Malnutrition leads to increased inflammation and expression of tuberculosis risk signatures in recently exposed household contacts of pulmonary tuberculosis.

Background: Most individuals exposed to Mycobacterium tuberculosis (Mtb) develop latent tuberculosis infection (LTBI) and remain at risk for progressing to active tuberculosis disease (TB). Malnutrition is an important risk factor driving progression from LTBI to TB. However, the performance of blood-based TB risk signatures in malnourished individuals with LTBI remains unexplored. The aim of this study was to determine if malnourished and control individuals had differences in gene expression, immune pathways and TB risk signatures. Methods: We utilized data from 50 tuberculin skin test positive household contacts of persons with TB - 18 malnourished participants (body mass index [BMI] < 18.5 kg/m2) and 32 controls (individuals with BMI ≥ 18.5 kg/m2). Whole blood RNA-sequencing was conducted to identify differentially expressed genes (DEGs). Ingenuity Pathway Analysis was applied to the DEGs to identify top canonical pathways and gene regulators. Gene enrichment methods were then employed to score the performance of published gene signatures associated with progression from LTBI to TB. Results: Malnourished individuals had increased activation of inflammatory pathways, including pathways involved in neutrophil activation, T-cell activation and proinflammatory IL-1 and IL-6 cytokine signaling. Consistent with known association of inflammatory pathway activation with progression to TB disease, we found significantly increased expression of the RISK4 (area under the curve [AUC] = 0.734) and PREDICT29 (AUC = 0.736) progression signatures in malnourished individuals. Conclusion: Malnourished individuals display a peripheral immune response profile reflective of increased inflammation and a concomitant increased expression of risk signatures predicting progression to TB. With validation in prospective clinical cohorts, TB risk biomarkers have the potential to identify malnourished LTBI for targeted therapy.

Early alveolar macrophage response and IL-1R-dependent T cell priming determine transmissibility of Mycobacterium tuberculosis strains.

Mechanisms underlying variability in transmission of Mycobacterium tuberculosis strains remain undefined. By characterizing high and low transmission strains of M.tuberculosis in mice, we show here that high transmission M.tuberculosis strain induce rapid IL-1R-dependent alveolar macrophage migration from the alveolar space into the interstitium and that this action is key to subsequent temporal events of early dissemination of bacteria to the lymph nodes, Th1 priming, granulomatous response and bacterial control. In contrast, IL-1R-dependent alveolar macrophage migration and early dissemination of bacteria to lymph nodes is significantly impeded in infection with low transmission M.tuberculosis strain; these events promote the development of Th17 immunity, fostering neutrophilic inflammation and increased bacterial replication. Our results suggest that by inducing granulomas with the potential to develop into cavitary lesions that aids bacterial escape into the airways, high transmission M.tuberculosis strain is poised for greater transmissibility. These findings implicate bacterial heterogeneity as an important modifier of TB disease manifestations and transmission.

Rapamycin modulates pulmonary pathology in a murine model of Mycobacterium tuberculosis infection.

Tuberculosis (TB) treatment regimens are lengthy, causing non-adherence to treatment. Inadequate treatment can lead to relapse and the development of drug resistance TB. Furthermore, patients often exhibit residual lung damage even after cure, increasing the risk for relapse and development of other chronic respiratory illnesses. Host-directed therapeutics are emerging as an attractive means to augment the success of TB treatment. In this study, we used C3HeB/FeJ mice as an experimental model to investigate the potential role of rapamycin, a mammalian target of rapamycin inhibitor, as an adjunctive therapy candidate during the treatment of Mycobacterium tuberculosis infection with moxifloxacin. We report that administration of rapamycin with or without moxifloxacin reduced infection-induced lung inflammation, and the number and size of caseating necrotic granulomas. Results from this study strengthen the potential use of rapamycin and its analogs as adjunct TB therapy, and importantly underscore the utility of the C3HeB/FeJ mouse model as a preclinical tool for evaluating host-directed therapy candidates for the treatment of TB.

A Parsimonious Host Inflammatory Biomarker Signature Predicts Incident Tuberculosis and Mortality in Advanced Human Immunodeficiency Virus.

BACKGROUND: People with advanced human immunodeficiency virus (HIV) (CD4 < 50) remain at high risk of tuberculosis (TB) or death despite the initiation of antiretroviral therapy (ART). We aimed to identify immunological profiles that were most predictive of incident TB disease and death. METHODS: The REMEMBER randomized clinical trial enrolled 850 participants with HIV (CD4 < 50 cells/µL) at ART initiation to receive either empiric TB treatment or isoniazid preventive therapy (IPT). A case-cohort study (n = 257) stratified by country and treatment arm was performed. Cases were defined as incident TB or all-cause death within 48 weeks after ART initiation. Using multiplexed immunoassay panels and ELISA, 26 biomarkers were assessed in plasma. RESULTS: In total, 52 (6.1%) of 850 participants developed TB; 47 (5.5%) died (13 of whom had antecedent TB). Biomarkers associated with incident TB overlapped with those associated with death (interleukin [IL]-1ß, IL-6). Biomarker levels declined over time in individuals with incident TB while remaining persistently elevated in those who died. Dividing the cohort into development and validation sets, the final model of 6 biomarkers (CXCL10, IL-1ß, IL-10, sCD14, tumor necrosis factor [TNF]-α, and TNF-ß) achieved a sensitivity of 0.90 (95% confidence interval [CI]: .87-.94) and a specificity of 0.71(95% CI: .68-.75) with an area under the curve (AUC) of 0.81 (95% CI: .78-.83) for incident TB. CONCLUSION: Among people with advanced HIV, a parsimonious inflammatory biomarker signature predicted those at highest risk for developing TB despite initiation of ART and TB preventive therapies. The signature may be a promising stratification tool to select patients who may benefit from increased monitoring and novel interventions. CLINICAL TRIALS REGISTRATION: NCT01380080.

Transmission phenotype of Mycobacterium tuberculosis strains is mechanistically linked to induction of distinct pulmonary pathology.

In a study of household contacts (HHC), households were categorized into High (HT) and Low (LT) transmission groups based on the proportion of HHC with a positive tuberculin skin test. The Mycobacterium tuberculosis (Mtb) strains from HT and LT index cases of the households were designated Mtb-HT and Mtb-LT, respectively. We found that C3HeB/FeJ mice infected with Mtb-LT strains exhibited significantly higher bacterial burden compared to Mtb-HT strains and also developed diffused inflammatory lung pathology. In stark contrast, a significant number of mice infected with Mtb-HT strains developed caseating granulomas, a lesion type with high potential to cavitate. None of the Mtb-HT infected animals developed diffused inflammatory lung pathology. A link was observed between increased in vitro replication of Mtb-LT strains and their ability to induce significantly high lipid droplet formation in macrophages. These results support that distinct early interactions of Mtb-HT and Mtb-LT strains with macrophages and subsequent differential trajectories in pathological disease may be the mechanism underlying their transmission potential.

Effect of malnutrition on radiographic findings and mycobacterial burden in pulmonary tuberculosis.

BACKGROUND: The relationship between malnutrition and tuberculosis (TB) severity is understudied. We investigated the effect of malnutrition on radiographic findings and mycobacterial burden. METHODS: Subjects included newly diagnosed, smear-positive, culture-confirmed, pulmonary TB cases enrolled in the Regional Prospective Observational Research for TB (RePORT) cohort. Multivariate regression models were used to evaluate the relationship at start of treatment between body mass index (BMI) and chest radiograph (CXR) findings of cavitation and percentage of lung affected and mycobacterial growth indicator tube (MGIT) time to positive (TTP). Severe malnutrition was defined as BMI<16 kg/m2, moderate malnutrition as 16-18.4kg/m2, and "normal"/overweight as ≥18.5 kg/m2. RESULTS: Of 173 TB cases with chest x-ray data, 131 (76%) were male. The median age was 45 years (range 16-82); 42 (24%) had severe malnutrition and 58 (34%) moderate malnutrition. Median percentage of lung affected was 32% (range 0-95), and 132 (76%) had cavitation. Individuals with severe malnutrition had, on average, 11.1% [95% CI: 4.0-13.3] more lung affected, compared to those with normal BMI, controlling for diabetes and cavitation. In multivariable analyses, cases with severe malnutrition had a 4.6-fold [95% CI, 1.5-14.1] increased odds of cavitation compared to those with normal BMI, controlling for smoking. Median MGIT TTP was 194.5 hours. Neither severe (aRR 0.99; 95% CI, 0.9-1.2) nor moderate (aRR 0.97; 95% CI, 0.8-1.1) malnutrition was associated with MGIT TTP. CONCLUSION: We found that malnutrition was associated with increased extent of disease and cavitation on CXR. These findings may reflect the immunomodulatory effect of malnutrition on pulmonary pathology.

Ambroxol Induces Autophagy and Potentiates Rifampin Antimycobacterial Activity.

Host-directed therapy in tuberculosis is a potential adjunct to antibiotic chemotherapy directed at Mycobacterium tuberculosis Ambroxol, a lead compound, emerged from a screen for autophagy-inducing drugs. At clinically relevant doses, ambroxol induced autophagy in vitro and in vivo and promoted mycobacterial killing in macrophages. Ambroxol also potentiated rifampin activity in a murine tuberculosis model.

"Tuberculosis in advanced HIV infection is associated with increased expression of IFNγ and its downstream targets".

BACKGROUND: Tuberculosis (TB) is the major cause of death in Human Immunodeficiency Virus (HIV)-infected individuals. However, diagnosis of TB in HIV remains challenging particularly when HIV infection is advanced. Several gene signatures and serum protein biomarkers have been identified that distinguish active TB from latent infection. Our study was designed to assess if gene expression signatures and cytokine levels would distinguish active TB in advanced HIV. METHODS: We conducted a case-control study of whole blood RNA-Seq and plasma cytokine/chemokine analysis in HIV-infected with CD4+ T cell count of ≤ 100 cells/µl, with and without active TB. Next, the overlap of the differentially expressed genes (DEG) with the published signatures was performed and then receiver operator characteristic (ROC) analysis was done on small gene discriminators to determine their performance in distinguishing TB in advanced HIV. ELISA was performed on plasma to evaluate cytokine and chemokine levels. RESULTS: Hierarchical clustering of the transcriptional profiles showed that, in general, HIV-infected individuals with TB (TB-HIV) clustered separately from those without TB. IPA indicated that the TB-HIV signature was characterized by an increase in inflammatory signaling pathways. Analysis of overlaps between DEG in our data set with published TB signatures revealed that significant overlap was seen with one TB signature and one TB-IRIS signature. ROC analysis revealed that transcript levels of FcGR1A (AUC = 0.85) and BATF2 (AUC = 0.82), previously reported as consistent single gene classifiers of active TB irrespective of HIV status, performed successfully even in advanced HIV. Plasma protein levels of IFNγ, a stimulator of FcGR1A and BATF2, and CXCL10, also up-regulated by IFNγ, accurately classified active TB (AUC = 0.98 and 0.91, respectively) in advanced HIV. Neither of these genes nor proteins distinguished between TB and TB-IRIS. CONCLUSIONS: Gene expression of FcGR1A and BATF2, and plasma protein levels of IFNγ and CXCL10 have the potential to independently detect TB in advanced HIV. However, since other lung diseases were not included in this study, these final candidates need to be validated as specific to TB in the advanced HIV population with TB.

Isoniazid Preventive Therapy for People With HIV Who Are Heavy Alcohol Drinkers in High TB-/HIV-Burden Countries: A Risk-Benefit Analysis.

BACKGROUND: Isoniazid preventive therapy (IPT) reduces mortality among people living with HIV (PLHIV) and is recommended for those without active tuberculosis (TB) symptoms. Heavy alcohol use, however, is contraindicated for liver toxicity concerns. We evaluated the risks and benefits of IPT at antiretroviral therapy (ART) initiation to ART alone for PLHIV who are heavy drinkers in 3 high TB-/HIV-burden countries. METHODS: We developed a Markov simulation model to compare ART alone to ART with either 6 or 36 months of IPT for heavy drinking PLHIV enrolling in care in Brazil, India, and Uganda. Outcomes included nonfatal toxicity, fatal toxicity, life expectancy, TB cases, and TB death. RESULTS: In this simulation, 6 months of IPT + ART (IPT6) extended life expectancy over both ART alone and 36 months of IPT + ART (IPT36) in India and Uganda, but ART alone dominated in Brazil in 51.5% of simulations. Toxicity occurred in 160/1000 persons on IPT6 and 415/1000 persons on IPT36, with fatal toxicity in 8/1000 on IPT6 and 21/1000 on IPT36. Sensitivity analyses favored IPT6 in India and Uganda with high toxicity thresholds. CONCLUSIONS: The benefits of IPT for heavy drinkers outweighed its risks in India and Uganda when given for a 6-month course. The toxicity/efficacy trade-off was less in Brazil where TB incidence is lower. IPT6 resulted in fatal toxicity in 8/1000 people, whereas even higher toxicities of IPT36 negated its benefits in all countries. Data to better characterize IPT toxicity among HIV-infected drinkers are needed to improve guidance.

Quest for correlates of protection against tuberculosis.

A major impediment to tuberculosis (TB) vaccine development is the lack of reliable correlates of immune protection or biomarkers that would predict vaccine efficacy. Gamma interferon (IFN-γ) produced by CD4(+) T cells and, recently, multifunctional CD4(+) T cells secreting IFN-γ, tumor necrosis factor (TNF), and interleukin-2 (IL-2) have been used in vaccine studies as a measurable immune parameter, reflecting activity of a vaccine and potentially predicting protection. However, accumulating experimental evidence suggests that host resistance against Mycobacterium tuberculosis infection is independent of IFN-γ and TNF secretion from CD4(+) T cells. Furthermore, the booster vaccine MVA85A, despite generating a high level of multifunctional CD4(+) T cell response in the host, failed to confer enhanced protection in vaccinated subjects. These findings suggest the need for identifying reliable correlates of protection to determine the efficacy of TB vaccine candidates. This article focuses on alternative pathways that mediate M. tuberculosis control and their potential for serving as markers of protection. The review also discusses the significance of investigating the natural human immune response to M. tuberculosis to identify the correlates of protection in vaccination.

Differential expression of Toll-like receptors on human alveolar macrophages and autologous peripheral monocytes.

BACKGROUND: Toll-like receptors (TLRs) are critical components in the regulation of pulmonary immune responses and the recognition of respiratory pathogens such as Mycobacterium Tuberculosis (M.tb). Through examination of human alveolar macrophages this study attempts to better define the expression profiles of TLR2, TLR4 and TLR9 in the human lung compartment which are as yet still poorly defined. METHODS: Sixteen healthy subjects underwent venipuncture, and eleven subjects underwent additional bronchoalveolar lavage to obtain peripheral blood mononuclear and bronchoalveolar cells, respectively. Surface and intracellular expression of TLRs was assessed by fluorescence-activated cell sorting and qRT-PCR. Cells were stimulated with TLR-specific ligands and cytokine production assessed by ELISA and cytokine bead array. RESULTS: Surface expression of TLR2 was significantly lower on alveolar macrophages than on blood monocytes (1.2 +/- 0.4% vs. 57 +/- 11.1%, relative mean fluorescence intensity [rMFI]: 0.9 +/- 0.1 vs. 3.2 +/- 0.1, p < 0.05). The proportion of TLR4 and TLR9-expressing cells and the rMFIs of TLR4 were comparable between alveolar macrophages and monocytes. The surface expression of TLR9 however, was higher on alveolar macrophages than on monocytes (rMFI, 218.4 +/- 187.3 vs. 4.4 +/- 1.4, p < 0.05) while the intracellular expression of the receptor and the proportion of TLR9 positive cells were similar in both cell types. TLR2, TLR4 and TLR9 mRNA expression was lower in bronchoalveolar cells than in monocytes.Pam3Cys, LPS, and M.tb DNA upregulated TLR2, TLR4 and TLR9 mRNA in both, bronchoalveolar cells and monocytes. Corresponding with the reduced surface and mRNA expression of TLR2, Pam3Cys induced lower production of TNF-alpha, IL-1beta and IL-6 in bronchoalveolar cells than in monocytes. Despite comparable expression of TLR4 on both cell types, LPS induced higher levels of IL-10 in monocytes than in alveolar macrophages. M.tb DNA, the ligand for TLR9, induced similar levels of cytokines in both cell types. CONCLUSION: The TLR expression profile of autologous human alveolar macrophages and monocytes is not identical, therefore perhaps contributing to compartmentalized immune responses in the lungs and systemically. These dissimilarities may have important implications for the design and efficacy evaluation of vaccines with TLR-stimulating adjuvants that target the respiratory tract.

Extracellular-regulated kinase activation regulates replication of Mycobacterium avium intracellularly in primary human monocytes.

Mycobacterium avium-intracellulare (MAI) is a ubiquitous environmental pathogen that causes disseminated infection in immunocompromised patients, such as those with human immunodeficiency virus, interleukin-12 deficiency, or interferon-gamma receptor mutation. Colony morphotypes are associated with MAI pathogenicity. Our previous studies have reported that smooth-transparent (SmT) morphotypes are more virulent and induce less cytokine (interleukin-1beta and tumor necrosis factor-alpha) production by human monocytes than the smooth-domed (SmD) morphotypes. Mitogen-activated protein (MAP) kinases such as extracellular-regulated kinase (ERK) are activated by the phagocytosis of particle antigens in macrophages, and this ERK activation subsequently influences cytokine expression and the control of intracellular pathogen growth. The influence of MAP kinase activation on MAI replication in human monocytes was examined. Peripheral blood monocytes isolated from healthy subjects by Ficoll-Hypaque sedimentation were infected with virulent SmT or avirulent SmD MAI without or with MAP kinase inhibitors. MAP kinase activities were determined by in vitro kinase assay, intracellular MAI growth by CFU assay, and cytokines by enzyme-linked immunosorbent assay. MAI infection induced ERK and p38 activation. Inhibition of ERK by PD98059, but not p38, significantly increased intracellular MAI growth. Tumor necrosis factor-alpha release and interleukin-1beta production in response to MAI were reduced by MAP kinase inhibition. p38 inhibition tended to reduce cytokine production more substantially. These data suggest that ERK activation limits intra-monocytic MAI replication and enhances monocytic cytokine release, whereas p38 activation influences only cytokine release. The effect of MAP kinases on MAI growth might thus be mediated by the modulation of cytokine production.

Down-regulation of MHC II in mesenchymal stem cells at high IFN-gamma can be partly explained by cytoplasmic retention of CIITA.

Mesenchymal stem cells (MSCs) are located in postnatal bone marrow, show plasticity, are linked to various bone marrow disorders, exhibit phagocytosis, exert Ag-presenting properties (APC), and are immune suppressive. Unlike professional APCs, MSCs respond bimodally to IFN-gamma in MHC-II expression, with expression at 10 U/ml and baseline, and down-regulation at 100 U/ml. The effects at high IFN-gamma could not be explained by down-regulation of its receptor, IFN-gammaRI. In this study, we report on the mechanisms by which IFN-gamma regulates MHC-II expression in MSCs. Gel shift assay and Western blot analyses showed dose-dependent increases in activated STAT-1, indicating responsiveness by IFN-gammaRI. Western blots showed decreased intracellular MHC-II, which could not be explained by decreased transcription of the master regulator CIITA, based on RT-PCR and in situ immunofluorescence. Reporter gene assays with PIII and PIV CIITA promoters indicate constitutive expression of PIII in MSCs and a switch to PIV by IFN-gamma, indicating the presence of factors for effect promoter responses. We explained decreased MHC-II at the level of transcription because CIITA protein was observed in the cytosol and not in nuclei at high IFN-gamma level. The proline/serine/threonine region of CIITA showed significant decrease in phosphorylation at high IFN-gamma levels. An understanding of the bimodal effects could provide insights on bone marrow homeostasis, which could be extrapolated to MSC dysfunction in hematological disorders.

Pathogenesis and management of HIV/TB co-infection in Asia.

HIV infection increases the risk of reactivation of latent tuberculosis (TB), progression of a new infection or re-infection to active disease, and acceleration of the natural course of the disease with a more rapid spread of strains, including those that are drug resistant, in the community. TB also accelerates the course of HIV-induced disease by activating viral replication and accentuating the decline in CD4 T cell counts. In this chapter, TB-HIV co-infection is discussed in the context of the situation in Vietnam, particularly Ho Chi Minh City, the creation of a well-integrated TB control and research programs in Cambodia, and the broad principles of the co-management of TB/HIV particularly in the context of the associated atypical forms of pulmonary TB (PTB), increased non-PTB and the frequency of acid fast smear negative cases.

Mycobacterium tuberculosis growth control by lung macrophages and CD8 cells from patient contacts.

RATIONALE: Healthy household contacts (HHCs) of patients with active pulmonary tuberculosis are exposed aerogenically to Mycobacterium tuberculosis (Mtb), thus permitting the study of protective local immunity. OBJECTIVES: To assess alveolar macrophage (AM) and autologous blood CD4 and CD8 T-cell-mediated Mtb growth control in HHCs and healthy, unexposed community control subjects (CCs). METHODS: AMs were infected with Mtb strains H(37)Ra and H(37)Rv at multiplicities of infection 0.1 and 1. Mtb colony-forming units were evaluated on Days 1, 4, and 7. MAIN RESULTS: CD8 T cells from HHCs in 1:1 cocultures with AMs significantly (p < 0.05) increased Mtb growth control by AMs. In CCs, no detectable contribution of CD8 T cells to Mtb growth control was observed. CD4 T cells did not increase Mtb growth control in HHCs or in CCs. IFN-gamma, nitric oxide, and tumor necrosis factor were determined as potential mediators of Mtb growth control in AMs and AM/CD8 and AM/CD4 cocultures. IFN-gamma production in AM/CD4 was twofold higher than that in AM/CD8 cocultures in both HHCs and CCs (p < 0.05). Nitric oxide production from AMs of HHCs increased on Days 4 and 7 and was undetectable in AMs from CCs. IFN-gamma and nitric acid concentrations and Mtb growth control were not correlated. Tumor necrosis factor levels were significantly increased in AM/CD8 cocultures from HHCs compared with AM/CD8 cocultures from CCs (p < 0.05). CONCLUSION: Aerogenic exposure to Mtb in HHCs leads to expansion of Mtb-specific effector CD8 T cells that limit Mtb growth in autologous AMs.

Mechanisms of apoptosis of T-cells in human tuberculosis.

The role of TGF-beta TNF-alpha FasL and Bcl-2 in apoptosis of CD4 T-cells during active TB was studied. Coculture of PBMC from TB patients with neutralizing antibodies to TGF-beta or TNF-alpha decreased spontaneous (P < or = 0.05) and MTB-induced (P < or = 0.02) T-cell apoptosis by 50-90%, but effects were not additive. Interestingly, only levels of TGF-beta in supernatants correlated with rates of spontaneous and MTB-induced apoptosis. FasL surface and mRNA expression were higher in unstimulated and MTB-stimulated PBMC from patients than controls, and neutralization of FasL abrogated apoptosis of T-cells from patients only. Intracellular Bcl-2 protein was lower among unstimulated CD4 T-cells from patients than those from controls (P < or = 0.02), and MTB stimulation reduced intracellular Bcl-2 content in CD4 T-cells from patients only (P < or = 0.001). These findings may indicate that, during TB, predisposition of CD4 T-cells to apoptosis may involve both low expression of Bcl-2, and excessive expression of TGF-beta TNF-alpha and FasL.

A randomized, double-blind, placebo-controlled trial of the use of prednisolone as an adjunct to treatment in HIV-1-associated pleural tuberculosis.

BACKGROUND: Active tuberculosis may accelerate progression of human immunodeficiency virus (HIV) infection by promoting viral replication in activated lymphocytes. Glucocorticoids are used in pleural tuberculosis to reduce inflammation-induced pathology, and their use also might reduce progression of HIV by suppressing immune activation. We examined the effect that prednisolone has on survival in HIV-1-associated pleural tuberculosis. METHODS: We conducted a randomized, double-blind, placebo-controlled trial of prednisolone as an adjunct to tuberculosis treatment, in adults with HIV-1-associated pleural tuberculosis. The primary outcome was death. Analysis was by intention to treat. RESULTS: Of 197 participants, 99 were assigned to the prednisolone group and 98 to the placebo group. The mortality rate was 21 deaths/100 person-years (pyr) in the prednisolone group and 25 deaths/100 pyr in the placebo group (age-, sex-, and initial CD4+ T cell count-adjusted mortality rate ratio, 0.99 [95% confidence interval, 0.62-1.56] [P =.95]). Resolution of tuberculosis was faster in the prednisolone group, but recurrence rates were slightly (though not significantly) higher, and use of prednisolone was associated with a significantly higher incidence of Kaposi sarcoma (4.2 cases/100 pyr, compared with 0 cases/100 pyr [P =.02]). CONCLUSIONS: In view of the lack of survival benefit and the increased risk of Kaposi sarcoma, the use of prednisolone in HIV-associated tuberculous pleurisy is not recommended.

Sputum cytokine levels in patients with pulmonary tuberculosis as early markers of mycobacterial clearance.

Sputum and serum from patients with active pulmonary tuberculosis (TB), healthy purified protein derivative-positive adults, and patients with bacterial pneumonia were collected to simultaneously assess local immunity in the lungs and peripheral blood. To determine whether cytokine profiles in sputum from TB patients and control subjects were a reflection of its cellular composition, cytospin slides were prepared in parallel and assessed for the presence of relative proportions of epithelial cells, neutrophils, macrophages, and T cells. Gamma interferon (IFN-gamma) in sputum from TB patients was markedly elevated over levels for both control groups. With anti-TB therapy, IFN-gamma levels in sputum from TB patients decreased rapidly and by week 4 of treatment were comparable to those in sputum from controls. Further, IFN-gamma levels in sputum closely followed mycobacterial clearance. Although detected at fourfold-lower levels, IFN-gamma immunoreactivities in serum followed kinetics in sputum. TNF-alpha, interleukin 8 (IL-8) and IL-6 also were readily detected in sputum from TB patients at baseline and responded to anti-TB therapy. In contrast to IFN-gamma, however, TNF-alpha and IL-8 levels also were elevated in sputum from pneumonia controls. These data indicate that sputum cytokines correlate with disease activity during active TB of the lung and may serve as potential early markers for sputum conversion and response to anti-TB therapy.

Bactericidal activity in whole blood as a potential surrogate marker of immunity after vaccination against tuberculosis.

The development of new tuberculosis (TB) vaccines will require the identification of correlates of human protection. This study examined the balance between immunity and virulence in a whole blood infection model in which intracellular mycobacterial survival was measured using BACTEC. In the blood of tuberculin-negative donors, counts of Mycobacterium tuberculosis H(37)Ra organisms fell by 0.14 log(10) CFU during 96 h of whole blood culture, whereas counts of Mycobacterium bovis BCG, M. tuberculosis H(37)Rv, and a clinical TB isolate's organisms increased by 0.13, 0.43, and 1.04 log(10) CFU, respectively (P < 0.001), consistent with their relative virulence. Inhibition of tumor necrosis factor alpha by the addition of methylprednisolone or pentoxifylline or removal of CD4(+) or CD8(+) T cells by magnetic beads had deleterious effects on immune control of intracellular growth only in the blood of tuberculin-positive donors. Repeated vaccination of eight tuberculin-negative volunteers with M. bovis BCG resulted in a 0.3 log (50%) reduction in BCG CFU counts in the model compared to baseline values (P < 0.05). Three of the volunteers responded only after the second vaccination. These experiments indicate that whole blood culture may be used to measure immunity to M. tuberculosis and that further studies of repeated BCG vaccination are warranted.