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The performance characteristics of the enzyme-linked immunospot assay (ELISPOT) assay (T-SPOT TB) for the diagnosis of latent tuberculosis infection in HIV-infected individuals are unknown. Given that ELISPOT enumerates Mycobacterium tuberculosis antigen-specific IFN-gamma-secreting T cells, HIV-associated immunosuppression might adversely affect test performance. However, we found that 28 out of 29 HIV-positive individuals (97%) gave evaluable test results, and performance was independent of the CD4 T-cell count. ELISPOT test performance appears to be independent of HIV-associated immunosuppression.

Background: Correcting the Th2 shift in HIV/AIDS represents a potential intervention strategy. However data on interleukin (IL)-4 expression in HIV or AIDS are un-interpretable because of failure to distinguish between IL-4 and its splice variant and natural antagonist, IL-4delta2., Objective: To determine Th1 [interferon (IFN)-gamma], IL-4delta2 and Th2 (IL-4) expression in whole blood and lung lavage from healthy volunteers and in HIV or HIV-tuberculosis (TB) co-infection., Design: Cross-sectional with prospective cohort., Methods: Expression of IL-4delta2, IL-4 and IFN-gamma were determined by quantitative real-time PCR, using unstimulated cells from whole blood and lung lavage, in 20 HIV-TB (pulmonary) co-infected patients, 20 matched HIV-positive controls and 20 HIV-negative healthy volunteers. Results were correlated with plasma viral load, CD4 cell counts, radiological scores and response to anti-TB treatment., Results: Compared to HIV negative donors, stable HIV-positive donors did not have increased levels of mRNA encoding IL-4, IL-4delta2 or IFN-gamma in blood or lavage. By contrast, the HIV-TB co-infected donors had increased IL-4 and IFN-gamma in both compartments. However the antagonist, IL-4delta2 was increased only in lavage. Consequently the dominant form was IL-4delta2 in lavage, but IL-4 itself in blood. The lung IL-4/IFN-gamma ratio correlated with radiological disease extent. With anti-TB treatment, IL-4 levels did not change whilst IL-4delta2 levels increased significantly., Conclusions: IL-4 and its natural antagonist, IL-4delta2 and are not upregulated in the absence of opportunistic infection. However in HIV-TB co-infection both cytokines increase in lung, but only IL-4 in the periphery. Further studies are required to determine if IL-4 facilitates systemic HIV progression.

Tuberculosis is a global public health catastrophe responsible for >8 million cases of illness and 2 million deaths annually. Pulmonary cavitation with cough-generated aerosol is the principle means of spread, and lung remodeling (healed cavitation, fibrosis, and bronchiectasis) is a major cause of lung disability, surpassing all other diffuse parenchymal lung diseases combined. Efficient granuloma turnover is mycobactericidal, and extracellular matrix is disbanded without scarring. In many with progressive disease, however, there is dysregulated granuloma turnover, liquefactive necrosis, and pathological scarring. The pathological mechanisms and the related immunological pathways underpinning these phenomena are reviewed in the present article. Further studies are needed to identify and develop specific immunotherapeutic interventions that target immunopathology, since they have the potential to substantially reduce spread.

Rationale: Tuberculosis progresses despite potent Th1 responses. A putative explanation is the simultaneous presence of a subversive Th2 response. However, interpretation is confounded by interleukin 4delta2 (IL-4delta2), a splice variant and inhibitor of IL-4., Objective: To study levels of mRNA encoding IL-4 and IL-4delta2, and their relationship to treatment and clinical parameters, in cells from lung lavage and blood from patients with pulmonary tuberculosis., Methods: IL-4delta2, IFN-gamma, IL-4, and soluble CD30 (sCD30) levels were measured by polymerase chain reaction and relevant immunoassays in 29 patients and matched control subjects lacking responses to tuberculosis-specific antigens., Results: mRNA levels for IL-4 and IL-4delta2 were elevated in unstimulated cells from blood and lung lavage of patients versus control subjects (p < 0.005). In control subjects, there were low basal levels of IL-4 and IL-4delta2 mRNA expressed mainly by non-T cells (p < 0.05). However, in patients, there were greater levels of mRNA for both cytokines in both T- and non-T-cell populations (p < 0.05 compared with control subjects). Radiologic disease correlated with the IL-4/IFN-gamma ratio and sCD30 (p < 0.005). After chemotherapy, IL-4 mRNA levels remained unchanged, whereas IL-4delta2 increased in parallel with IFN-gamma (p < 0.05). Sonicates of Mycobacterium tuberculosis upregulated expression of IL-4 relative to IL-4delta2 in mononuclear cell cultures from patients (p < 0.05)., Conclusions: A Th2-like response, prominent in T cells and driven by tuberculosis antigen, is present in tuberculosis and modulated by treatment, suggesting a role for IL-4 and IL-4delta2 in the pathogenesis of tuberculosis and their ratio as a possible marker of disease activity. The specific antigens inducing the IL-4 response require identification to facilitate future vaccine development strategies.

Tuberculosis is out of control in developing countries, where it is killing millions of people every year. In these areas, the present vaccine - Mycobacterium bovis bacillus Calmette-Guérin (BCG) - is failing. Progressive tuberculosis occurs because the potentially protective T helper 1 (T(H)1)-cell response is converted to an immunopathological response that fails to eliminate the bacteria. Here, we discuss the data indicating that the problem in developing countries is not a lack of adequate T(H)1-cell responses but, instead, an exaggerated tendency to switch to immunopathological responses. We propose that a successful vaccine needs to block this immunopathology, because it is not the quantity of T(H)1-cell activity that matters but, rather, its context.

Real-time RT-PCR has become a common technique, no longer limited to specialist core facilities. It is in many cases the only method for measuring mRNA levels of vivo low copy number targets of interest for which alternative assays either do not exist or lack the required sensitivity. Benefits of this procedure over conventional methods for measuring RNA include its sensitivity, large dynamic range, the potential for high throughout as well as accurate quantification. To achieve this, however, appropriate normalisation strategies are required to control for experimental error introduced during the multistage process required to extract and process the RNA. There are many strategies that can be chosen; these include normalisation to sample size, total RNA and the popular practice of measuring an internal reference or housekeeping gene. However, these methods are frequently applied without appropriate validation. In this review we discuss the relative merits of different normalisation strategies and suggest a method of validation that will enable the measurement of biologically meaningful results.

Purpose of Review: The tuberculin skin test, now over a century old, is prone to reader variability, and outcomes are influenced by cross-reactivity with environmental mycobacteria, previous bacillus Calmette-Guerin (BCG) vaccination, and anergy in immunosuppressed individuals. More recently, T-cell-based interferon-gamma responses to Mycobacterium tuberculosis - specific antigens have been investigated for their role in diagnosing latent tuberculosis infection., Recent Findings: We review the evidence supporting the utility of the interferon-gamma assay for the diagnosis of latent tuberculosis infection (LTBI) in low-prevalence countries. We discuss the principle of the test, technical factors related to performance, and its utility in active tuberculosis, in specialised subgroups such as immunocompromised patients, and its applicability in developing countries., Summary: Compared with the tuberculin skin test, the antigen-specific interferon-gamma assay, when used in a standardised protocol (overnight incubation assay using a combination of two antigens) for the diagnosis of LTBI, has greater specificity in BCG-vaccinated individuals, displays a stronger association with exposure, and is less biased by environmental mycobacteria such as Mycobacterium avium. Prospective studies are required, however, to confirm that treating LTBI, as defined by the interferon-gamma assay, will reduce the tuberculosis burden in low-prevalence countries and whether interferon-gamma responses are predictive of those who have a high risk of progression to active tuberculosis. Further studies are required to address the utility of the interferon-gamma assay in specialised subgroups of patients, in developing countries, and as a marker of disease activity.

Tuberculosis vaccine candidates are entering clinical studies in areas where BCG fails. This is a high-risk strategy. We suggest that geographical variation in the efficacy of BCG is related to the presence in developing countries of a cross-reactive background Th2-like response, probably attributable to exposure of mother and infant to helminths and environmental mycobacteria. Such Th2-like activity can stop Mycobacterium tuberculosis from being pushed into a latent state by the Th1 response, impair bactericidal functions and cause toxicity of TNF-alpha and pulmonary fibrosis. A successful vaccine, rather than driving a Th1 response, might need to suppress this pre-existing subversive Th2-like component.

There is little data about the use of different oxygen sources during assessment for long-term oxygen therapy (LTOT) and how this impacts upon blood oxygen tensions and prescribed flow rates. Patients with chronic obstructive pulmonary disease (COPD), n=30, had assessments for LTOT using both an oxygen-concentrator and piped hospital oxygen (wall-oxygen) as supply sources. In addition, a random survey of 64 hospitals was conducted to determine what source of oxygen supply was used during assessments. Wall-oxygen was used by 89% of hospitals to perform assessments. During assessments, the median oxygen flow required to achieve an arterial oxygen tension (Pa,O2) >8 kPa was significantly greater for an oxygen-concentrator than for wall-oxygen, with a median difference (range) in flow of 1 (0-3) L. This difference was most likely in those with an forced expiratory volume <30% of predicted. At an oxygen flow of 1 L.min(-1), the mean P(a,O2) using an oxygen-concentrator was significantly lower than that of the wall-oxygen value, with a difference of 1.32+/-1.19 kPa (mean+/-SD). The common practice of using wall-oxygen to perform assessments significantly underestimates the required oxygen-concentrator flow rate. This may have implications for the long-term effect of domiciliary oxygen therapy.

Background: The benefit of highly active antiretroviral therapy (HAART) in the treatment of patients coinfected with tuberculosis (TB) and human immunodeficiency virus (HIV) is unclear because of concerns about treatment-related complications., Methods: We compared outcomes in patients starting TB treatment during the pre-HAART era (before 1996; n=36) with those in patients starting treatment during the HAART era (during or after 1996; n=60)., Results: During a median of 3.6 years of follow-up, 49 patients died or had an AIDS event. Compared with patients in the pre-HAART group, those in the HAART group had a lower risk of death (cumulative at 4 years, 43% vs. 22%; P=.012) and of death or having an AIDS event (69% vs. 43%; P=.023). Event risk within the first 2 months of TB treatment was exceptionally high in patients with CD4(+) cell counts <100 cells/mm(3) and declined thereafter. HAART use during follow-up was associated with a marked reduction in event risk (adjusted hazard ratio, 0.38 [95% confidence interval, 0.16-0.91])., Conclusions: HAART substantially reduces new AIDS events and death in coinfected patients. Those with a CD4(+) cell count <100 cells/mm(3) have a high event risk during the intensive phase of anti-TB treatment. These data should be taken into account when deciding to delay HAART in coinfected patients with CD4(+) cell counts <100 cells/mm(3).

Current attempts to find a vaccine for tuberculosis (TB) are based on the assumption that it must drive a Th1 response. We review the evidence that progressive disease might not be due to absence of Th1, but rather to the subversive effect of an unusual Th2-like response, involving interleukin-4 (IL-4) and IL-4delta2. This Th2-like response can impair bactericidal function and lead to toxicity of tumour necrosis factor-alpha (TNF-alpha) and to pulmonary fibrosis. If this is important, effective vaccines will need to suppress pre-existing Th2-like activity. Such vaccines are feasible and are active therapeutically in mouse TB.

The aim of this study was to evaluate the safety and efficacy of azathioprine in the treatment of interstitial lung disease (ILD) associated with systemic sclerosis (SSc). The records of patients with SSc with ILD who were treated with azathioprine were reviewed. Patients were treated with azathioprine and low-dose prednisone if they had progressive pulmonary symptoms (deterioration in the dyspnea score) or poor or deteriorating lung function. Response was classified as improved if the FVC increased more than 10% from baseline, and stable if it remained within 10% of baseline. Serial dyspnea scores were recorded. Eleven patients were treated with azathioprine, three of whom received treatment for 6 months or less owing to adverse effects (nausea, leukopenia and pulmonary tuberculosis in one patient each). The remaining eight patients received at least 12 months' treatment and the results suggested an improvement in the mean percent predicted FVC from a baseline value of 54.25+/-3.53 to 63.38+/-6.15 after 12 months ( p=0.101). Overall, five patients improved and three remained stable. The mean dyspnea score ( n=8) improved from a baseline of 1.55+/-0.19 to 0.50+/-0.19 at 12 months ( p=0.011). This is the first case series of patients with SSc-associated ILD treated with azathioprine. Our results suggest that azathioprine may have a role in stabilizing lung function and improving symptoms in SSc, although this needs confirmation by a randomized controlled trial.

Background: Virtual bronchoscopy software is now available to district general hospitals (DGHs). There is limited information on the clinical utility of virtual bronchoscopy and whether it offers any additional information over conventional axial computed tomography in the setting of a busy DGH chest unit., Methods: Virtual bronchoscopy and computed tomography findings were compared in all patients who had a virtual bronchoscopy study over a 12 month period., Results: Eighteen consecutive patients had virtual bronchoscopy for a specific clinical indication over the study period. Additional information was conveyed by virtual bronchoscopy in five patients (in four patients the airways distal to an obstruction were better visualised thereby influencing decisions about airway stenting and in one patient the virtual bronchoscopy study showed an endobronchial lesion missed on computed tomography). In nine patients who were unfit for fibreoptic bronchoscopy (FOB) the radiologist was more confident in excluding an obstructive airway lesion. The main indication for performing a virtual bronchoscopy study was to rule out an obstructive airway lesion in patients who were unfit for FOB (n = 11)., Conclusion: Virtual bronchoscopy is feasible and useful in the management of a few selected patients in a DGH chest unit. Virtual bronchoscopy may convey additional information over computed tomography when the distal airways need to be visualised and for discrete endoluminal lesions.

Analysis of RNA expression using techniques like real-time PCR has traditionally used reference or housekeeping genes to control for error between samples. This practice is being questioned as it becomes increasingly clear that some housekeeping genes may vary considerably in certain biological samples. We used real-time reverse transcription PCR (RT-PCR) to assess the levels of 13 housekeeping genes expressed in peripheral blood mononuclear cell culture and whole blood from healthy individuals and those with tuberculosis. Housekeeping genes were selected from conventionally used ones and from genes reported to be invariant in human T cell culture. None of the commonly used housekeeping genes [e.g., glyceraldehyde-phosphate-dehydrogenase (GAPDH)] were found to be suitable as internal references, as they were highly variable (>30-fold maximal variability). Furthermore, genes previously found to be invariant in human T cell culture also showed large variation in RNA expression (>34-fold maximal variability). Genes that were invariant in blood were highly variable in peripheral blood mononuclear cell culture. Our data show that RNA specifying human acidic ribosomal protein was the most suitable housekeeping gene for normalizing mRNA levels in human pulmonary tuberculosis. Validations of housekeeping genes are highly specific for a particular experimental model and are a crucial component in assessing any new model.

Objective: To compare the views of doctors and patients on whether doctors should wear white coats and to determine what shapes their views., Methods: A questionnaire study of 400 patients and 86 doctors was performed., Results: All 86 of the doctors' questionnaires were included in the analysis but only 276 of the patients were able to complete a questionnaire. Significantly more patients (56%) compared with their doctors (24%) felt that doctors should wear white coats (p<0.001). Only age (>70 years) (p<0.001) and those patients whose doctors actually wore white coats (p<0.001) were predictive of whether patients favoured white coats. The most common reason given by patients was for easy identification (54%). Less than 1% of patients believed that white coats spread infection. Only 13% of doctors wore white coats as they were felt to be an infection risk (70%) or uncomfortable (60%). There was no significant difference between doctor subgroups when age, sex, grade, and specialty were analysed., Conclusion: In contrast to doctors, who view white coats as an infection risk, most patients, and especially those older than 70 years, feel that doctors should wear them for easy identification. Further studies are needed to assess whether this affects patients' perceived quality of care and whether patient education will alter this view.

Background: The British Thoracic Society (BTS) guidelines have not been examined collectively for their impact on chronic obstructive pulmonary disease (COPD). Whether intensive outpatient follow up of COPD patients after acute admission, using these guidelines, improved quality of life compared to the "usual practice" of primary care follow up was investigated., Methods: Altogether 103 patients with a new diagnosis of COPD were admitted and screened over a four year period. Seventy patients were excluded because of another dominant medical condition or a mandatory requirement for intervention. Patients were randomised to regular primary care (control group, n = 15) or chest clinic follow up (intervention group, n = 10). Spirometry, oxygen saturation, St George's Respiratory Questionnaire (SGRQ), and Short Form 36 questionnaire were measured at baseline and six months. The intervention group was reviewed at least four times in the six month period and received spirometry, ambulatory oxygen assessment, smoking cessation advice, nebuliser assessment, a steroid trial, advice about nutrition/exercise, and introduction to a patient support group., Results: There was no significant difference between baseline measurements in the two groups. There was a significant mean (SD) improvement in the SGRQ symptom score from baseline to six months in the intervention group [20.98 (20.36)] compared with the controls [0.23 (12.55)] (p = 0.004). At six months the SGRQ symptom score, impact score, and total score was significantly better in the intervention than the control group (p = 0.01, 0.02, and 0.02)., Conclusion: Aggressive implementation of BTS guidelines after initial hospitalisation may improve respiratory health specific quality of life scores in patients with COPD. Larger studies are needed to confirm this finding.