Misdiagnosis of tuberculosis and the clinical relevance of non—tuberculous mycobacteria in Zambia

ObjectiveTo determine the accuracy of TB diagnosis of TB in Zambia in the era of increasing HIV prevalence.MethodsSputum of the clinically diagnosed TB cases was additionally subjected to liquid culture and molecular identification. This study distinguished between TB cases confirmed by positive Mycobacterium tuberculosis (M. tuberculosis) cultures and mycobacterial disease caused by non-tuberculous mycobacteria (NTM).ResultsOnly 49% of the 173 presumptively diagnosed TB cases was M. tuberculosis cultured, while in 13% (22) cases, a combination of M. tuberculosis and NTM was found. In 18% of the patients only NTM were cultured. In 28%, no mycobacteria was cultivable. HIV positive status was correlated with the isolation of NTM (P <0.05).ConclusionsThe diagnosis of tuberculosis based on symptoms, sputum smear and/or chest X-ray leads to significant numbers of false-positive TB cases in Zambia, most likely due to the increased prevalence of HIV. The role of NTM in tuberculosis-like disease also seems relevant to the false diagnosis of TB in Zambia.     

Interleukin-6 and Mycobacterium tuberculosis dormancy antigens improve diagnosis of tuberculosis

ObjectivesIFNγ-release assays (IGRAs) used for diagnosis of Mycobacterium (M.) tuberculosis infection have limited sensitivity. Alternative cytokines and M. tuberculosis latency-associated antigens may improve immune-based tests.MethodsMultiplex cytokine analyses was done in culture supernatants after 6-day in vitro restimulation with M. tuberculosis IGRA and latency-associated antigens (i.e. Rv2628, Rv1733) in tuberculosis patients (n = 22) and asymptomatic contacts (AC)s (n = 20) from Ghana.ResultsFour cytokines (i.e. IFNγ, IP-10, IL-22 and IL-6) were significantly increased after IGRA-antigen specific restimulation. IFNγ, IP-10, and IL-22 correlated positively and showed no differences between the study groups whereas IGRA-antigen induced IL-6 was significantly higher in tuberculosis patients. Using adjusted IGRA criteria, IL-6 showed the highest sensitivity for detection of tuberculosis patients (91%) and ACs (85%) as compared to IFNγ, IP-10, and IL-22. Rv2628 and Rv1733 restimulation induced significantly higher IFNγ, IP-10, and IL-22 concentrations in ACs. Combined antigen/cytokine analyses identified study group specific patterns and a combination of Rv2628/Rv1733 induced IFNγ with IGRA-antigen induced IL-6 was optimal for classification of tuberculosis patients and ACs (AUC: 0.92, p<0.0001).ConclusionsWe demonstrate the potency of alternative cytokines, especially IL-6, and latency-associated antigens Rv1733/Rv2628 to improve detection of M. tuberculosis infection and to classify tuberculosis patients and healthy contacts.     

Non tuberculosis mycobacteria isolates among new and previously treated pulmonary tuberculosis patients in Nigeria

ObjectiveTo determine the prevalence of non tuberculosis mycobacteria (NTM) among new and previously treated tuberculosis (TB) patients in Nigeria.MethodsIt was a retrospective study. A total of 102 sputum smear positive samples/culture isolates from pulmonary TB patients (41 new smear positive and 61 smear positive retreatment cases) were sent to the Institute of Tropical Medicine, Antwerp Belgium between 2007-2009. Data on patients' characteristics were retrieved from their treatment cards.ResultsAmong the 102 samples, 25 isolates results (20 were culture negative while 5 were contaminated) were excluded from the study. Data were available for 77 mycobacterium isolates. 70 (90.9%) were identified as Mycobacterium tuberculosis and 7 (9.1%) as atypical mycobacteria. Among the atypical mycobacteria, three of them were Mycobacterium fortuitum, two Mycobacterium intracellulare and two Mycobacterium chelonae. Of the seven isolates with atypical mycobacteria, 4 (57.1%) were from previously treated patients, while 3 (42.9%) were new sputum positive patients. There was no statistically significant difference in NTM infection between new and previously treated pulmonary TB patients (P =0.97).ConclusionsThe study shows the involvement of atypical mycobacterium in pulmonary infection in both new and previously treated TB patients. Therefore, there is a need to carry out culture and drug susceptibility testing in all pulmonary TB patients especially those who had failed conventional DOTS treatment to rule out NTM infections.     

The relationship between 25-hydroxyvitamin D levels and treatment course of pulmonary tuberculosis

BackgroundLow serum vitamin D level is associated with a high risk of developing active tuberculosis (TB). We investigated the relationships between serum vitamin D levels and clinical course of TB after standard chemotherapy in hospitalized non-HIV patients with TB.MethodsHospitalized patients with TB were recruited between February 2008 and July 2010. Confirmatory tests were performed using sputum smear and culture positivity tests for Mycobacterium tuberculosis. Drug sensitivity testing was performed for all the subjects and those not showing drug resistances for the first-line anti-TB drugs were included in the study. These patients were treated with the standard first-line anti-TB drugs. Serum 25-hydroxyvitamin D [25(OH)D] levels were measured on admission, and the relationships between 25(OH)D and clinical characteristics (laboratory data on admission and treatment outcomes) were examined. We defined vitamin D deficiency as a condition where serum level of 25(OH)D was lower than 20 ng/ml.ResultsA total of 38 patients were included in the study. Mean (±SD) 25(OH)D levels were 13.7±5.9 ng/ml. The prevalence of vitamin D deficiency was 87%. In 23 patients treated with the standard first-line 4-drug regimen (Age <80 years) serum 25(OH)D levels showed significant negative correlation with time taken to obtain 3 consecutive negative sputum smears or TB bacteria cultures. This relationship suggests that low serum vitamin D level may not only increase the risk of developing active TB but may also be related to the poor treatment outcomes in these patients.ConclusionsLow serum vitamin D level is a good predictor of prolonged clinical course in patients with active pulmonary TB.     

Impact of vitamin D in the treatment of tuberculosis

Tuberculosis (TB) is a major global health problem and often coincides with nutritional deficiency. In fact, vitamin D deficiency has been reported among TB patients, and vitamin D receptor polymorphisms are associated with susceptibility to Mycobacterium tuberculosis. High doses of vitamin D were widely used to treat TB patients in the preantibiotic era. This approach was successful: vitamin D can suppress intracellular growth of M tuberculosis in vitro. Vitamin D also induced the expression of cathelicidin, which is involved in the first line of defense in TB patients. Thus, vitamin D may have a role in TB treatment, and further investigation is needed.     

Evaluation of cytokine levels using QuantiFERON-TB Gold Plus in patients with active tuberculosis

ObjectivesA recently released new QuantiFERON (QFT) product, QFT TB Gold plus (QFT-plus), is optimized for both CD4 and CD8 responses and reported to have higher sensitivity compared to the former QFT-3 G. Previously, using supernatants of QFT-3 G, we and others have demonstrated that cytokines other than IFN-γ may be useful in diagnosing tuberculosis. The present study aimed to identify cytokines that are useful for accurately diagnosing active tuberculosis by using QFT-plus and compared the data to those with QFT-3 G.MethodsEighty-three active tuberculosis patients and 70 healthy control subjects who were examined by QFT at Tokyo National Hospital from June 2017 to July 2018 were enrolled. QFT-3 G and QFT-plus were performed according to the manufacturer's instructions. At the same time, blood cell culture supernatants were collected and assayed for their cytokine levels using R&D Systems Luminex Assay and MAGPIX System. The levels of cytokines were compared between different antigen-containing tubes (3 G Ag, TB1 and TB2 tubes), as well as between the patients and the control subjects. ROC curves were drawn, and the AUCs were calculated.ResultsFive cytokines, i.e., IL-2, IL-6, IL-8, IP-10 and MIP-1β, produced by human blood cells in three independent tubes containing different tuberculosis antigens were higher in the 3 G Ag tube compared to both the TB1 and TB2 tubes. Further, when the TB1 and TB2 tubes were compared, TB2 showed greater production of only PDGF-BB, and less production of IL-6 and TNF-α. For diagnosing active tuberculosis, the levels of IP-10 were superior to the level of IFN-γ based on showing a larger AUC for ROC curves in our present study setting. Finally, the levels of IFN-γ, IL-1RA, IL-2, IP-10, MCP-1 and MIP-1β were distinctly different between the active tuberculosis patients and healthy controls.ConclusionsIn summary, there was no cytokine that was higher in the tubes of QFT-plus compared to the tube of QFT-3 G, suggesting inferiority of QFT-plus antigens to 3 G Ag in terms of elicitation of cytokine production. Our results also suggest the usefulness of cytokines that showed a significant difference between the active tuberculosis patients and the healthy controls—namely, IFN-γ, IL-1RA, IL-2, IP-10, MCP-1 and MIP-1β—for diagnosing tuberculosis, but the roles of these cytokines in the pathogenesis of tuberculosis need to be elucidated (UMIN000035253).     

Drug resistance pattern of Mycobacterium tuberculosis isolates from patients of five provinces of Iran

Objective:To determine the patterns of resistance to first line anti-tuberculosis(TB)drugs among a collection of Mycobacterium tuberculosis(MTB)isolates from 5 provinces of Iran.Methods:A total of the 6 426 clinical specimens from patients suspected of active TB were collected from March 2010 to June 2012.All specimens were subjected for microscopy and culture tests in the TB centers of studies provinces.Drug susceptibility testing to the first line anti-TB drugs for culture positive MTB was performed on Lwenstein-Jensen(LJ)medium using proportion method.Results:Of 6 426 clinical specimens,261 were culture positive for mycobacteria,of which 252 were MTB and 9 were MOTT(mycobacteria other than tuberculosis).Of 252 MTB isolates.211(83.7%)were pan-susceptible and 41(16.3%)were resistant to at least one drug.Resistance was most common to streptomycin.30 isolates(12.0%),followed by isoniuzid,20isolates(8.0%),rifampin,15 isolates(6.0%)and ethambutol,14 isolates(5.5%).Sixteen(6.3%)MTB isolates were MDR.A clear evidence of heterogeneity amongst the 5 provinces in the proportions with resistance to one or more drugs was observed[χ~2=12.209(4 degrees of freedom),P values=0.015 9].Conclusions:The prevalence of drug resistance in this study area underscoring the need for further enforcement of TB control strategies in the Iran.Drug susceptibility testing for all TB cases to provide optimal treatment,establishing advanced diagnostic facilities for rapid detection of MDR-TB and continuous monitoring of drug resistance are recommended for prevention and control of drug-resistant TB.     

The antioxidant status and lipid peroxidation product of newly diagnosed and 6 weeks follow–up patients with pulmonary tuberculosis in Owerri,Imo state,Nigeria

ObjectiveTo determine the levels of antioxidants vitamins C and E as well as lipid peroxidation product malondialdehyde (MDA) in Mycobacterium tuberculosis (M. tuberculosis) patients.MethodsSixty two M. tuberculosis positive patients and fifty five healthy controls within the age of twenty five to forty years without any systemic disease attending General Hospital Owerri were involved in this study. Forty one cases were longitudinally followed up with standard anti-tuberculosis chemotherapy (ATT) for six weeks.ResultsThe results obtained showed that the levels of vitamin C (0.91±0.42 mg/dL) and vitamin E (0.84±0.31 mg/dL) were significantly decreased in M. tuberculosis patients before treatment when compared to the healthy controls [(1.64±0.41 mg/dL) and (1.46±0.38 mg/dL)] respectively at P<0.05, while the level of MDA (8.7±1.81 nM/mL) in M. tuberculosis patients was significantly higher (P<0.05) before anti-tuberculosis treatment as compared with the healthy control (4.91±1.9 nM/mL). Also, there was a significant increase in vitamin C and E levels after 6 weeks of ATT, while MDA levels was decreased when compared with the control (P<0.05).ConclusionsThe depletion of vitamins C and E as well as elevation of MDA in tuberculosis patients is suggestive of lipid peroxidation and oxidative stress. The increase in vitamin C and E as well as decrease in MDA after 6 weeks of anti-tuberculosis treatment is suggestive of good response to treatment with standard ATT. Hence, vitamin C and E supplementation improves the quality of life of tuberculosis patients.     

A single dose of vitamin D enhances immunity to mycobacteria

RATIONALE: Vitamin D was used to treat tuberculosis (TB) in the preantibiotic era. Prospective studies to evaluate the effect of vitamin D supplementation on antimycobacterial immunity have not previously been performed. OBJECTIVES: To determine the effect of vitamin D supplementation on antimycobacterial immunity and vitamin D status. METHODS: A double-blind randomized controlled trial was conducted in 192 healthy adult TB contacts in London, United Kingdom. Participants were randomized to receive a single oral dose of 2.5 mg vitamin D or placebo and followed up at 6 weeks. MEASUREMENTS AND MAIN RESULTS: The primary outcome measure was assessed with a functional whole blood assay (BCG-lux assay), which measures the ability of whole blood to restrict luminescence, and thus growth, of recombinant reporter mycobacteria in vitro; the readout is expressed as a luminescence ratio (luminescence postinfection/baseline luminescence). IFN-gamma responses to the Mycobacterium tuberculosis antigens early secretory antigenic target-6 and culture filtrate protein 10 were determined with a second whole blood assay. Vitamin D supplementation significantly enhanced the ability of participants' whole blood to restrict BCG-lux luminescence in vitro compared with placebo (mean luminescence ratio at follow-up, 0.57, vs. 0.71, respectively; 95% confidence interval for difference, 0.01-0.25; p=0.03) but did not affect antigen-stimulated IFN-gamma secretion. CONCLUSIONS: A single oral dose of 2.5 mg vitamin D significantly enhanced the ability of participants' whole blood to restrict BCG-lux luminescence in vitro without affecting antigen-stimulated IFN-gamma responses. Clinical trials should be performed to determine whether vitamin D supplementation prevents reactivation of latent TB infection. Clinical trial registered with www.clinicaltrials.gov (NCT 00157066).     

Efficacy and safety of cholecalciferol-augmented anti-tuberculosis therapy for treatment of naïve patients with pulmonary tuberculosis: A randomized,controlled, clinical study

Background/objectiveVitamin D deficiency may contribute to the therapeutic failure of antituberculosis therapy (ATT). The aim of this study is to explore the role of adding cholecalciferol to the standard ATT in improving the therapeutic outcome among the naïve patients with pulmonary tuberculosis (TB).MethodsA randomized, controlled, clinical study, which included 496 naïve patients with pulmonary TB, was carried out. The patients were randomly allocated to two groups. Group-A included 247 patients who received ATT, while group-B included 249 patients who received ATT with cholecalciferol.ResultsThe rate of therapeutic failure among the study population was 29.4%; it was significantly lower among patients of group-B compared to those of group-A (22.1% (95% CI 14.7–26.2) vs 38.1% (95% CI 31.5–46.1), p 0.036). In addition, the rate of early therapeutic response was significantly higher among patients of group-B compared to those of group-A (35.3% (95% CI 29.6–42.3) vs 19.4% (95% CI 15.1–24.6), p 0.041). Incidence rate of adverse effects was 19.3%; it was higher (although not statistically significant) among patients of group-A compared to those of group-B (21.9% vs 16.9%).ConclusionsIn conclusion, cholecalciferol-augmented ATT can be more efficacious in treating naïve patients with pulmonary TB compared to the standard ATT. In addition, adding vitamin D3 to ATT provides extra protection against the hepatic and muscular adverse effects of ATT.     

Th2 immune response by the iron-regulated protein HupB of Mycobacterium tuberculosis

BackgroundHupB is an iron-regulated protein essential for the growth of Mycobacterium tuberculosis inside macrophages. To investigate if HupB induced a dominant Th2 type immune response, we studied the effect of rHupB on PBMCs from TB patients and by infecting mouse macrophages with wild type and hupB KO mutants.MethodsPBMCs from pulmonary TB (n = 60), extra pulmonary TB (n = 23) and healthy controls (n = 30) were stimulated with purified HupB and the cytokines secreted were assayed. The sera were screened for anti-HupB antibodies by ELISA. Mouse macrophages cell line (RAW 264.7) was infected with wild type, hupB KO and hupB-complemented strains of M. tuberculosis grown in high and low iron medium and the expression of cytokines was assayed by qRT-PCR.ResultsMurine macrophages infected with the hupB KO strain produced low levels of the pro-inflammatory cytokines IFN-γ, TNF-α, IL-1, and IL-18 and high levels of IL-10. HupB induced IL-6 and IL-10 production in PBMCs of TB patients and down-regulated IFN-γ and TNF-α production. The influence of HupB was remarkable in the EPTB group.ConclusionHupB shifted the immune response to the Th2 type. Low IFN-γ and elevated IL-10 in EPTB patients is noteworthy.     

Effects of vitamin D supplementation on circulatory YKL-40 and MCP-1 biomarkers associated with vascular diabetic complications: A randomized,placebo-controlled,double-blind clinical trial

AimDiabetic patients predispose to vascular diseases such as nephropathy, and retinopathy. Poor adherence to medical treatment and dietary recommendations in uncontrolled diabetes leads to vascular damages. Vitamin D has been extensively studied and found to be protective against diabetes mellitus. YKL-40 and Monocyte chemoattractant protein-1 (MCP-1) are considered to exert crucial role in diabetes and its complications. Therefore, this study was designed to investigate effects of vitamin D supplementation on serum levels of YKL-40 and MCP-1 involved in the development of diabetic complications.MethodsFor 12 weeks, 48 type 2 diabetic patients enrolled in the trial and randomly were divided into two groups (n = 24 per group), receiving one of the following: 100 μg (4000 IU) vitamin D or placebo. Before and after intervention, serumYKL-40, MCP-1, insulin, IL-6, TNF-α, 25- (OH) vitamin D and HbA1c were measured.ResultsOur results revealed that serum levels of 25 (OH) vitamin D significantly increased in vitamin D group (p < 0.001). Vitamin D supplementation also significantly reduced serum YKL-40 levels (−22.7 vs. −2.4 ng/ml; (p-value = 0.003)). There was a significant decline in MCP-1 concentration in intervention group at the end of the study (−45.7 vs. −0.9 pg/ml; (p = 0.001)). Furthermore, there was a significant decrease in IL-6, fasting insulin and HOMA-IR in intervention group after 3 months supplementation.ConclusionsDaily vitamin D supplementation effectively reduced circulatory YKL-40 and MCP-1 levels in patients with type-2 diabetes and vitamin D deficiency. Vitamin D might contribute in reducing diabetic complications via modulating YKL-40 and MCP-1 signaling pathways.     

Vitamin D accelerates resolution of inflammatory responses during tuberculosis treatment

Calcidiol, the major circulating metabolite of vitamin D, supports induction of pleiotropic antimicrobial responses in vitro. Vitamin D supplementation elevates circulating calcidiol concentrations, and thus has a potential role in the prevention and treatment of infection. The immunomodulatory effects of administering vitamin D to humans with an infectious disease have not previously been reported. To characterize these effects, we conducted a detailed longitudinal study of circulating and antigen-stimulated immune responses in ninety-five patients receiving antimicrobial therapy for pulmonary tuberculosis who were randomized to receive adjunctive high-dose vitamin D or placebo in a clinical trial, and who fulfilled criteria for per-protocol analysis. Vitamin D supplementation accelerated sputum smear conversion and enhanced treatment-induced resolution of lymphopaenia, monocytosis, hypercytokinaemia, and hyperchemokinaemia. Administration of vitamin D also suppressed antigen-stimulated proinflammatory cytokine responses, but attenuated the suppressive effect of antimicrobial therapy on antigen-stimulated secretion of IL-4, CC chemokine ligand 5, and IFN-α. We demonstrate a previously unappreciated role for vitamin D supplementation in accelerating resolution of inflammatory responses during tuberculosis treatment. Our findings suggest a potential role for adjunctive vitamin D supplementation in the treatment of pulmonary infections to accelerate resolution of inflammatory responses associated with increased risk of mortality.     

Interleukin-6 Blood Levels in Sensitive and Multiresistant Tuberculosis

Abstract Background:   Interleukin-6 (IL-6) is a proinflammatory cytokine implicated in the immunopathogenesis of tuberculosis (TB). Multidrug resistance in tuberculosis is recognized worldwide as an important public health issue. The mechanism underlying TB pathogenesis in general and drug resistance in particular is not well understood, but it may be that IL-6 is one factor that enhances pathology in drug-resistant TB. The purpose of this study was to identify patterns of IL-6 production in active pulmonary TB with different sensitivity to standard drug therapy. Patients and Methods:   IL-6 blood levels were studied in 38 patients with active pulmonary TB: 23 patients were very sensitive to specific chemotherapy (STB), and 15 were multiresistant (MDRTB). An ELISA assay (Biossource) was used to quantify IL-6 in the sera of 38 TB patients and 63 healthy blood donors. Results:   The STB group was composed of 16 males (73.9%) and 7 females, MDRTB by 9 males (60%) and 6 females, and control group by 51 males (81%) and 12 females. The results showed a significant increase in IL-6 concentration in TB (median = 4.3 pg/ml, range 0.5–24) compared to that of healthy individuals (median = 0.5 pg/ml, range 0–2.8, p < 0.001). Additionally, IL-6 concentrations were increased in both STB (median = 4.1 pg/ml, range 0.5–24) and MDRTB (median = 5.1 pg/ml, range 0.5–12) groups in relation to controls (p < 0.001). In contrast, significant differences were not observed between STB and MDRTB groups (p > 0.05). Conclusion:   IL-6 levels were increased in pulmonary tuberculosis, independent of drug resistance.     

The relative frequency of Mycobacterium tuberculosis and Mycobacterium avium infections in HIV positive patients, Ahvaz, Iran

ObjectiveTo estimate the prevalence of Mycobacterium tuberculosis (M. tuberculosis) and Mycobacterium avium (M. avium) infections in HIV-positive patients suspected to have pulmonary and extrapulmonary mycobacterial co-infection using PCR technique.MethodsTotally 50 samples comprising sputum, pleural fluid and CSF taken from HIV positive patients suspected to have mycobacterial infection, were processed. The demographic information and results of acid fast staining and culture were recorded for each patient. The PCR for detecting of M. tuberculosis comprised of specific primers targeting IS6110 gene sequence. For detecting of M. avium, PCR with primers that amplifies the mig gene were used.ResultsFrom 50 samples processed, 45 were sputum (90%), 3 pleural fluid (6%) and 2 CSF (4%). In total, 8 (16%) were culture positive, 7 had positive acid fast staining (14 %) and 13 samples (26%) were positive using PCR technique. All the positive samples were sputum and belonged to patients with pulmonary infection. Of these, 9 were positive for M. tuberculosis (69.2%) and 4 were identified as M. avium (30.8%), which 2 out of 13 positive samples showed mixed infections by both mycobacteria.ConclusionsThe PCR shows the highest detection rate (26%) of mycobacteria compared with culture and acid fast staining. The majority of infections were with M. tuberculosis (18%) and this shows the importance of this mycobacterial co-infection in HIV positive patients in the region of study.     

Vitamin D receptor genetic polymorphisms in severe and recurrent tuberculosis in children

AimTo analyze the genetic polymorphisms of vitamin D receptor FokI, TaqI, ApaI and BsmI gene polymorphisms in children with severe and recurrent tuberculosis (TB).MethodsA prospective, observational study was conducted in 35 children with severe and recurrent TB referred to our Pediatric TB clinic at a tertiary referral center for children. The blood samples were analysed for genetic polymorphisms of Vitamin D receptor with respect to FokI, TaqI, ApaI and BsmI genotypes and their individual alleles and association of various clinical and laboratory parameters were analysed.ResultTen (28.6%) children had recurrent TB and 26 (74.3%) had severe TB. The severity of TB was not associated with Ff and ff polymorphism of FokI (Odd's ratio 7.88) as compared to no FokI polymorphism. Absence of FokI polymorphism was associated with recurrent lymph node TB (Odds ratio 3.429). Presence of Tt polymorphism of TaqI (p = 0.04) and Fok1 Polymorphism [Odds ratio 7.88] were not associated with recurrent TB.ConclusionRecurrent TB was absent in presence of Tt polymorphism of TaqI. Severe TB was not associated polymorphism of Vitamin D receptor polymorphisms.     

Th1 and Th17 immune responses to viable Propionibacterium acnes in patients with sarcoidosis

BackgroundPropionibacterium acnes and Mycobacterium tuberculosis have emerged as probable candidates responsible for sarcoidosis. This study was conducted to investigate the Th1/Th17 responses elicited by these pathogens in sarcoidosis and to clarify the causative role of these pathogens.MethodsPeripheral blood mononuclear cells (PBMCs) obtained from patients with sarcoidosis and from healthy volunteers were, respectively, co-cultured with viable P. acnes, with Bacille de Calmette et Guérin (BCG) as a viable M. tuberculosis complex, and with the early secretory antigenic target (ESAT)-6. Th1 cytokine production was measured using RT-PCR and enzyme-linked immunospot (ELISPOT) assays, and interleukin (IL)-17 mRNA expression was measured by RT-PCR.ResultsIL-2 secretion from PBMCs after stimulation with P. acnes was significantly higher in patients with sarcoidosis than in the controls. Similarly, IL-2 and IL-12 mRNA expression after stimulation with P. acnes was significantly higher in PBMCs from patients with sarcoidosis than in PBMCs from controls. In contrast, IL-17 mRNA expression was significantly lower in PBMCs from patients with sarcoidosis than in PBMCs from controls. No significant differences between the groups were observed in the responses to stimulation with BCG or ESAT-6.ConclusionSarcoidosis may arise from an imbalance of Th1/Th17 immune responses against viable P. acnes, but not M. tuberculosis complex.     

Serodiagnosis of tuberculosis: Detection of mycobacterial antibodies and antigens

Setting: The diagnosis of tuberculosis is based primarily on identification of mycobacteria and on clinical evidence. Recently, serological studies have been widely used experimentally as a diagnostic approach.Objective: The aim of our study was to optimize serodiagnosis of tuberculosis by detecting mycobacterial antigens and antibodies in sera from patients with lung tuberculosis, non-related diseases and healthy controls.Design: Mycobacterium tuberculosis H37Rv was disintegrated by pressure. Cell walls were extracted with 3 M KCL and were subjected to gel filtration in Toyopearl gel. Immune sera were prepared by immunization of rabbits with cell wall material. Anti H37Rv antibodies were purified by affinity chromatography. The reagents obtained were used to detect serum antibodies and antigens (following immune complex dissociation) using ELISA.Results: Using fraction 6 of cell wall extract, antibodies were detected in 72.2% of TB patients; there were no positive reactions in control subjects. By use of affinity-purified antibodies, antigens were detected in 77.1% of TB patients, 10% of patients with unrelated diseases and 6.7% of healthy controls.Conclusion: Effective serodiagnosis of tuberculosis can be achieved only by combining detection of both circulating antibodies and antigens using highly specific purified reagents and immune complex-dissociated sera.     

Effect of cytokine modulation by thalidomide on the granulomatous response in murine tuberculosis

Setting: Experimental murine tuberculosis.Objective: To evaluate the effect of cytokine modulation by thalidomide on the progression of the lung granulomatous response following aerosol tuberculosis infection in mice.Design: Mice infected by the respiratory route with 200–500 viable Mycobacterium tuberculosis Erdman were treated with daily subcutaneous injections of thalidomide (30 mg/kg) or saline for 4 weeks. The bacillary load, granulomatous response and cytokine production in the lungs were evaluated.Results: Aerosol M. tuberculosis infection resulted in a progressive granulomatous response in the lungs. At 28 days after infection, large granulomata with central necrosis and no apoptosis were observed. The infection induced high serum and lung cytokine mRNA levels. Thalidomide treatment resulted in a significant reduction in tumor necrosis factor-a, interleukin 6 (IL-6) and IL-10 protein levels (blood) and mRNA expression (lungs). IL-12 and interferon-γ were unaffected. The lungs of thalidomide-treated mice had smaller granulomata with apoptotic cells and no necrosis. Thalidomide treatment did not change the bacillary load.Conclusion: Thalidomide immunomodulation reduces inflammatory cytokines and concomitant lung pathology following acute aerosol M. tuberculosis infection, without increasing the bacillary load.     

Identification of mycobacteria and other acid fast organisms associated with pulmonary disease

ObjectiveTo identify Mycobacterium tuberculosis (M. tuberculosis) and other acid fast organisms isolated from sputum of HIV positive adult patients with pulmonary disease in Jos, Nigeria.MethodsAcid fast organisms isolated from 80 acid fast bacilli (AFB) positive sputa of HIV positive adult patients suspected for tuberculosis in Jos, Nigeria were identified for members of M. tuberculosis complex (M. tuberculosis, Mycobacterium bovis, Mycobacterium africanum, Mycobacterium canetti, Mycobacterium microti and Mycobacterium caprae) by use of spoligootyping, Multiplex Gen Probe, Hain genotype assay and gene sequencing for spoligotype negative isolates.ResultsSeven different spoligotypes of M. tuberculosis complex were identified from 70/80 (87.5%) total number of isolates. Mycobacterium kansasii (1), Mycobacterium dulvalii (1) Nocardia species (1) and Tsukamurella species (2) were detected from 5/10 spoligotype negative isolates.ConclusionsAlthough M. tuberculosis is the dominant AFB associated with chronic pulmonary disease in Jos, Nigeria, other clinically relevant mycobacteria were also observed in the study. This suggests that other AFB positive microorganisms associated with tuberculosis-like symptoms might be misdiagnosed and incorrectly treated as M. tuberculosis. It is therefore necessary for laboratories in tuberculosis high burden countries to step up diagnostic procedures beyond routine smear microscopy.