An Overview of Genetic Information of Latent Mycobacterium tuberculosis / 결핵및호흡기질환

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.

Evaluation of hepatoprotective effect of silymarin among under treatment tuberculosis patients: a randomized clinical trial

Hepatic toxicity is the most serious adverse effect of anti tuberculosis drugs. This study was performed to evaluate efficacy of silymarin as a hepatoprotective herbal agent. In a randomized double blind clinical trial 70 new cases of pulmonary tuberculosis were divided into two groups. The intervention group was assigned to receive silymarin and the control group received placebo. Tuberculosis was treated by classic regimen consisting isoniazid, rifampin, pyrazinamide and ethambutol. No statistically significant difference was found between the two groups concerning the frequency of drug induced liver injury or mild elevation of liver enzymes. Silymarin was safe without any major side effect. Our results showed no significant hepatoprotective effect of silymarin among patients on tuberculosis treatment

Populations of latent Mycobacterium tuberculosis lack a cell wall: Isolation, visualization, and whole-genome characterization

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 [0.3-01 microm], lose their acid fastness [85-90%] and change their shape. Spoligopatterns of latent cells were identical to original H[37]R[v], with differences observed at spacers two and 14. WGS revealed only a few genetic changes relative to the already published H[37]R[v] reference genome. Among these was a large 2064-hp insertion [RvD6], which was originally detected in both H[37]R[a] and CDC1551, but not H[37]R[v]

Radiologic manifestations of pulmonary tuberculosis in patients of intensive care units

Tuberculosis [TB] is a serpent disease with various pulmonary manifestations, and timely diagnosis of the disease is paramount, since delayed treatment is associated with severe morbidity, particularly in intensive care units [ICU]. Therefore, it is imperative that intensivists understand the typical distribution, patterns, and imaging manifestations of TB. To describe different manifestations of pulmonary TB in patients in the ICU. In a retrospective study, all patients with a clinical and a laboratory-confirmed diagnosis of TB who were admitted to the ICU were entered in the study. All patients had a confirmatory laboratory diagnosis of TB including positive smears. The patterns of parenchymal lesions, involved segments and presence of cavity, bronchiectasis and bronchogenic spread of the lesions with computed tomography [CT] and chest/X-ray [CXR] were recorded and analyzed. Data of 146 patients with TB were entered in the study. The most common finding in CT was acute respiratory distress syndrome [ARDS]-like radiologic manifestations [17.1%], followed by parenchymal nodular infiltration [13.6%] and cavitation [10.9%], consolidation [10.2%], interstitial involvement [9.5%], calcified parenchymal mass [8.3%], ground-glass opacities [7.5%], and pleural effusion or thickening [6.9%]. Radiologic evidence of lymphadenopathy was seen in up to 43% of adults. Miliary TB was observed in 2.3% of patients, mostly in those older than 60 years of age. ARDS-like [64.5%] manifestations on CT and miliary TB [85.5%] had the highest mortality rates among other pulmonary manifestations. ARDS, interstitial involvement, and Parenchymal nodular infiltration are the most common manifestations of pulmonary TB. Various features of TB in ICU patients could be misleading for intensivists

Comparison of Rifampicin Resistance in Mycobacterium tuberculosis Isolates by Multiplex Allele Specific PCR (MAS-PCR) with Enzyme Linked Immunosorbent Assay (PCR-ELISA).

Aims: Detection of drug resistance M. tuberculosis isolates is one of the most important strategies to control the disease. Nowadays, with advances in molecular technology, various methods are available to detect drug resistant M. tuberculosis strains such as those based on capture specific probes. In this study, we aimed to investigate the frequency of mutation in the M. tuberculosis -rpoB gene by Polymerase Chain Reaction based on Enzyme Linked Immuno Sorbent Assay (PCR-ELISA) detection. Methodology: Thirty-three culture positive isolates were randomly selected for this study. All the isolates were subjected to a drug Susceptibility Test (DST) using the proportion method. Then the ability and the efficiency of Multiplex Allele Specific PCR (MAS-PCR) and PCR-ELISA to detect Rif resistant (Rifr) M. tuberculosis isolates was compared and evaluated. Results: Mutation of rpoB gene was detected in 19/33 isolates (57.6%) by PCR-ELISA. Hybridization with the specific mutant probes 516 and 526 codon occurred in 1/33 isolates each (3% respectively). Hybridization with the specific mutant probe 531 occurred in 13/33 isolates (39.4%). Three isolates (9.2%) showed simultaneous mutation in codons 516 and 531. The sensitivity and specificity of MAS-PCR in comparison to the Proportional Method was 100%. On the other hand, PCR-ELISA showed 75% sensitivity and 69.2% specificity. The positive predictive value for the PCR-ELISA method was 78.9% and the negative predictive value was 64.3%.The general efficacy of test was 72.7%. Conclusion: The study showed that the sensitivity and specificity of PCR-ELISA to detect mutations in the rpoB gene in Drug Resistant strains was low. Furthermore, this proved to be a complex, time consuming and expensive test. Therefore, this test is not recommended for determining Rifampicin resistance in M. tuberculosis strains.

[Survey of serum fat-soluble vitamins, zinc, copper and selenium levels in patients with cystic fibrosis]

Cystic fibrosis [CF] is a genetically disease with different respiratory and gastrointestinal organ involvement. Because of gastrointestinal tract involvement, patients have mal-absorption for vitamins and mineral elements; for example vitamin A, D, E and trace elements such as Selenium [Se], Zinc [Zn] and Copper [Cu] have low serum level in CF patients in different studies. The goal of this study is to define relation between serum level of this elements and severity of respiratory and gastrointestinal tract involvement in these patients. This cross sectional study performed on 39 cystic fibrosis patients admitted in pediatric ward of Masih Daneshvari hospital, Tehran, Iran. Serum levels of Vitamin A, D and E, Selenium, Zinc and Copper deficiency were studied. Serum level of Vitamin A and D were lower than normal values. Also, 3 cases [7.68%] had serum level of Zinc lower than normal values. CF patients are susceptible to have deficiencies of fat soluble vitamins and trace elements such as Selenium and Zinc because of mal-absorption. It seems that a perfect diet and nutritional components are helpful in prevention of these problems

effect of Garcin[registered sign] in preventing antiTB-induced hepatitis in newly diagnosed tuberculosis patients

Adverse effects of antituberculosis agents such as hepatotoxicity may reduce treatment effectiveness, because they significantly contribute to nonadherence and eventually result in treatment failure, relapse or the emergence of drug resistance. Garlic is an ancient herbal substance, which its effectiveness on isoniazid and rifampicin-induced hepatic injury in animal models has been demonstrated [1]. In the present study a randomized, double blind, placebo-controlled, parallel group clinical trial was designed to assess the effect[s] of garlic tablets [1000 mg daily] administered for two weeks orally. Fifty eight newly diagnosed, smear positive pulmonary tuberculosis patients, with age ranges between 18-65 years old, were randomly allocated into two groups. Each patient received either garlic or placebo tablets for the first two weeks of tuberculosis treatment. Of total 58 patients, 31 received garlic tablets while 27 received placebo. No significant difference was found between the two groups regarding age, sex, nationality, smoking, underlying diseases and opium usage. During 8 weeks of anti-TB [antituberculosis] treatment, 8 [13.0%] patients developed drug-induced hepatotoxicity [DIH]. Of them, 6 [75%] occurred in the first two weeks of treatment. Fifty percent of the patients who developed DIH were in garlic group. Results indicated no significant difference between groups in developing DIH [p=1.000]. We could not show a significant role in preventing DIH by 1000 mg daily garlic administration

Rapid detection of isoniazid resistance in mycobacterium tuberculosis by a single multiplex allele-specific polymerase chain reaction assay

Global surveillance has shown that drug resistant [DR] tuberculosis [TB] is widespread. Prompt detection of Mycobacterium tuberculosis drug resistance is essential for effective control of TB. The most frequent mutations associated with Isoniazid [INH] resistance in Mycobacterium are substitutions at codons 315 in the katG gene and the mabA-inhA promoter region [15]. This survey evaluated INH resistant-associated mutations in order to determine rapid detection of TB resistance. Through a descriptive cross-sectional study total of 96 sputum specimens were digested, examined microscopically for acid-fast bacilli and inoculated into L wenstein-Jensen slants. Thereafter, the susceptibility and identification tests were performed on culture positive specimens. Subsequently, the strains were subjected to multiplex allele-specific polymerase chain reaction [MAS-PCR] targeting in the codons 315 in the katG gene and the mabA-inhA promoter region. Distinct PCR banding patterns were observed for different mutation profiles. Drug susceptibility testing revealed that out of 96 available isolates, 30 [31.3%] were susceptible, 36 [37.5%] had multi-drug resistance [MDR-TB] and 30 [31.3%] showed mono-drug resistance. In comparison with the culture-based phenotypic drug susceptibility test, the sensitivity and specificity of MAS-PCR assay for drug resistance-related genetic mutations were 76.7% and 71.4%, respectively. The correlation between MAS-PCR and culture-based phenotypic drug susceptibility testing findings was 99. 4%. The profile of the isolates suggests a significant number of different DR strains with a high frequency of mutations at codon 315 of the katG gene. MAS-PCR provides a rapid, simple and cost-effective method for detecting MDR-TB

Clinical and demographic characteristics of confirmed cases in H1N1 [2009] influenza

Presentation of pandemic H1N1 influenza [H1N1] is widely evolving as it continues to involve different geographic locations and populations. This study was conducted to improve the precision of clinical diagnosis of H1N1 [2009] influenza infection in an outpatient setting. A prospective cross-sectional study was conducted among adult patients [age >15 years] with influenza-like illnesses [ILI] from November 2009 to February 2010. Clinical, laboratory and epidemiological findings in the first week of illness were collected using a standardized datasheet. Influenza testing was performed by real-time reverse-transcriptase polymerase chain reaction [rRT-PCR]. Thirty nine [24%] patients were positive for H1N1 and 123 [76%] were negative for any subtype of influenza A virus. Whilst otalgia [14% vs. 0 p= 0.01] was more prevalent in non-influenza A cases, cough [90% vs. 72% p = 0.03] and shortness of breath [67% vs. 47% p = 0.02] were more often associated with H1N1-infection. Comparative analysis of coexisting conditions and demographic factors of patients revealed no other significant differences between the two groups. The clinical presentation of H1N1 [2009] infection is largely indistinguishable from other acute respiratory diseases. Although previous studies suggested significant differences in demographic and co-existing conditions of H1N1 infected patients, our study shows that as the pandemic spreads worldwide and affects the majority of the population, H1N1 diagnosis based on clinical presentation and demographic characteristics has become less practical and much more difficult in tertiary care centers

Primary immunodeficiency in children: report of seven years study

Primary immunodeficiency diseases [PIDs] are a genetically heterogeneous group of disorders that affect distinct components of the innate and adaptive immune system, such as neutrophils, macrophages, dendritic cells, complement proteins, natural killer cells, T and B lymphocytes. These disorders are rare, with an estimated prevalence of 1:10,000 live births. This study aimed at describing the clinical features, disease complications, treatment modalities and overall outcome of patients with Primary Immunodeficiency Diseases [PID] in Masih Daneshvari hospital during a 7-year period [2001-2008]. This was a retrospective study based on the review of patients' medical records. Clinical, laboratory, and epidemiological data including personal and family history were obtained by reviewing records of patients admitted to the Pediatric Pulmonary Ward of NRITLD, a referral center for tuberculosis and lung diseases. The diagnosis was made based on WHO criteria for primary immunodeficiency disorders. Data collected from 59 patients were evaluated and analyzed. There were 35 [59.3%] males and 24 [40.69%] females. The age of patients ranged from 6 months to 14.5 years and the mean age was 7.4 years. Positive family history was detected in 20 [33.9%] cases and parents of 36 patients [61.2%] were consanguineous. Twenty patients [33.9%] had a family history of PID. Phagocytic disorder [57.2%] was the most common form of PID, followed by antibody deficiency [33.7%] and T-cell or combined deficiency [8.2%]. No case of complement deficiency was detected. In this group of under study patients, 2 cases expired as the result of respiratory failure due to drug resistant pneumonia [chronic granulomatous disease cases]. Based on studied results, Phagocytic disorders [57.2%] were the most common disorders among our PID patients. This may be due to the large number of CGD patients referred with the pathologic finding of granuloma misdiagnosed with tuberculosis. Considering the high prevalence of PID in this study, cases with unusual, chronic, severe or recurrent infections should be evaluated for immunodeficiency disorders

Prevalence of oseltamivir-resistant 2009 H1N1 influenza virus among patients with pandemic 2009 H1N1 influenza infection in NRITLD, Tehran, Iran

Oseltamivir-resistant cases were reported during the 2009 pandemic influenza outbreak and therefore, widespread emergence of oseltamivir-resistant 2009 H1N1 virus is imaginable. Underlying medical conditions like immunosuppression increase the chance of oseltamivir resistance. In a retrospective cross-sectional study, respiratory tract specimens of confirmed cases of 2009 H1N1 influenza referred to the Masih Daneshvari Hospital were analyzed for presence of H275Y mutation. From November 2009 through March 2010, oseltamivir-resistant 2009 H1N1 infection was observed and confirmed in 4 patients [including 2 immunocompromised patients] by performing H275Y mutation molecular testing. Close monitoring of resistance to neuraminidase inhibitors is essential in tertiary care centers. The H275Y mutation [oseltamivir-resistant genotype] could appear in the absence or presence of selective drug pressure

[Occurrence pulmonary tuberculosis in a patient with Bernard soulier syndrome: a case report]

Bernard soulier syndrome [BSS] is a rare congenital bleeding disorder characterized by thrombocytopenia. BSS causes bruising, epistaxis, gingival bleeding, menorrhagia, post partum bleeding, gastrointestinal bleeding and post traumatic hemorrhage, but there is no report of hemoptysis in BSS. A 14 year-old girl was referred to our center due to massive hemoptysis. Her chest x-ray revealed complete collapse of the right lung. Rigid bronchoscopy was preformed and the intrabronchial clots were removed. Smear and culture of direct sputum was positive for Mycobacterium tuberculosis. She received anti-tuberculosis treatment. During treatment, she developed a massive vaginal bleeding, because of drug interaction between rifampin and low dose contraceptive [LD], which she had been taking for control of massive menstrual bleeding. Her vaginal bleeding was controlled by platelet infusion and recombinant factor 7 infusion. After 2 months of anti-TB treatment, sputum smear and culture for BK converted to negative. One year after treatment, the pulmonary tuberculosis was completely cured and no hemoptysis occurred. When hemoptysis occurs in patients with Bernard soulier syndrome, we should consider other differential diagnoses, and further diagnostic evaluation is recommended

First insight into the drug resistance pattern of mycobacterium tuberculosis in Dohuk, Iraq: using spoligotyping and MIRU-VNTR to characterize multidrug resistant strains

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 L?wenstein-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

Comparison of polymerase chain reaction single-strand conformation polymorphism with DNA sequencing to detect drug resistance of mycobacterium tuberculosis isolates

Advancements in molecular technology increased our understanding of genetic mechanism of drug resistance. Nowadays, the chance of rapid detection of resistant Mycobacterium tuberculosis [M. tuberculosis] strains is increased. In the present study, we aimed to investigate the sensitivity and specificity of PCR-SSCP for detecting susceptible and resistant strains of M. tuberculosis compared with DNA sequencing. To calculate the sensitivity and specificity of PCR-SSCP assay to detect drug resistance in M. tuberculosis, respiratory samples were collected from suspected patients referred to Mycobacteriology Research Center [Masieh Daneshvary Hosptial] since 2002. Susceptibility testing against first line drugs was performed on 74 culturepositive specimens. Consequently, PCR-SSCP and DNA sequencing were performed on katG, inhA, ahpC and rpoB genes. Drug-susceptibility testing by the proportional method in selected samples revealed 16 MDR [21.6%], 23 mono-drug resistant [31%] and 35 susceptible strains [47.3%]. In comparison with DNA sequencing as a gold standard for molecular methods, the sensitivity of PCR-SSCP assay for detecting of mutation in 315 codon of katG gene was 94.74% [CI=73.97%-99.87%] with 100% [CI=93.51%-100%] specificity. In contrast, the sensitivity and specificity of this assay in detecting of rpoB gene were 70.8% [CI=48.91%-87.38%] and 88% [CI=75.69%-95.47%], respectively. PCR-SSCP in combination with DNA sequencing can be used as screening method to detect MDR-TB and mono-drug resistant cases

Shape variation in Mycobacterium tuberculosis

Over several decades, morphological variation of Mycobacterium tuberculosis [M. tuberculosis] has engaged the attention of numerous investigators. The single point on which all investigators have agreed is that tubercle bacillus does not always manifest itself in the classical rod shape. While most commonly the organism appears as a granular rod, the other forms i.e., coccid, filament and club shapes are also present. Aside from the more purely academic aspect of the subject, the possible significance of variant forms in the etiology, prognosis, and control of tuberculosis infection were objects of heated controversies, even before 1900. These differences have never been resolved, and have been ignored by most recent workers. The main questions were centered on the following points: [1] Dose the tubercle bacillus produce endospore? [2] Does it normally undergo a complicated life cycle? [3] What is the importance of the non-acid-fast forms? [4] And what happens to the bacteria during latent infection? Today, based on various in-vitro and in-vivo models, the researchers agreed to consider M. tuberculosis as a two-phase microorganism which can appear either in its metabolically active acid-fast or in its inactive forms. It is the purpose of this chapter to review and discuss morphological variation and its challenges in M. tuberculosis. Furthermore, the cell shape and cell division were illustrated using atomic force microscopy. The present information will discuss the adaptation mechanism in M. tuberculosis and may help scientists to identify targets for novel therapies

Synchronous comparison of mycobacterium tuberculosis epidemiology strains by "MIRU-VNTR" and "MIRU-VNTR and spoligotyping" technique

Molecular epidemiology analyses are frequently used in determining epidemiology of tuberculosis. Recently, Mycobacterial Interspersed Repetitive Unit Variable Number Tandem Repeat [MIRU-VNTR] and Spoligotyping has become an important method, as it allows high-through put, discriminatory and reproducible analysis of clinical isolate. The purpose of this study is to compare techniques of "MIRU-VNTR" versus "MIRU-VNTR and Spoligotyping" together for study of genetic pattern of Mycobacterium tuberculosis [M. tuberculosis] strains. Sixty M. tuberculosis [MTB] isolates were selected [30 susceptible, 30 multi-drug resistant] for this study. Thereafter, the "MIRU-VNTR and spoligotyping" were performed to identify their genetic patterns. The frequency of unknown genetic pattern of MTB was compared using technique of "MIRU-VNTR" alone versus "MIRU-VNTR and Spoligotyping" together. The MIRU-VNTR allelic diversity at each of the loci was calculated by Hunter - Gaston Discriminatory Index [HGDI]. Based on differentiation index of all strains 10, 16, 26, 31 and 40 loci were identified as the most distinctive [HGI >/= 0.6] and 2, 4, 20 and 24 as the weakest distinctive locus [HGI