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Background: The potential available market (PAM) for new diagnostics for tuberculosis that meet the specifications of the high-priority target product profiles (TPPs) is currently unknown., Methods: We estimated the PAM in 2020 in 4 high-burden countries (South Africa, Brazil, China, and India) for tests that meet the specifications outlined in the TPPs. The yearly PAM was estimated for the most likely application of each TPP., Results: In 2020 the PAM for all 4 countries together was estimated to be (1) 12M tests/year with a value of 48M-71M USD for a sputum smear-replacement test; (2) 16M tests/year with a value of 65M-97M USD for a biomarker test; (3) 18M tests/year with a value of 18M-35M USD for a triage test; (4) 12M tests/year with a value of 59M-2238M USD for a tuberculosis detection plus drug susceptibility test (DST) all-in-one or 1.5M tests/year for a DST that follows a positive tuberculosis detection test with a corresponding value of 75M-121M for both tuberculosis detection and DST., Conclusions: Although there is a considerable potential market for novel tuberculosis diagnostics that fit the specification of the TPPs in the 4 high-burden countries, the actual market for an individual product remains uncertain., (© The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Background: Four priority target product profiles for the development of diagnostic tests for tuberculosis were identified: 1) Rapid sputum-based (RSP), 2) non-sputum Biomarker-based (BMT), 3) triage test followed by confirmatory test (TT), and 4) drug-susceptibility testing (DST)., Methods: We assessed the cost of the new tests in suitable strategies and of the conventional diagnosis of tuberculosis as per World Health Organization guidelines, in 36 high tuberculosis and MDR burden countries. Costs were then compared to the available funding for tuberculosis at country level., Results: Costs of diagnosing tuberculosis using RSP ranged US$93-187 million/year; if RSP unit cost is of US$2-4 it would be lower/similar cost than conventional strategy with sputum smear microscopy (US$ 119 million/year). Using BMT (with unit cost of US$2-4) would cost US$70-121 million/year and be lower/comparable cost than conventional diagnostics. Using TT with TPP characteristics (unit cost of US$1-2) followed by Xpert would reduce diagnostic costs up to US$36 million/year. Costs of using different novel DST strategies for the diagnosis of drug resistance would be higher compared with conventional diagnosis., Conclusions: Introducing a TT or a biomarker test with optimal characteristics would be affordable from a cost and affordability perspective at the current available funding for tuberculosis. Additional domestic or donor funding would be needed in most countries to achieve affordability for other new diagnostic tests., (© The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

To accelerate the fight against tuberculosis, major diagnostic challenges need to be addressed urgently. Post-2015 targets are unlikely to be met without the use of novel diagnostics that are more accurate and can be used closer to where patients first seek care in affordable diagnostic algorithms. This article describes the efforts by the stakeholder community that led to the identification of the high-priority diagnostic needs in tuberculosis. Subsequently target product profiles for the high-priority diagnostic needs were developed and reviewed in a World Health Organization (WHO)-led consensus meeting. The high-priority diagnostic needs included (1) a sputum-based replacement test for smear-microscopy; (2) a non-sputum-based biomarker test for all forms of tuberculosis, ideally suitable for use at levels below microscopy centers; (3) a simple, low cost triage test for use by first-contact care providers as a rule-out test, ideally suitable for use by community health workers; and (4) a rapid drug susceptibility test for use at the microscopy center level. The developed target product profiles, along with complimentary work presented in this supplement, will help to facilitate the interaction between the tuberculosis community and the diagnostics industry with the goal to lead the way toward the post-2015 global tuberculosis targets., (© The Author 2014. Published by Oxford University Press on behalf of the Infectious Diseases Society of America. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.)

Background: Extrapulmonary tuberculosis (EPTB) and most pediatric TB cannot be diagnosed using sputum-based assays. The epidemiological impact of different strategies to diagnose EPTB and pediatric TB is unclear., Methods: We developed a dynamic epidemic model of TB in a hypothetical population with epidemiological characteristics similar to India. We evaluated the impact of four alternative diagnostic test platforms on adult EPTB and pediatric TB mortality over 10 years: (1) Nucleic acid amplification test optimized for diagnosis of EPTB ("NAAT-EPTB"); (2) NAAT optimized for pediatric TB ("NAAT-Peds"); (3) more deployable NAAT for sputum-based diagnosis of adult pulmonary TB ("point-of-care (POC) sputum NAAT"); and (4) more deployable NAAT capable of diagnosing all forms of TB using non-invasive, non-sputum specimens ("POC non-sputum NAAT")., Results: NAAT-EPTB lowered adult EPTB mortality by a projected 7.6% (95% uncertainty range [UR]: 6.5-8.8%). NAAT-Peds lowered pediatric TB mortality by 6.8% (UR: 4.9-8.4%). POC sputum NAAT, though only able to diagnose pulmonary TB, reduced projected pediatric TB deaths by 13.3% (UR: 4.6-15.7%) and adult EPTB deaths by 8.4% (UR 2.0-9.3%) simply by averting transmission of disease. POC non-sputum NAAT had the greatest effect, lowering pediatric TB mortality by 34.7% (UR: 26.8-38.7), and adult EPTB mortality by 38.5% (UR: 30.7-41.2). The relative impact of a POC sputum NAAT (i.e., enhanced deployability) versus NAAT-EPTB (i.e., enhanced ability to specifically diagnose TB-NSP) on adult EPTB mortality depends most strongly on factors that influence transmission, with settings of higher transmission (e.g., higher per-person transmission rate, lower diagnostic rate) favoring POC sputum NAAT., Conclusion: Although novel tests for pediatric TB and EPTB are likely to reduce TB mortality, major reductions in pediatric and EPTB incidence and mortality also require better diagnostic tests for adult pulmonary TB that reach a larger population.

Xpert MTB/RIF (Cepheid, Sunnyvale, CA, USA) is endorsed for the detection of pulmonary tuberculosis (TB). We performed a systematic review and meta-analysis to assess the accuracy of Xpert for the detection of extrapulmonary TB. We searched multiple databases to October 15, 2013. We determined the accuracy of Xpert compared with culture and a composite reference standard (CRS). We grouped data by sample type and performed meta-analyses using a bivariate random-effects model. We assessed sources of heterogeneity using meta-regression for predefined covariates. We identified 18 studies involving 4461 samples. Sample processing varied greatly among the studies. Xpert sensitivity differed substantially between sample types. In lymph node tissues or aspirates, Xpert pooled sensitivity was 83.1% (95% CI 71.4-90.7%) versus culture and 81.2% (95% CI 72.4-87.7%) versus CRS. In cerebrospinal fluid, Xpert pooled sensitivity was 80.5% (95% CI 59.0-92.2%) against culture and 62.8% (95% CI 47.7-75.8%) against CRS. In pleural fluid, pooled sensitivity was 46.4% (95% CI 26.3-67.8%) against culture and 21.4% (95% CI 8.8-33.9%) against CRS. Xpert pooled specificity was consistently >98.7% against CRS across different sample types. Based on this systematic review, the World Health Organization now recommends Xpert over conventional tests for diagnosis of TB in lymph nodes and other tissues, and as the preferred initial test for diagnosis of TB meningitis., (©ERS 2014.)

Background: Xpert MTB/RIF, the first automated molecular test for tuberculosis, is transforming the diagnostic landscape in low-income countries. However, little information is available on its performance in low-incidence, high-resource countries., Methods: We evaluated the accuracy of Xpert in a university hospital tuberculosis clinic in Montreal, Canada, for the detection of pulmonary tuberculosis on induced sputum samples, using mycobacterial cultures as the reference standard. We also assessed the potential reduction in time to diagnosis and treatment initiation., Results: We enrolled 502 consecutive patients who presented for evaluation of possible active tuberculosis (most with abnormal chest radiographs, only 18% symptomatic). Twenty-five subjects were identified to have active tuberculosis by culture. Xpert had a sensitivity of 46% (95% confidence interval [CI], 26%-67%) and specificity of 100% (95% CI, 99%-100%) for detection of Mycobacterium tuberculosis. Sensitivity was 86% (95% CI, 42%-100%) in the 7 subjects with smear-positive results, and 28% (95% CI, 10%-56%) in the remaining subjects with smear-negative, culture-positive results; in this latter group, positive Xpert results were obtained a median 12 days before culture results. Subjects with positive cultures but negative Xpert results had minimal disease: 11 of 13 had no symptoms on presentation, and mean time to positive liquid culture results was 28 days (95% CI, 25-47 days) compared with 14 days (95% CI, 8-21 days) in Xpert/culture-positive cases., Conclusions: Our findings suggest limited potential impact of Xpert testing in high-resource, low-incidence ambulatory settings due to lower sensitivity in the context of less extensive disease, and limited potential to expedite diagnosis beyond what is achieved with the existing, well-performing diagnostic algorithm.

Background: Multidrug-resistant tuberculosis (MDR-TB) is resistant to both rifampicin (RIF) and isoniazid (INH). Whereas many TB diagnostics detect RIF-resistance, few detect INH-monoresistance, which is common and may increase risk of acquired MDR-TB. Whether inclusion of INH-resistance in a first-line rapid test for TB would have an important impact on MDR-TB rates remains uncertain., Methods: WE DEVELOPED A TRANSMISSION MODEL TO EVALUATE THREE TESTS IN A POPULATION SIMILAR TO THAT OF INDIA: a rapid molecular test for TB, the same test plus RIF-resistance detection ("TB+RIF"), and detection of RIF and INH-resistance ("TB+RIF/INH"). Our primary outcome was the prevalence of INH-resistant and MDR-TB at ten years., Results: Compared to the TB test alone and assuming treatment of all diagnosed MDR cases, the TB+RIF test reduced the prevalence of MDR-TB among all TB cases from 5.5% to 3.8% (30.6% reduction, 95% uncertainty range, UR: 17-54%). Despite using liberal assumptions about the impact of INH-monoresistance on treatment outcomes and MDR-TB acquisition, expansion from TB+RIF to TB+RIF/INH lowered this prevalence only from 3.8% to 3.6% further (4% reduction, 95% UR: 3-7%) and INH-monoresistant TB from 15.8% to 15.1% (4% reduction, 95% UR: (-8)-19%)., Conclusion: When added to a rapid test for TB plus RIF-resistance, detection of INH-resistance has minimal impact on transmission of TB, MDR-TB, and INH-monoresistant TB.

Identification and treatment of latent tuberculosis infection (LTBI) can substantially reduce the risk of developing active disease. However, there is no diagnostic gold standard for LTBI. Two tests are available for identification of LTBI: the tuberculin skin test (TST) and the gamma interferon (IFN-γ) release assay (IGRA). Evidence suggests that both TST and IGRA are acceptable but imperfect tests. They represent indirect markers of Mycobacterium tuberculosis exposure and indicate a cellular immune response to M. tuberculosis. Neither test can accurately differentiate between LTBI and active TB, distinguish reactivation from reinfection, or resolve the various stages within the spectrum of M. tuberculosis infection. Both TST and IGRA have reduced sensitivity in immunocompromised patients and have low predictive value for progression to active TB. To maximize the positive predictive value of existing tests, LTBI screening should be reserved for those who are at sufficiently high risk of progressing to disease. Such high-risk individuals may be identifiable by using multivariable risk prediction models that incorporate test results with risk factors and using serial testing to resolve underlying phenotypes. In the longer term, basic research is necessary to identify highly predictive biomarkers.

Existing diagnostic tests for pleural tuberculosis (TB) have inadequate accuracy and/or turnaround time. Interferon-gamma (IFNg) has been identified in many studies as a biomarker for pleural TB. Our objective was to develop a lateral flow, immunochromatographic test (ICT) based on this biomarker and to evaluate the test in a clinical cohort. Because IFNg is commonly present in non-TB pleural effusions in low amounts, a diagnostic IFNg-threshold was first defined with an enzyme-linked immunosorbent assay (ELISA) for IFNg in samples from 38 patients with a confirmed clinical diagnosis (cut-off of 300 pg/ml; 94% sensitivity and 93% specificity). The ICT was then designed; however, its achievable limit of detection (5000 pg/ml) was over 10-fold higher than that of the ELISA. After several iterations in development, the prototype ICT assay for IFNg had a sensitivity of 69% (95% confidence interval (CI): 50-83) and a specificity of 94% (95% CI: 81-99%) compared to ELISA on frozen samples. Evaluation of the prototype in a prospective clinical cohort (72 patients) on fresh pleural fluid samples, in comparison to a composite reference standard (including histopathological and microbiologic test results), showed that the prototype had 65% sensitivity (95% CI: 44-83) and 89% specificity (95% CI: 74-97). Discordant results were observed in 15% of samples if testing was repeated after one freezing and thawing step. Inter-rater variability was limited (3%; 1 out of 32). In conclusion, despite an iterative development and optimization process, the performance of the IFNg ICT remained lower than what could be expected from the published literature on IFNg as a biomarker in pleural fluid. Further improvements in the limit of detection of an ICT for IFNg, and possibly combination of IFNg with other biomarkers such as adenosine deaminase, are necessary for such a test to be of value in the evaluation of pleural tuberculosis.

The use of mobile phones has substantially increased throughout the world over the last decade. This has opened up opportunities for the integration of mobile phones as health intervention tools in many aspects of health care, including prevention, diagnosis, data collection, treatment and adherence monitoring and surveillance. Several applications have been explored in human immunodeficiency virus care. The field of tuberculosis (TB) has not exploited the potential of mobile health (m-health) to the same extent, although the opportunities have been recognized. A number of proof-of-concept and pilot studies have been published on m-health in TB care, and an even larger number of studies are available in the grey literature. This article summarizes publications and recent developments at the intersection of TB care and m-health. We show that more rigorous studies evaluating different applications and implementation strategies are needed to establish an evidence base that serves to inform policy and decision making. We outline further areas of research that should be addressed and potential challenges that lie ahead if m-health applications are to enhance the accessibility and quality of TB care.

Few studies have correlated the results of interferon (gamma interferon) release assays (IGRAs) with known markers of tuberculosis (TB) treatment response. We report the results of serial QuantiFERON-TB gold in-tube assay (QFT) testing on 149 patients with active tuberculosis and correlate the results with smear and culture conversion. We show that QFT results do not offer much value for treatment monitoring of TB disease.

Resistance to antimicrobials continues to increase worldwide. Data suggest that older patients are among the main reservoirs of multidrug-resistant organisms (MDROs) in the hospital. We hypothesized that older patients (≥ 65 years of age) are more likely to harbor MDRO at hospital admission. We compared rates of methicillin-resistant Staphylococcus aureus (MRSA), vancomycin-resistant enterococci (VRE) and multidrug-resistant gram-negative bacteria (MDRGN) recovered from clinical cultures within the first 48 h of admission to an adult acute care hospital between the elderly (≥ 65 years old) and young per 1000 age-stratified admissions over a 12-year study period. Trends in antimicrobial resistance, sites of recovery and species for MDRGN were also characterized. An average of 7534 positive bacterial cultures were collected per year. The admission prevalence per 1000 age-stratified admissions was consistently higher among the elderly for all three MDRO under investigation. Among the elderly, the admission prevalence increased significantly for VRE (0.89 in 1998 to 3.62 in 2009 per 1000 admissions; p < 0.001) and MDRGN (1.41 in 1998 to 11.33 in 2009 per 1000 admissions; p < 0.001). Percentage resistant for all three MDRO increased as well. These data suggest that elderly patients are contributing substantially to the influx of MDRO into the hospital setting., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Laboratorians and clinicians often rely on package inserts of diagnostic tests to assess their accuracy. We compared test accuracy for tuberculosis diagnostics reported in 19 package inserts against estimates in published meta-analyses and found that package inserts generally report overoptimistic accuracy estimates. However, package inserts of most tests approved by the U.S. Food and Drug Administration (FDA) or endorsed by the World Health Organization provide more realistic estimates that agree with meta-analyses.

Undiagnosed and mismanaged tuberculosis (TB) continues to fuel the global TB epidemic. Rapid, accurate and early diagnosis of TB is therefore a priority to improve TB case detection and interrupt transmission. Although considerable improvements have been made in TB diagnostics, there are two major gaps in the existing diagnostics pipeline: (1) lack of a simple accurate point-of-care test that can be used for rapid diagnosis at the primary care level; (2) lack of a biomarker (or combination of biomarkers) that can be used to identify latently infected individuals who will benefit most from preventive therapy. Currently available commercial serological (antibody detection) tests are inaccurate and do not improve patient outcomes. Despite this evidence, dozens of serological tests are sold and used in countries (e.g. India) with weak regulatory systems, especially in the private sector. Recognizing the threat posed by these suboptimal tests, a World Health Organization (WHO) Expert Group has strongly recommended against the use of serological tests for the diagnosis of pulmonary and extra-pulmonary TB. Another WHO Expert Group has discouraged the use of interferon-γ release assays for active pulmonary TB diagnosis in low- and middle-income countries. All existing tests for latent TB infection appear to have only modest predictive value and further research is needed to identify highly predictive biomarkers., (Copyright © 2011 S. Karger AG, Basel.)

Identification of latent tuberculosis (TB) infection and preventive therapy is important for TB control, especially in high-risk populations. Since the advent of interferon-γ release assays (IGRAs), many studies have evaluated their role in the diagnosis of active and latent TB. With the growing evidence base, many guidelines now include IGRAs. We surveyed the literature and contacted experts to identify 33 guidelines and position papers from 25 countries and two supranational organizations. The results show considerable diversity in the recommendations on IGRAs, with four approaches commonly proposed: (i) two-step approach of tuberculin skin test (TST) first, followed by IGRA either when the TST is negative (to increase sensitivity, mainly in immunocompromised individuals), or when the TST is positive (to increase specificity, mainly in bacillus Calmette-Guérin-vaccinated individuals); (ii) Either TST or IGRA, but not both; (iii) IGRA and TST together (to increase sensitivity); and (iv) IGRA only, replacing the TST. Overall, the use of IGRAs is increasingly recommended, but most of the current guidelines do not use objective, transparent methods to grade evidence and recommendations, and do not disclose conflicts of interests. Future IGRA guidelines must aim to be transparent, evidence-based, periodically updated, and free of financial conflicts and industry involvement., (2011 The Authors. Clinical Microbiology and Infection; 2011 European Society of Clinical Microbiology and Infectious Diseases.)

Cryptosporidium parvum causes severe long-standing diarrhea in immunocompromised patients. Sclerosing cholangitis caused by C. parvum is a rare complication in transplant recipients. We report herein the presentation of Cryptosporidium-associated cholangitis in an adult liver transplant patient diagnosed by liver biopsy. The patient improved on treatment with azithromycin and paromomycin.

Pertussis toxin (PTX) has potent immunologic adjuvant activity in vivo and concomitantly enhances both T helper type (Th1) and Th2 cytokine responses. The PTX-induced enhancement of Th1 and Th2 immunity is mediated via the activation of antigen presenting cells (APCs), but the underlying mechanism is not known. Here we asked whether the adjuvant activity of PTX on T cell immunity was mediated by cytokines and/or costimulatory signals. The results show that in vivo blockade of CD28-CD80/86 costimulation essentially abrogated PTX-mediated enhancement of antigen-specific Th1 and Th2 responses. Blockade of CD40L-CD40 interactions was less efficient in inhibiting PTX-mediated enhancement of Th1 and Th2 responses. In contrast, the adjuvant activity of PTX was not mediated via cytokines, because neither Th1 nor Th2 responses were substantially impaired in mice deficient for IL-12, IFN-gamma, IL-4, IL-5, or IL-6. Collectively, the data suggest that PTX mediates its adjuvant effects on T cell cytokine differentiation and clonal expansion via the modulation of costimulatory molecules on APCs. Understanding the costimulatory pathways targeted by PTX could lead to the design of novel adjuvants that selectively induce Th1 or Th2 immunity.

After several decades of research into the macrophage migration inhibitory factor (MIF), its diverse actions in the immune system are yet to be fully revealed. What has become clear is that MIF plays an important role in both innate and adaptive immunity. However, while several pathways mediating the function of MIF in the immune system have been established, its role in pathogenic states such as autoimmune diseases has remained unresolved. MIF has been implicated in different autoimmune diseases, including rheumatoid arthritis, glomerulonephritis, and multiple sclerosis, but knowledge about the underlying cellular and molecular mechanisms is just emerging. However, overall it appears that the inhibition of its proinflammatory action is likely to be a successful new therapeutic strategy for some autoimmune diseases, possibly by reducing the need for steroids. As more aspects of the role of this cytokine in the pathogenesis of autoimmune diseases are elucidated, better strategies to target it therapeutically can be expected.

Macrophage migration inhibitory factor (MIF) is a cytokine that plays a critical role in the regulation of macrophage effector functions and T cell activation. However, its role in the pathogenesis of T cell-mediated autoimmune diseases, such as experimental autoimmune encephalomyelitis (EAE), has remained unresolved. In this study, we report that anti-MIF Ab treatment of SJL mice with acute EAE improved the disease severity and accelerated the recovery. Furthermore, the anti-MIF treatment impaired the homing of neuroantigen-reactive pathogenic T cells to the CNS in a VCAM-1-dependent fashion. Interestingly, MIF blockade also decreased the clonal size of the neuroantigen-specific Th1 cells and increased their activation threshold. Taken together, the results demonstrate an important role for MIF in the pathogenesis of EAE/multiple sclerosis and suggest that MIF blockade may be a promising new strategy for the treatment of multiple sclerosis.