Beneficial effect of long-chain n-3 polyunsaturated fatty acid supplementation on tuberculosis in mice.

Intakes of the omega-3 essential fatty acids (n-3 EFAs) are low in the general adult population, with high n-6/n-3 polyunsaturated fatty acid (PUFA) ratios and the accompanying suboptimal n-3 PUFA status. Eicosapentaenoic (EPA) and docosahexaenoic acid (DHA) have antibacterial and inflammation-resolving effects in tuberculosis (TB). However, whether switching to a diet with optimum n-3 EFA intake after the infection has comparable benefits has not been investigated. We aimed to compare the effects of a diet with sufficient n-3 EFA content in an acceptable n-6/n-3 PUFA ratio for rodents ((n-3)eFAS group) with those on the same diet supplemented with EPA and DHA (EPA/DHA group) in Mycobacterium tuberculosis (Mtb)-infected C3HeB/FeJ mice with a low n-3 PUFA status. Mice were conditioned on an n-3 PUFA-deficient diet with a high n-6/n-3 PUFA ratio for 6 weeks before Mtb infection and randomized to either (n-3)eFAS or EPA/DHA diets 1 week post-infection for 3 weeks. At endpoint, EPA and DHA compositions were higher and arachidonic acid, osbond acid, and total n-6 LCPUFAs lower in all lipid pools measured in the EPA/DHA group (all P < 0.001). Percentage body weight gain was higher (P = 0.017) and lung bacterial load lower (P < 0.001) in the EPA/DHA group. Additionally, the EPA/DHA group had a more pro-resolving lung lipid mediator profile and lower lung in IL-1α and IL-1ß concentrations (P = 0.023, P = 0.049). Inverse correlations were found between the lung and peripheral blood mononuclear cell EPA and DHA and selected pro-inflammatory cytokines. These are the first findings that indicate that EPA/DHA supplementation provides benefits superior to a diet with sufficient n-3 EFAs concerning bacterial killing, weight gain and lung inflammation resolution in Mtb-infected mice with a low n-3 PUFA status. Therefore, EPA and DHA may be worth considering as adjunct TB treatment.

A model-based analysis identifies differences in phenotypic resistance between in vitro and in vivo: implications for translational medicine within tuberculosis.

Proper characterization of drug effects on Mycobacterium tuberculosis relies on the characterization of phenotypically resistant bacteria to correctly establish exposure-response relationships. The aim of this work was to evaluate the potential difference in phenotypic resistance in in vitro compared to murine in vivo models using CFU data alone or CFU together with most probable number (MPN) data following resuscitation with culture supernatant. Predictions of in vitro and in vivo phenotypic resistance i.e. persisters, using the Multistate Tuberculosis Pharmacometric (MTP) model framework was evaluated based on bacterial cultures grown with and without drug exposure using CFU alone or CFU plus MPN data. Phenotypic resistance and total bacterial number in in vitro natural growth observations, i.e. without drug, was well predicted by the MTP model using only CFU data. Capturing the murine in vivo total bacterial number and persisters during natural growth did however require re-estimation of model parameter using both the CFU and MPN observations implying that the ratio of persisters to total bacterial burden is different in vitro compared to murine in vivo. The evaluation of the in vitro rifampicin drug effect revealed that higher resolution in the persister drug effect was seen using CFU and MPN compared to CFU alone although drug effects on the other bacterial populations were well predicted using only CFU data. The ratio of persistent bacteria to total bacteria was predicted to be different between in vitro and murine in vivo. This difference could have implications for subsequent translational efforts in tuberculosis drug development.

Study of IL-6 and vitamin D3 in patients of pulmonary tuberculosis.

BACKGROUND: Mycobacterium tuberculosis can grow in hostile intracellular environment of macrophages by actively evading macrophage-associated antibacterial activities. The stress response factor contributes this process by releasing inflammatory cytokine Interleukin 6 (IL-6). IL-6 screening of patients with TB may be useful to monitor the progress of infection and to infer the risk of progression to active disease. Vitamin D has a critical role in the innate immune system, in the circulating metabolite and supports induction of pleiotropic antimicrobial responses, through the production of antimicrobial peptides, particularly cathelicidin and its active metabolite. 1,25-dihydoxyvitamin D, has long been known to enhance immune response to mycobacteria. In this study, we have studied the role of IL-6 and Vitamin D3 in M. tuberculosis. MATERIALS AND METHODS: Three groups involved in this study are Control, Category I (newly diagnosed TB) and MDR TB patients. The serum levels of IL-6 and vitamin D3 were measured using chemiluminescence and fully-automated enzyme-linked immunosorbent assay respectively. RESULTS: The serum levels of IL-6 were significantly increased, whereas vitamin D3 decreased in TB multidrug-resistant group of patients compared to the newly diagnosed TB patients. CONCLUSION: IL-6 appears to be the major cytokine elaborated by mycobacteria infection as well as play a role in the clinical manifestations and pathological events and hence may function as a potent biomarker of tuberculosis. Since, Vitamin D increases activity of cell-mediated immunity; it can be used as a supplementation during tuberculosis therapy.

Status of vitamin D and the associated host factors in pulmonary tuberculosis patients and their household contacts: A cross sectional study.

Innate immunity plays an important role in pathophysiology of tuberculosis which is influenced by various host factors. One such factor is vitamin D which, along with its associated molecule, can alter the host defense against Mycobacterium Tuberculosis (M.Tb.) via altered production of cathelicidin and nitric oxide, both having bactericidal effect. Therefore, assessment of vitamin D and its associated molecules in tuberculosis patients and household contacts as compared to healthy controls were done and the implication of these findings in susceptibility to tuberculosis (TB) was studied. 80 active TB patients, 75 household contacts and 70 healthy controls were included. Vitamin D receptor (VDR), vitamin D binding protein (VDBP) and inducible nitric oxide synthase (iNOS) mRNA levels were studied using quantitative PCR. Serum VDR, cathelicidin, and iNOS levels were measured using ELISA. Vitamin D and NO levels were measured in serum using chemiluminescence based immunoassay and greiss reaction based colorimetry kit respectively. Decreased serum levels of vitamin D were observed in active TB patients as compared to healthy controls (p < 0.001). VDR and iNOS mRNA levels were found to be significantly lower in active TB patients compared to household contacts and healthy controls (p < 0.0001 and 0.005 respectively). VDBP mRNA expression was found to be lower in active TB group as compared to household contacts and healthy controls however the difference was not found to be significant (p > 0.21). Although, mRNA expression of VDR, VDR protein and iNOS along with vitamin D levels were significantly (p < 0.05) higher in household contacts compared to active TB group. However, levels of iNOS, NO and cathelicidin were found to be higher in TB patients as compared to household contacts and healthy controls (p < 0.01, 0.05 and 0.01 respectively). Higher levels of Vitamin D along with VDR and iNOS expression in household contacts as compared to active TB patients suggest vitamin D might have a protective role against TB plausibly decreasing disease susceptibility. Low vitamin D levels in active TB patients warrants further studies to determine the role of vitamin D supplementation in prevention and treatment of TB.

Mitogen-Induced Interferon Gamma Production in Human Whole Blood: The Effect of Heat and Cations.

BACKGROUND: Interferon-gamma release assays (IGRAs) are blood tests used to measure the amount of interferon-γ (IFN-γ) released by T lymphocytes after stimulation by antigens specific for the diagnosis of latent tuberculosis infection. A mitogen serves as a positive control to assess the immune function in IGRAs. METHODS: This in vitro study was conducted to evaluate IFN-γ production by human whole blood stimulated with heat-treated and/or cation-supplemented phytohemagglutinin (PHA), concanavalin A (Con A) and pokeweed mitogen (PWM), using QuantiFERON-TB Gold Kit ELISA tests. RESULTS: The optimal concentrations of PWM, Con A and PHA for IGRAs were 2 µg/mL, 5 µg/mL and 10 µg/mL, respectively. The results showed that IFN-γ production in response to PWM was the highest and PHA was the lowest amount. The median values of three mitogens were in the following order: PWM≥Con A≥ positive control>>PHA-P>>negative control. PWM and PHA were heat stable, while Con A was heat sensitive. The mitogen response of lymphocytes to untreated or heat-treated PWM and heat-treated Con A was increased in 1 mM Ca2+-supplemented groups, whereas the response to heat-treated PHA was decreased. Exposure to 1 mM Mg2+ had no effect on untreated or heat-treated PWM, and a concentration of 1 mM Zn2+ inhibited the stimulation of un-treated PWM. We found that calcium supplementation improved the PWM-induced production of IFN-γ. CONCLUSION: Therefore, PWM is an appropriate mitogen for use as a positive control in IGRAs. It is a potential indicator of cytokine production in the diagnostic as well as research settings, and calcium supplementation improved stimulation.

Mucosal-Associated Invariant T Cell Levels Are Reduced in the Peripheral Blood and Lungs of Children With Active Pulmonary Tuberculosis.

Mucosal associated invariant T (MAIT) cells are unconventional, semi-invariant T lymphocytes that recognize microbial-derived vitamin B2 (riboflavin) biosynthesis precursor derivatives presented by the monomorphic MHC class 1-related (MR1) molecule. Upon microbial infection, MAIT cells rapidly produce cytokines and cytotoxic effectors, and are thus important players in anti-microbial defense. MAIT cells are protective in experimental models of infection and are decreased in the blood of adult patients with bacterial infections, including Mycobacterium tuberculosis (Mtb). In children, the risk of rapid progression to active tuberculosis (TB) following Mtb infection is higher than in adults. Whether MAIT cells influence the outcome of Mtb infection in children is therefore, an important issue. We analyzed MAIT cell numbers and phenotype in 115 children investigated for pulmonary TB and determined their potential correlation with disease progression. MAIT cells were reduced in numbers and activated in the peripheral blood of children with active TB as compared to those with latent TB infection (LTBI) and healthy children. Moreover, MAIT cells did not accumulate and did not proliferate in the lung of children with active TB. These results suggest that MAIT cells may be important in preventing progression of Mtb infection to active TB in children.

Changes in serum level of trace elements in pulmonary tuberculosis patients during anti-tuberculosis treatment.

INTRODUCTIONS: Tuberculosis is spreading throughout the globe, while it is a crucial cause of death in developing countries. In this study, trace elements concentrations and their alterations were determined in TB patients during anti-tuberculosis treatment period. MATERIALS AND METHODS: We have collected blood samples from a total of 180 TB patients with pulmonary Tuberculosis, and 180 healthy controls in Sistan, Iran. The serum iron, copper, lead, calcium, arsenic and selenium concentrations were detected at the beginning of anti-TB chemotherapy, at the end of 2nd, 4th and 6th month after treatment initiation. Data were then analyzed using SPSS version 20. RESULTS AND DISCUSSIONS: Although Ca, Pb, and As levels did not change during the treatment period, serum concentrations of Fe, Zn, Cu, and Se were diminished in TB patients significantly during treatment in comparison with controls (P < 0.001).We also found that there was a significant difference in the Cu/Se and Cu/Zn ratios in tuberculosis patients in comparison with healthy individuals (P < 0.001). CONCLUSIONS: Trace elements serum concentrations are affected by TB infection and anti-TB therapy. Their serum levels were strongly perturbed during infection as well as anti-TB treatment.

Serum Zinc Concentrations in Patients with Pulmonary Tuberculosis.

Bangladesh is a tuberculosis (TB) burden country. It is one of the most important causes of mortality and morbidity and a major barrier of social and economic development. Zinc is a major trace element and an essential component of the body immune system. It's an important determinant of resistance to infection by maintaining cell mediated immunity. This analytical case control study was conducted in the Department of Internal Medicine, Bangabandhu Sheikh Mujib Medical University, Dhaka, Bangladesh to see the association of serum zinc concentrations with pulmonary tuberculosis (PTB) in adult population (18-60 years) from January 2015 to January 2016. Freshly diagnosed PTB patients before initiating anti-TB chemotherapy as cases (N=43) and TB negative subjects as controls (N=48) were included conveniently in this study with a rigid selection criteria. Serum zinc concentrations were estimated by using atomic absorption spectrophotometer. The mean±SD age and BMI of the case group and control group were 33.30±14.71 and 32.69±11.60 years, 19.88±2.31 and 22.08±2.80 kg/m2 respectively. The concentrations of serum zinc were significantly lower (P=0.01) in PTB group (840.9±230.0 µgm/l) compared with the control group (965.6±219.9 µgm/l). There was marked variation of mean±SD serum zinc concentrations between male (1008.95±246.16 µgm/l) and female (937.24±200.35 µgm/l) in control group (P=0.182) though the variation is minimal in PTB group (P=0.724). The serum zinc concentrations showed positive correlation with BMI (P=0.642) but negative correlation with age (P=0.023) in both case and control. The lower serum zinc concentrations (12.06%) in PTB patients indicate relative immune deficiency. Routine assessment of serum zinc concentration of PTB patients should be considered and further outcome should be assessed with zinc supplementation.

Higher serum 25-hydroxyvitamin D concentrations are associated with active pulmonary tuberculosis in hospitalised HIV infected patients in a low income tropical setting: a cross sectional study.

BACKGROUND: The inherent risk of developing tuberculosis (TB) in HIV- infected individuals is further enhanced by hypovitaminosis D. Interventions that offset HIV-associated immune deterioration potentially arrest disease progression and incidence of opportunistic infections including TB. Despite conflicting reports on association between vitamin D deficiency (VDD) and risk of TB, vitamin D (VD) supplementation remains a promising intervention. METHODS: We conducted a comparative cross-sectional study on 145 HIV+/pulmonary TB+ (PTB) and 139 HIV+/PTB- hospitalised patients to investigate association of vitamin D status and risk of PTB. Stratified random sampling was used to select archived serum specimens from participants enrolled in a randomised controlled trial (RCT) conducted to investigate the impact of using a point-of-care urine lipoarabinomannan strip test for TB diagnosis. PTB status was confirmed using sputum smear microscopy, culture or GeneXpert MTB/RIF. Serum 25-hydroxyvitamin D [25(OH) D] concentrations were assayed by competitive chemiluminescent immunoassay prior to commencement of anti-TB treatment. Effect of VD status on duration of hospital stay and patient outcomes on follow up at 8 weeks were also investigated. Median serum 25(OH) D concentrations were compared using Mann-Whitney test and covariates of serum VD status were assessed using logistic regression analysis. RESULTS: Overall VDD prevalence in the cohort was 40.9% (95% CI: 35.1-46.8). Median serum 25(OH)D concentrations were significantly higher in HIV+/PTB+ group (25.3 ng/ml, IQR:18.0-33.7) compared to the HIV+/PTB- group (20.4 ng/ml, IQR:14.6-26.9), p = 0.0003. Patients with serum 25(OH) D concentration ≥ 30 ng/ml were 1.9 times more likely to be PTB+ compared to those with serum 25(OH) D concentrations < 30 ng/ml (odds ratio (OR) 1.91; 95% CI 1.1-3.2). PTB-related death was associated with higher odds of having 25(OH) D levels≥30 ng/ml. Age, gender, CD4+ count, combination antiretroviral therapy (cART) status, efavirenz based cART regimen and length of hospital stay were not associated with vitamin D status. CONCLUSIONS: The finding of an association between higher serum 25(OH) D concentrations and active PTB and TB-related mortality among hospitalised HIV-infected patients in the present study is at variance with the commonly reported association of hypovitaminosis and susceptibility to TB. Our findings though, are in concordance with a small pool of reports from other settings.

Immune modulation properties of herbal plant leaves: Phyllanthus niruri aqueous extract on immune cells of tuberculosis patient - in vitro study.

Disease progression in Tuberculosis (TB) is dependent on host's immune system. Phyllanthus niruri, a traditional herb, has long been used to boost immune system in Indonesian society. This study aimed to observe the potential role of P. niruri in inducing immune cells activity in TB patients by in vitro approach. Peripheral blood mononuclear cells (PBMCs) and macrophages were collected from active pulmonary TB patients. After stimulation with graded doses of P. niruri aqueous extract, cell proliferation, phagocytic activity and nitric oxide (NO) release were analysed. P. niruri aqueous extract induced proliferation of PBMCs, increased NO release, and improved macrophages phagocytic activity. These effects were observed in a dose-dependent manner. This may lead to further research for the potential role of P. niruri as immunomodulatory adjuvant therapy for TB patients.

Trace elements in blood samples of smoker and nonsmoker active pulmonary tuberculosis patients from Jamshoro, Pakistan.

Pulmonary tuberculosis (PTB) is a serious public threat throughout the world. PTB and smoking have a strong correlation. Malnutrition, poverty, addiction, overcrowding, illiteracy, unemployment, and poor hygienic conditions are the collective aspects for the disease progress. Pakistan is the fifth among 22 high tuberculosis (TB) burden countries and the fourth regarding multidrug-resistant tuberculosis (MDR-TB). The aim of study was to determine the concentration of essential and toxic elements from blood samples of smoker and nonsmoker PTB patients by inductively coupled plasma mass spectrometry (ICP-MS) followed by microwave acid digestion and compared with control subjects (n = 30). Eighty PTB patients were selected from different hospitals with age ranging 20-70 years. It was interpreted that the mean age among males and females was found to be 35.6 ± 1.4 and 33.5 ± 1.2, respectively, and the male patients were highly affected in contrast to females. Essential elements such as Mn, Fe, Zn, and Se were statistically found to be lower while Ca, Co, and Cu were found to be higher compared to the control group (p = 0.00). However, toxic elements like Al, Cr, Ni, As, Cd, and Pb were statistically elevated in smokers than nonsmokers. Further research is needed to understand the degree of the impact of essential trace elements on treatment outcome (follow-up) followed by balanced healthy nutritional supplementation along with medical therapy, consequently improving the pulmonary tuberculosis outcome and survival as well.

Impact of Vitamin A and Carotenoids on the Risk of Tuberculosis Progression.

BACKGROUND: Low and deficient levels of vitamin A are common in low- and middle-income countries where tuberculosis burden is high. We assessed the impact of baseline levels of vitamin A and carotenoids on tuberculosis disease risk. METHODS: We conducted a case-control study nested within a longitudinal cohort of household contacts (HHCs) of pulmonary tuberculosis case patients in Lima, Peru. We defined case patients as human immunodeficiency virus (HIV)-negative HHCs with blood samples in whom tuberculosis disease developed ≥15 days after enrollment of the index patient. For each case patient, we randomly selected 4 controls from among contacts in whom tuberculosis disease did not develop, matching for sex and year of age. We used conditional logistic regression to estimate odds ratios for incident tuberculosis disease by vitamin A and carotenoids levels, controlling for other nutritional and socioeconomic factors. RESULTS: Among 6751 HIV-negative HHCs with baseline blood samples, 192 had secondary tuberculosis disease during follow-up. We analyzed 180 case patients with viable samples and 709 matched controls. After controlling for possible confounders, we found that baseline vitamin A deficiency was associated with a 10-fold increase in risk of tuberculosis disease among HHCs (adjusted odds ratio, 10.53; 95% confidence interval, 3.73-29.70; P < .001). This association was dose dependent, with stepwise increases in tuberculosis disease risk with each decreasing quartile of vitamin A level. CONCLUSIONS: Vitamin A deficiency strongly predicted the risk of incident tuberculosis disease among HHCs of patients with tuberculosis. Vitamin A supplementation among individuals at high risk of tuberculosis may provide an effective means of preventing tuberculosis disease.

Anti-inflammatory and antioxidant effect of ginger in tuberculosis.

BACKGROUND: Tuberculosis (TB) has reemerged to become the world's leading cause of death from a single infectious agent. Inflammatory cytokines play an important role during the course of the disease and may be responsible for tissue damage by lipid peroxidation. The study was aimed to explore the anti-inflammatory and antioxidant effect of ginger in pulmonary TB patients. METHODS: A total of 69 pulmonary TB patients participated in a randomized and placebo-controlled study. The intervention group received 3 g of ginger extract daily for 1 month and placebo group was supplemented with starch capsule. Participants of both groups were taking standard antitubercular treatment during the study. The concentrations of tumor necrosis factor (TNF) alpha, ferritin and malondialdehyde (MDA) in blood samples were analyzed before and after the intervention by using enzyme-linked immunosorbent assay for TNF alpha and ferritin and spectrophotometry for MDA. RESULTS: Ginger supplementation significantly reduced the levels of TNF alpha, ferritin and MDA in ginger supplemented group in comparison to baseline. Ginger supplementation with antitubercular treatment significantly lowered TNF alpha, ferritin and MDA concentrations in comparison to control group. CONCLUSIONS: Ginger was found to be effective as an anti-inflammatory and antioxidant supplement along with anti-TB therapy as it possesses strong free radical scavenging property.

Pharmacokinetics of Rifampin, Isoniazid, Pyrazinamide, and Ethambutol in Infants Dosed According to Revised WHO-Recommended Treatment Guidelines.

There are limited pharmacokinetic data for use of the first-line antituberculosis drugs during infancy (<12 months of age), when drug disposition may differ. Intensive pharmacokinetic sampling was performed in infants routinely receiving antituberculosis treatment, including rifampin, isoniazid, pyrazinamide, and ethambutol, using World Health Organization-recommended doses. Regulatory-approved single-drug formulations, including two rifampin suspensions, were used on the sampling day. Assays were conducted using liquid chromatography-mass spectrometry; pharmacokinetic parameters were generated using noncompartmental analysis. Thirty-nine infants were studied; 14 (36%) had culture-confirmed tuberculosis. Fifteen (38%) were premature (<37 weeks gestation); 5 (13%) were HIV infected. The mean corrected age and weight were 6.6 months and 6.45 kg, respectively. The mean maximum plasma concentrations (Cmax) for rifampin, isoniazid, pyrazinamide, and ethambutol were 2.9, 7.9, 41.9, and 1.3 µg/ml, respectively (current recommended adult target concentrations: 8 to 24, 3 to 6, 20 to 50, and 2 to 6 µg/ml, respectively), and the mean areas under the concentration-time curves from 0 to 8 h (AUC0-8) were 12.1, 24.7, 239.4, and 5.1 µg · h/ml, respectively. After adjusting for age and weight, rifampin exposures for the two formulations used differed inCmax(geometric mean ratio [GMR],2.55; 95% confidence interval [CI], 1.47 to 4.41;P= 0.001) and AUC0-8(GMR, 2.52; 95% CI, 1.34 to 4.73;P= 0.005). HIV status was associated with lower pyrazinamideCmax(GMR, 0.85; 95% CI, 0.75 to 0.96;P= 0.013) and AUC0-8(GMR, 0.79; 95% CI, 0.69 to 0.90;P< 0.001) values. No other important differences were observed due to age, weight, prematurity, ethnicity, or gender. In summary, isoniazid and pyrazinamide concentrations in infants compared well with proposed adult target concentrations; ethambutol concentrations were lower but similar to previously reported pediatric studies. The low rifampin exposures require further investigation. (This study has been registered at ClinicalTrials.gov under registration no. NCT01637558.).

Correlation between Resistin, Tuberculosis and Khat Addiction: A Study from South Western Province of Saudi Arabia.

Tuberculosis(TB) is a disease of global significance, which accounts for a death in every 15 seconds. Recent studies shows TB is rising in certain parts of the world, and Saudi Arabia is one of them. Several factor contribute in predisposing the subjects for infection including but not limited to addiction to various compounds which have immune modulation properties, such as amphetamines and Heroin etc. Khat a plant whose leaves are chewed for its euphoric effect in east Africa and Arabian Peninsula including Saudi Arabia, is considered as mildly addictive, and its principle compound, Cathinone shares structural and functional similarity with amphetamine a known immunomodulator. Tuberculosis being a disease of immune modulation has a varied spectrum of complex interplay of proinflammatory molecules, resistin is one of them. In the present study, we try to explore the trinity of khat addiction, serum resistin level and tuberculosis by correlating the serum resistin level in non khat addicted healthy subjects, khat addicted healthy subjects, and in patients, both khat addicted and non khat addicted, with active tuberculosis. We observed significantly higher resistin level among the apparently healthy khat addicted subjects as compared to non addicted healthy controls. Thereafter, when we compare the resistin levels between khat addicted and non khat addicted TB patients we did not found significant difference between the two groups. However bacillary load was observe to be significantly higher among the khat addicted TB patient as compare to non addicted one. Validation of above results in animal model revealed dose dependant increase in bacillary growth in the Wistar rats treated with khat. Taken together these results suggest the role of khat in immune modulation albeit in the limited frame of resistin level.

Serum protein gamma-glutamyl hydrolase, Ig gamma-3 chain C region, and haptoglobin are associated with the syndromes of pulmonary tuberculosis in traditional Chinese medicine.

BACKGROUND: Traditional Chinese Medicine (TCM) has been applied in treating tuberculosis (TB) based on the TCM syndromes with the effects of inhibiting Mycobacterium, strengthening the body immune system, and reducing the pulmonary toxicity. We used bioinformatic methods to study the clinical and pathological characteristics of pulmonary TB patients with TCM syndromes. Isobaric tags for relative and absolute quantification - coupled two dimensional liquid chromatography-tandem mass spectrometry (iTRAQ-2DLC-MS/MS) methods were applied to screen differentially expressed serum proteins. METHODS: Pulmonary TB cases were divided into four distinctive TCM syndromes: pulmonary Yin deficiency (PYD) syndrome, hyperactivity of fire due to Yin deficiency (HFYD) syndrome, deficiency of Qi and Yin (DQY) syndrome, and deficiency of Yin and Yang (DYY) syndrome. The serum samples from 214 pulmonary TB patients were collected, and the clinical and pathological data was analyzed by using iTRAQ-2DLC-MS/MS. Finally, the differentially expressed proteins were screened and tested by ELISA. Only 5 patients with DYY syndrome were recruited in 3 years, which were not enough for further research. RESULTS: The DQY cases had higher erythrocyte sedimentation rate (ESR) compared to the PYD and HFYD cases (P=0.0178). 94.44% (12 PYD, 18 HFYD, and 4 DQY before anti-TB treatment) of 36 treated TB cases were transformed to PYD accompanied with the reduction of ESR and absorption of pulmonary lesions. A total of 39 differentially expressed proteins (ratios of >1.3 or <0.75) were found among the three TCM syndromes. Proteomic studies revealed that gamma-glutamyl hydrolase (GGH), Ig gamma-3 chain C region (IGHG3), and haptoglobin (HPT) were specifically over-expressed in PYD (P<0.01), HFYD (P<0.001), and DQY cases (P<0.01), respectively. Furthermore, GGH was significantly higher in PYD cases compared to the HFYD and DQY cases (P<0.01, P<0.001, respectively), whereas IGHG3 was significantly higher in HFYD cases than PYD and DQY cases (P<0.001, P<0.01, respectively). CONCLUSIONS: The results suggest that TCM syndromes are significantly correlated with the pulmonary lesions and ESR. GGH was associated with folate metabolism in PYD cases, IGHG3 was linked to the control of Mycobacterium infection in HFYD patients, and HPT was involved in hypoxia in DQY patients. The present study provides new biological basis to understand the pathological changes and proteomic differences of TB syndromes.

Improvement in plasma drug activity during the early treatment interval among Tanzanian patients with multidrug-resistant tuberculosis.

BACKGROUND: Individual pharmacokinetic variability may be common in patients treated for multidrug-resistant tuberculosis (MDR-TB) but data are sparse from resource-limited settings and across the early treatment interval. METHODS: Plasma drug activity, as measured by the TB Drug Activity (TDA) assay at 2 and 4 weeks of treatment with a standardized MDR-TB regimen was performed in patients with pulmonary MDR-TB from Tanzania. TDA values were correlated with measures of early treatment outcome including every two week collection of sputum for time-to-positivity (TTP) in liquid culture from the MGIT 960 automated system. Patients were evaluated at 24 weeks and those surviving without delayed sputum culture conversion (>8 weeks), culture reversion after previously negative, or weight loss were defined as having a favorable outcome. RESULTS: Twenty-five patients were enrolled with a mean age of 37 ±12 years. All were culture positive from the pretreatment sputum sample with a mean TTP in MGIT of 257 ±134 hours, and the median time to culture conversion on treatment was 6 weeks. Twenty patients (80%) had an increase in TDA, with the overall mean TDA at 2 weeks of 2.1 ±0.7 compared to 2.4 ±0.8 at 4 weeks (p = 0.005). At 2 weeks 13 subjects (52%) had a TDA value > 2-log killing against their own M. tuberculosis isolate compared to 17 subjects (68%) at 4 weeks (McNemar's exact test p = 0.29). An interim treatment outcome was able to be determined in 23 patients (92%), of whom 7 had a poor outcome (30%). An increase in TDA from week 2 to week 4 was associated with favorable outcome, [unadjusted OR = 20.0, 95% CI: 1.61-247.98, exact p = 0.017 and adjusted OR = 19.33, 95% CI: 1.55-241.5, exact p = 0.023]. CONCLUSIONS: The majority of patients with MDR-TB in Tanzania had an increase in plasma drug activity from week 2 to week 4 of treatment as measured by the TDA assay. Understanding the etiology and full impact of this dynamic may inform therapeutic intervention.

Vitamin D levels in Indian children with intrathoracic tuberculosis.

BACKGROUND & OBJECTIVES: Deficiency of vitamin D, an immunomodulator agent, is associated with increased susceptibility to tuberculosis in adults, but only limited studies are available in the paediatric age group, especially regarding association of vitamin D with type and outcome of tuberculosis. We conducted this study to determine the baseline 25-hydroxy vitamin D levels in children suffering from intrathoracic tuberculosis and its association with type and outcome of tuberculosis. METHODS: Children with intrathoracic tuberculosis, diagnosed on the basis of clinico-radiological criteria, were enrolled as part of a randomized controlled trial on micronutrient supplementation in paediatric tuberculosis patients. Levels of 25-hydroxy vitamin D were measured in serum samples collected prior to starting antitubercular therapy by chemiluminescent immunoassay technology. RESULTS: Two hundred sixty six children (mean age of 106.9 ± 43.7 months; 57.1% girls) were enrolled. Chest X-ray was suggestive of primary pulmonary complex, progressive disease and pleural effusion in 81 (30.5%), 149 (56%) and 36 (13.5%) subjects, respectively. Median serum 25-hydroxy vitamin D level was 8 ng/ml (IQR 5, 12). One hundred and eighty six (69.9%) children were vitamin D deficient (serum 25-hydroxy vitamin D <12 ng/ml), 55 (20.7%) were insufficient (12 to <20 ng/ml) and 25 (9.4%) were vitamin D sufficient (≥ 20 ng/ml). Levels of 25-hydroxy vitamin D were similar in all three types of intrathoracic tuberculosis, and in microbiologically confirmed and probable cases. Levels of 25-hydroxy vitamin D did not significantly affect outcome of the disease. Children who were deficient or insufficient were less likely to convert (become smear/culture negative) at two months as compared to those who were 25-hydroxy vitamin D sufficient ( p <0.05). INTERPRETATION & CONCLUSIONS: Majority of Indian children with newly diagnosed intrathoracic tuberculosis were deficient in vitamin D. Type of disease or outcome was not affected by 25-hydroxy vitamin D levels in these children. However, children who did not demonstrate sputum conversion after intensive phase of antitubercular therapy had lower baseline 25-hydroxy vitamin D levels as compared to those who did.

Screening and identification of potential biomarkers and establishment of the diagnostic serum proteomic model for the Traditional Chinese Medicine Syndromes of tuberculosis.

ETHNOPHARMACOLOGICAL RELEVANCE: Chemotherapy is the mainstay of modern tuberculosis (TB) control. Traditional Chinese Medicine (TCM) can enhance the effect of anti-TB drug, promote the absorption of the foci in the lung and reduce drug toxicity. In TCM, the determination of treatment is based on ZHENG (also called TCM syndrome). To establish a diagnostic model by using proteomics technology in order to identify potential biomarkers for TCM syndromes of TB. MATERIALS AND METHODS: The surface-enhanced laser desorption ionization time of flight mass spectrometer (SELDI-TOF MS) combined with weak cation exchange (WCX) magnetic beads was used to screen serum samples from 71 cases of deficiency of lung yin syndrome (DLYS), 64 cases of fire (yang) excess yin deficiency syndrome (FEYDS) and 45 cases of deficiency of both qi and yin syndrome (DQYS). A classification model was established by Biomarker Pattern Software (BPS). Candidate protein biomarkers were purified by reverse phase-high performance liquid chromatograph (RP-HPLC), identified by MALDI-TOF MS, LC-MS/MS and validated by ProteinChip Immunoassays. RESULTS: A total of 74 discriminating m/z peaks (P<0.001) among three TCM syndromes of TB were detected. A diagnostic model for the TCM syndrome of TB based on the five biomarkers (3961.7, 4679.7, 5646.4, 8891.2 and 9416.7 m/z) was established which could discriminate DLYS, FEYDS and DQYS patients with an accuracy of 74.0%, 72.5%, and 96.7%, respectively. The candidate biomarker with m/z of 9416.7 was identified as a fragment of apolipoprotein C-III (apoC-III) by MALDI-TOF-MS and LC-MS/MS. CONCLUSION: The TCM syndrome diagnostic model of TB could successfully distinguish the three TCM syndromes of TB patients. This provided a biological basis for the determination of treatment based on different TCM syndromes of TB. ApoC-III was identified as a potential biomarker for TCM syndromes of TB and revealed the biochemical basis and pathogenesis of TCM syndromes in TB.

Association between serum selenium level and conversion of bacteriological tests during antituberculosis treatment.

OBJECTIVE: To determine whether serum selenium levels are associated with the conversion of bacteriological tests in patients diagnosed with active pulmonary tuberculosis after eight weeks of standard treatment. METHODS: We evaluated 35 healthy male controls and 35 male patients with pulmonary tuberculosis, the latter being evaluated at baseline, as well as at 30 and 60 days of antituberculosis treatment. For all participants, we measured anthropometric indices, as well as determining serum levels of albumin, C-reactive protein (CRP) and selenium. Because there are no reference values for the Brazilian population, we used the median of the serum selenium level of the controls as the cut-off point. At 30 and 60 days of antituberculosis treatment, we repeated the biochemical tests, as well as collecting sputum for smear microscopy and culture from the patients. RESULTS: The mean age of the patients was 38.4 ± 11.4 years. Of the 35 patients, 25 (71%) described themselves as alcoholic; 20 (57.0%) were smokers; and 21 (60.0%) and 32 (91.4%) presented with muscle mass depletion as determined by measuring the triceps skinfold thickness and arm muscle area, respectively. Of 24 patients, 12 (39.2%) were classified as moderately or severely emaciated, and 15 (62.5%) had lost > 10% of their body weight by six months before diagnosis. At baseline, the tuberculosis group had lower serum selenium levels than did the control group. The conversion of bacteriological tests was associated with the CRP/albumin ratio and serum selenium levels 60 days after treatment initiation. CONCLUSIONS: Higher serum selenium levels after 60 days of treatment were associated with the conversion of bacteriological tests in pulmonary tuberculosis patients.