while 88.33% and 91.66% of cases were susceptible to ciprofloxacin and both amikacin and kanamycin, respectively. Conclusion: Regarding the given result, first-line tuberculosis drugs should be excluded from the treatment regimen of M. simiae patients. In addition, subtype I seems to be the most or even the only isolated M. simiae subtype among all Iranian patients. While both amikacin and kanamycin drugs showed better efficacy in the treatment M. simiae, the most susceptible antibiotic is still indeterminate.

Introduction: Nontuberculous mycobacteria (NTM) are opportunistic pathogens that have increased in recent years. The distribution of NTM species is not similar, and it seems to depend on geographic and environmental regions. In this study, the distribution of NTM species from environmental and clinical samples in the Middle East was reviewed. Methods: To provide an overview of NTM, all studies addressing NTM in the Middle East were searched from 1984 to 2014; 111 articles were found describing the prevalence of NTM in this region. Results: A total of 1994 and 886 NTM were included in the clinical and environmental samples, respectively. Mycobacterium fortuitum was the predominant NTM identified among the rapidly growing mycobacteria (RGM) in both environmental (32.6%) and clinical (53.1%) samples. The most common slowly growing mycobacteria (SGM) were MAC 297 (25.3%) in clinical cases. Among SGM in environmental samples, the most common species was Mycobacterium MAC 49 (11%) which reported from Iraq, Iran and Saudi Arabia. Conclusion: This is the first study reporting NTM species in the Middle East. In summary, this review demonstrates that the frequency of NTM has increased in this region. In addition to the limitations of the NTM study in some countries of the Middle East, there is an urgent need to develop laboratory methods for NTM identification to prepare a comprehensive report for this region.

Background: Limited evidence showed the persistence of Mycobacterium tuberculosis (MTB) in soil and water of TB sanatoriums or hospitals, but no reports suggest the occurrence of MTB in nature. Here, this study investigates the probability of isolating MTB from the environment with no clear source of contamination. To further highlight the transmission vehicle from human to nature and vice versa, the typing patterns of isolated MTB strains from the environment was compared with clinical MTB isolates. Additionally, the viability of MTB after 3 and 6 months of contamination was investigated. Methods: The soil (n = 700) and water (n = 800) from different locations of suburban towns of Tehran was examined for the presence of MTB. The digested and decontaminated samples were inoculated in three LJ (Lowenstein Jensen) mediums and incubated at 37 °C, 25 °C and 42 °C for 12 weeks. Spoligotypes and MIRU-VNTR typing methods were performed on DNA extracted from single colonies. Thereafter, the obtained typing patterns were compared with the genotyping of TB patients that were residents of Robat Karim (n = 25), Firuzkuh (n = 10), Shahr-e-Ray (n = 20) and Tehran (n = 413). Result: MTB accounts for 14.4% of total mycobacterium that was isolated from soil and water in suburban towns of Tehran. Based on polymerase chain reaction (PCR) typing methods, T family (56/82; 68%) followed by Delhi/CAS (11/82; 13.4%) were the most frequent MTB super families in both water and soil samples. Overall, 27.7% of isolates in clusters were intra-connected strains, i.e., found in different regions; however, no similar typing pattern between soil, water and clinical isolates was observed. The most frequent super family of MTB in clinical isolates was Delhi/CAS (142; 30.3%) followed by NEW-1 (127; 27.1%). The environment and clinical isolates had a minimum of 2 and a maximum of 10loci difference (5–7 difference on average). The bacilli in contaminated soil (36%) and damp water (8.4%) remained alive for a considerable period of time, i.e., up to 9 months. Conclusion: Persistence of MTB in soil and water highlights the risk of transmission, which needs further investigation. This study was supported by the National Research Institute of Tuberculosis and Lung Disease of Iran, Iran.

Introduction: This study was conducted to assess the rate of human P2X7 and TNF-α gene polymorphisms among Iranian pulmonary tuberculosis (TB) cases. P2X7 gene is highly polymorphic, and it is up-regulated by an inflammatory cytokine. Recently, a correlation between the expression of the P2X7 receptor and the elevated levels of TNF-α have been shown, but the apparent clinical correlation could not be demonstrated. Methodology: This study included 80 confirmed pulmonary tuberculosis (PTB) cases. Fifty control subjects were selected from the TB patients (clinical and laboratory) who had positive tuberculin tests (10–15 mm) and showed no sign of diseases (laboratory, radiological and clinical parameter). Single nucleotide polymorphisms (SNPs) in P2X7 (+1513, −762) and TNF-α (at −238, −308, −244, −857, −863) genes were assessed using PCR-RFLP and allele-specific PCR. Thereafter, haplotype and diplotype variability were compared and analyzed. Results: For 1513 loci, the heterozygosity was higher in patients (35%; 44.3%) than control subjects (12%; 24%) [(p = 0.026) ORS; 2.45 CI 95% (1.13–5.33)]. For −762 loci, the frequency of mutant alleles between patients and controls were not statistically significant. No statistical difference was observed in allele frequencies of TNF −308 and −857. However, the frequency of −238 allele were more in TB cases (72.1%) (p = 0.000)[ORs: 5.85(2.70–12.64). Data analysis showed more frequencies of haplotypes, i.e., TGGA-CA and CGGA-TA in patients (21.5%; 14.6%) than the control group (2.0%; 6.0%), respectively. Additionally, the diplotype “CCGGGGGG-CCAA” was significantly associated with susceptibility to PTB (1.9 [0.08–48.3]). Conclusions: In the studied population, polymorphisms in P2X7 (1513) and TNF-α (−238) gene was associated with the risk of developing PTB. Additionally, distribution of haplotype and diplotype variables did appear to be more specific than SNPs.

Background and objective: Nontuberculosis mycobacteria (NTM) are free-living saprophytes that are found in water, soil, animals and dairy products. During recent years, advances in molecular biology improved the recovery and accuracy of NTM identification. In the present study, similar approaches were used to differentiate environmental NTM. For this reason, the prevalence of environmental NTM in soil and water in two densely populated areas of Tehran were investigated. Material and Methods: A total of 2,540 samples were collected from soil (n = 1237) and water (n = 1303) resources. For the soil sample: digestion and decontamination were performed using modified Engbaeks method, and for the water sample: decontamination followed with ethylpyridinium chloride (CPC; final concentration of 0.05%) and standard protocol. The phenotypic and genotyping tests were performed on all positive cultures. For molecular tests, both 16S-23S RNA and hsp65 genes spacer PCR-RFLP were used. Results: Of the 2,540 collected samples, 285 (12%) were positive for mycobacterium. One hundred and sixty-nine (169; 60%) belonged to a slow growing mycobacteria (SGM) and 116 (41%) were rapid growing mycobacteria (RGM). The most frequent NTM were identified as Mycobacterium farcinogenes (51/485; 10.5%) and Mycobacterium fortuitum (38/485; 7.8%). However, M. farcinogenes is found to be dominant in both studied areas, but the frequency of RGM was different. In Shahre-ray, Mycobacterium senegalense was identified as the most frequent RGM (22/65; 34%), while in Varamin, M. fortuitum (20/51; 40%) had the most frequency. Conclusion: The molecular genotyping methods facilitate the rapid identification of environmental NTM. As the higher prevalence of NTM within a region increases the risk of human infection, this study outlines the importance of further investigation.

Introduction: In many developing countries, like Iran, newborns receive the BCG vaccination for prophylaxis against mycobacterial infections. Although in many children or infants BCG infection after BCG vaccination is rare, children with primary immunodeficiency are at high risk of developing BCG-osis. According to many studies, some cytokines such as IL-12 play a major role in immunological responses in dealing with mycobacterial infections. Therefore, this research studies the relationship between polymorphism of genes encoding this cytokine and BCG-osis in Iranian children. Method: Thirty-four children up to 7 years old with BCG-osis and 34 healthy children vaccinated with BCG during the first two days of life were chosen for this analysis. DNA samples were extracted from both groups. Single nucleotide polymorphisms (SNPs) in IL-12 (at +705, +1158, +1196 and +1637) genes were assessed using PCR-RFLP. Allele and genotype frequencies in cases and controls were compared using the Pearson Chi-Square test. Result: The prevalence of IL-12 705 AG (heterozygote mutant) genotype was 50% in patients, but in the control group this rate was 38.2%. Likewise, AA (frequent hemozygote) genotypes were 52.9% and 38.2% in patients and control groups, respectively (p-Value >0.05). It was observed only homozygote alleles for 1158, 1196, 1637 and 1664 positions in both patients and control groups. However, there was no significant association between case and control groups. Conclusions: BCG vaccination in infants can cause protective immunity, and IL-12 plays an important role in immune responses. However, this study did not show any significant association between polymorphisms in IL-12 (at +705, +1158, +1196 and +1637) and risk of developing BCG-osis.

Background: Scaling-up in treatment for Mycobacterium tuberculosis (MTB) is a health priority. Currently, treatment regimens are long with complications, and default rates are very high. Tuberculosis (TB) patients who do not complete treatment pose a potential public health risk and are categorized as default to follow-up (DFU). Methods: In 2014, an anti-TB drug-resistance survey in Tehran, Iran, enrolled 1718 new and re-treatment patients who were TB, MDR-TB (multidrug-resistant) and old TB. TB was detected in these patients with positive acid fast bacilli (AFB) in smear and culture of sputum. Results: All patients received treatment with standardized first-line or second-line regimens (based on the type of TB infection). Patients are followed from the time of registration and treatment until the completion of treatment. This study reports the treatment outcomes of the retrospective study which was assessed from 2004 up to 2014. The reported cure rate for TB patients was 97.5% among new and old cases; 85.9% of patients (n = 1476) had no relapse; 13.7% of them (n = 236) had two or less than two relapses, and only 6 cases (0.4%) had more than two relapses; 10.5% of patients (n = 183) defaulted from anti-TB treatment and have not come back for follow-up after 52 weeks. Conclusions: These results showed the achievement of the National Tuberculosis Control Program implementation. It can be concluded that the treatment plans and appropriate follow-up of therapy have the greatest success in improving TB treatment outcomes.

Background: The infections due to rapidly growing mycobacteria (RGM) are becoming an important health problem in many countries in the world. Globally, an increase in RGM infections is being reported from several regions worldwide. However, there is limited information about the prevalence of these kinds of organisms in Iran. Methods: The relevant data of the prevalence of RGM were retrieved by searching several databases, such as PubMed, Web of Science, Cochrane Library, Embase, Scopus, Iranmedex, and Scientific Information Database. Meta-analysis was performed by Comprehensive Meta-Analysis (V2.0, Biostat) software. Results: The meta-analyses showed that the Mycobacterium fortuitum (22.7% [95% CI 16.1–30.9]), Mycobacterium abscessus (14.0% [95% CI 6.4–27.8]) and Mycobacterium chelonae (7.6% [95% CI 2.8–18.8]) were the most prevalent RGM among the conducted studies in Iran. Conclusions: The relatively high prevalence of RGM underlines the need for greater enforcement of infection control strategies. Establishment of appropriate diagnostic criteria and management guidelines for diseases caused by RGM and expanding the number and quality of regional reference laboratories may facilitate more accurate action for prevention and control of this kind of bacteria.

Background and objective: During recent years, the World Health Organization (WHO) proposed new software for improving the tuberculosis (TB) laboratory services. The protocol is known as “quality stepwise implementation tool” and is based on enforcement of quality assurance services through accreditation by the International Organization for Standardization (ISO) 15189. As a national reference TB laboratory (NRL) of Iran, the benefit and challenges of implementing this standard were analyzed. Material and Methods: The investigation was a cross-sectional study and was underlined by the committee of the Quality Management System (QMS) in the NRL. The required information was collected in two steps: first, the QC form that contains 334 quality questions (designed by WHO) was filled out. These questions determine 12 main organizational blocks of laboratory (facilities and safety1, organization and personnel2, documents and records3, management reviews4, client management and customer service5, equipment6, internal audit7, purchasing and inventory8, information management9, process control and internal / external quality management10, corrective actions occurrence11, and incident management and process improvement12). The minimum rating of laboratories is stated to be from 55% to a maximum of 95%. In the second step, using the quality stepwise implementation tool, these blocks were in four phases, and 41 steps were analyzed. The data were compared using Fisher's Exact test and Pearson Chi-Square. Results: In the period 2013–2014, the rate of OMS in three focal points (pre-analytical, analytical, and post-analytical) in the NRL was in the range of 80–100%; whereas in previous years the rate ranged from 48% to 79%. Therefore, through QMS the NRL scores had reached a higher level. Based on available results, the NRL-assigned score was a 5-star rating. Conclusions: As the internal score of NRL reaches 5, an international evaluation of accreditation scheme is sought. This is the first investigation that used the quality stepwise implementation tool in Iran.