From Bench to Clinic: A Nitroreductase Rv3368c-Responsive Cyanine-Based Probe for the Specific Detection of Live Mycobacterium tuberculosis.

Tuberculosis (TB), characterized by high mortality and low diagnosis, is caused by a single pathogen, Mycobacterium tuberculosis (Mtb). Imaging tools that can be used to track Mtb without pre-labeling and to diagnose live Mtb in clinical samples can shorten the gap between bench and clinic, fuel the development of novel anti-TB drugs, strengthen TB prevention, and improve patient treatment. In this study, we report an unprecedented novel nitroreductase-responsive cyanine-based fluorescent probe (Cy3-NO2-tre) that rapidly and specifically labels Mtb and detects it in clinical samples. Cy3-NO2-tre generated fluorescence after activation by a specific nitroreductase, Rv3368c, which is conserved in the Mycobacteriaceae. Cy3-NO2-tre effectively imaged mycobacteria within infected host cells, tracked the infection process, and visualized Mycobacterium smegmatis being endocytosed by macrophages. Cy3-NO2-tre also detected Mtb in the sputum of patients with TB and exhibited excellent photostability. Furthermore, the Cy3-NO2-tre/auramine O percentage change within 7 ± 2 days post drug treatment in the sputum of inpatients was closely correlated with the reexamination results of the chest computed tomography, strongly demonstrating the clinical application of Cy3-NO2-tre as a prognostic indicator in monitoring the therapeutic efficacy of anti-TB drugs in the early patient care stage.

Multiple cytokine analysis based on QuantiFERON-TB gold plus in different tuberculosis infection status: an exploratory study.

BACKGROUND: More efficient and convenient diagnostic method is a desperate need to reduce the burden of tuberculosis (TB). This study explores the multiple cytokines secretion based on QuantiFERON-TB Gold Plus (QFT-Plus), and screens for optimal cytokines with diagnostic potential to differentiate TB infection status. METHODS: Twenty active tuberculosis (ATB) patients, fifteen patients with latent TB infection (LTBI), ten patients with previous TB and ten healthy controls (HC) were enrolled. Whole blood samples were collected and stimulated by QFT-Plus TB1 and TB2 antigens. The levels of IFN-γ, TNF-α, IL-2, IL-6, IL-5, IL-10, IP-10, IL-1Ra, CXCL-1 and MCP-1 in supernatant were measured by Luminex bead-based multiplex assays. The receiver operating characteristic curve was used to evaluate the diagnostic accuracy of cytokine for distinguishing different TB infection status. RESULTS: After stimulation with QFT-Plus TB1 and TB2 antigens, the levels of all cytokines, except IL-5 in TB2 tube, in ATB group were significantly higher than that in HC group. The levels of IL-1Ra concurrently showed the equally highest AUC for distinguishing TB infection from HC, followed by the levels of IP-10 in both TB1 tube and TB2 tube. Moreover, IP-10 levels displayed the largest AUC for distinguishing ATB patients from non-ATB patients. Meanwhile, the levels of IP-10 also demonstrated the largest AUC in both TB1 tube and TB2 tube for distinguishing ATB patients from LTBI. CONCLUSIONS: In addition to conventional detection of IFN-γ, measuring IP-10 and IL-1Ra based on QFT-Plus may have the more tremendous potential to discriminate different TB infection status.

A case report about a child with drug-resistant tuberculous meningitis.

BACKGROUND: Hematogenous disseminated tuberculosis predisposes to concurrent tuberculous meningitis (TBM), the most devastating and disabling form of tuberculosis. However, children often have atypical clinical symptoms, difficulty in specimen collection, low specimen content, and an increasing incidence of drug-resistant tuberculosis. Thus, the accurate diagnosis and timely treatment of childhood tuberculosis face monumental challenges. CASE PRESENTATION: The 14-year-old female presented to the hospital with intermittent fever, headache, and blurred vision. Her cerebrospinal fluid (CSF) showed a lymphocytic pleocytosis, an elevated protein level, and a decreased chloride level. And her CSF tested positive for TB-RNA. Xpert MTB/RIF detected Mycobacterium tuberculosis in her CSF, but the rifampin resistance test was unknown. Subsequently, her CSF culture was positive for Mycobacterium tuberculosis. The drug sensitivity test (DST) revealed resistance to isoniazid, rifampin, and fluoroquinolones. A computed tomography (CT) of the chest showed diffuse miliary nodules in both lungs. Intracranial enhanced magnetic resonance imaging (MRI) showed "multiple intensified images of the brain parenchyma, cisterns, and part of the meninges." The final diagnosis is miliary pulmonary tuberculosis and pre-extensive drug-resistant TBM. After 19 months of an oral, individualized antituberculosis treatment, she recovered with no significant neurological sequelae. CONCLUSION: For patients with miliary pulmonary tuberculosis, especially children, even if there are no typical clinical symptoms, it is necessary to know whether there is TBM and other conditions. Always look for the relevant aetiological basis to clarify whether it is drug-resistant tuberculosis. Only a rapid and accurate diagnosis and timely and effective treatment can improve the prognosis and reduce mortality and disability rates.

Clinical characteristics and predictive model of pulmonary tuberculosis patients with pulmonary fungal coinfection.

BACKGROUND: In clinical settings, pulmonary tuberculosis (PTB) patients were often found to have pulmonary fungal coinfection. This study aimed to assess the clinical characteristics of patients suffering from coinfection with TB and pulmonary fungal and construct a predictive model for evaluating the probability of pulmonary fungal coinfection in patients with pulmonary tuberculosis. METHODS: The present case-control study retrospectively collected information from 286 patients affected by PTB who received treatment from December 6,2016- December 6,2021 at Beijing Chest Hospital, Capital Medical University. As control subjects, patients with sex and address corresponding to those of the case subjects were included in the study in a ratio of 1:1. These 286 patients were randomly divided into the training and internal validation sets in a ratio of 3:1. Chi-square test and logistic regression analysis were performed for the training set, and a predictive model was developed using the selected predictors. Bootstrapping was performed for internal validation. RESULTS: Seven variables [illness course, pulmonary cavitation, broad-spectrum antibiotics use for at least 1 week, chemotherapy or immunosuppressants, surgery, bacterial pneumonia, and hypoproteinemia] were validated and used to develop a predictive model which showed good discrimination capability for both training set [area under the curve (AUC) = 0.860, 95% confidence interval (CI) = 0.811-0.909] and internal validation set (AUC = 0.884, 95% CI = 0.799-0.970). The calibration curves also showed that the probabilities predicted using the predictive model had satisfactory consistency with the actual probability for both training and internal validation sets. CONCLUSIONS: We developed a predictive model that can predict the probability of pulmonary fungal coinfection in pulmonary tuberculosis patients. It showed potential clinical utility.

Spontaneous Mutational Patterns and Novel Mutations for Delamanid Resistance in Mycobacterium tuberculosis.

Delamanid (DLM) and pretomanid (PTM) are recent additions to the anti-tuberculosis (TB) drug armamentarium, and they offer more effective options for drug-resistant TB treatment. In particular, DLM is included in Group C, which is recommended for use in longer multidrug-resistant (MDR)-TB regimens. Previous studies have shown that resistance to DLM/PTM is caused by mutations in the ddn, fgd1, fbiA, fbiB, fbiC, and fbiD genes, which are related to the F420-dependent bioactivation pathway. Herein, we conduct in vitro selection of DLM-resistant strains using clinical Mycobacterium tuberculosis (MTB) isolates with various drug resistance profiles. The spontaneous resistance frequency of drug-susceptible (DS) MTB (1.14 × 10-6 to 1.04 × 10-4) to DLM was similar to that of H37Rv (8.88 × 10-6 to 9.96 × 10-6) but higher than those of multidrug-resistant MTB (2.03 × 10-7 to 3.18 × 10-6) and extensively drug-resistant (XDR) MTB (4.67 × 10-8 to 3.60 × 10-6). Of the 100 independently selected DLM-resistant MTB mutants, 65% harbored mutations in genes associated with either DLM prodrug activation (ddn, 39.73%; fgd1, 16.44%) or the F420 biosynthetic pathway (fbiA, 16.44%; fbiB, 5.48%; fbiC, 21.92%). Of the 45 mutations we identified, 38 were not previously reported. A structure analysis revealed that several point mutations affected the ligand binding or structural stability of enzymes related to DLM resistance, which would block the enzyme activity required for prodrug activation. Our results elucidate the in vitro spontaneous DLM-resistance patterns of different clinical strains, which could improve the understanding of the causes of DLM resistance in clinical strains and of the effects on drug resistance of different mutations in genes that are related to the DLM activation pathway.

Development and Validation of a Nomogram for Prediction of QT Interval Prolongation in Patients Administered Bedaquiline-Containing Regimens in China: a Modeling Study.

Corrected QT duration (QTc) interval prolongation is the most frequent adverse event associated with bedaquiline (BDQ) use. It may affect the safety of antituberculosis therapy, which leads to the consequent demands of needing to monitor during therapy. Our objective was to establish and validate a prediction model for estimating the risk of QTc prolongation after initiation of BDQ-containing regimens to multidrug-resistant tuberculosis (MDR-TB) patients. We constructed an individualized nomogram model based on baseline demographic and clinical characteristics of each patient within a Chinese cohort during BDQ treatment. The generalizability of this model was further validated through use of externally acquired data obtained from Beijing Chest Hospital from 2019 to 2020. Overall, 1,215 and 165 patients were included in training and external validation cohorts, respectively, whereby during anti-TB drug treatment, QTc prolongation was observed in 273 (22.5%) and 29 (17.6%) patients within these respective cohorts, for whom QTc values were >500 ms in 86 (31.5%) and 10 (34.7%) patients, respectively. Next, a total of four Cox proportional hazards models were created and assessed; then, nomograms derived from the models were plotted based on independent predictors of clofazimine, baseline QTc interval, creatinine, extensive drug-resistance (XDR), moxifloxacin, levofloxacin, and sex. Nomogram analysis revealed concordance index values of 0.723 (95% confidence interval [CI], 0.695 to 0.750) for the training cohort and 0.710 (95% CI, 0.627 to 0.821) for the external validation cohort, thus indicating relatively fair agreement between predicted and observed probabilities of QTc prolongation occurrence based on data obtained during 8-week, 16-week, and 24-week anti-TB treatment of both cohorts. Taken together, results obtained using these models demonstrated that coadministration of clofazimine and abnormal baseline QTc interval significantly contributed to QTc prolongation development during MDR-TB patient treatment with a BDQ-containing anti-TB treatment regimen.

Reduced Susceptibility of Mycobacterium tuberculosis to Bedaquiline During Antituberculosis Treatment and Its Correlation With Clinical Outcomes in China.

BACKGROUND: We aimed to assess the proportion of multidrug-resistant tuberculosis (MDR-TB) cases with initial bedaquiline (BDQ) resistance, monitor the dynamics of BDQ susceptibility of Mycobacterium tuberculosis isolates during therapy, and correlate susceptibility with MDR-TB patient clinical outcomes in China. METHODS: A retrospective, cohort study of MDR-TB patients was conducted, with positive cultures collected from cases at 13 sites. Patients with nontuberculous mycobacterial infection during anti-TB therapy were excluded. BDQ minimal inhibitory concentrations (MICs) were determined using a 7H9 Middlebrook broth-based microdilution method. Mutations that conferred BDQ resistance were detected via Sanger sequencing. RESULTS: A total of 277 patients receiving BDQ treatment were studied, with BDQ resistance noted in isolates from 2.2% (6/277) of MDR-TB cases, sputum conversion observed in 5 cases, and culture conversion observed in 138 cases within 2 weeks. Another 15 and 30 isolates were excluded from final analysis due to failures in obtaining subcultures and serial isolates, respectively. Of 94 cases that yielded serial isolates, 11 exhibited reduced BDQ susceptibility, while 3 of 5 cases with acquired resistance failed to culture-convert. Sequence analysis revealed that 6 of 11 BDQ-resistant isolates harbored Rv0678 mutations; no mutations were detected in 3 other BDQ resistance-associated genes. No significant intergroup difference in culture conversion time was observed. CONCLUSIONS: MDR-TB patients in China exhibited a low initial BDQ resistance rate. MDR-TB cases with acquired BDQ resistance were at greater risk of treatment failure relative to initially BDQ-resistant cases. Rv0678 mutations accounted for BDQ resistance in this cohort.

Membrane Feature-Inspired Profiling of Extracellular Vesicles for Pancreatic Cancer Diagnosis.

Extracellular vesicles (EVs) have recently emerged as a promising tumor biomarker, and EV phenotyping offers many benefits for cancer diagnosis. However, the practicality of EV assays remains a challenge due to macromolecule disturbances, biomarker heterogeneities, and EV abundance limitations. Here, we demonstrate a membrane-based biosensor for precise and sensitive EV identification. The sensor synergistically integrates EV capture and detection by virtue of EV membrane features (membrane protein and lipid bilayer), comprising antibody-conjugated magnetic beads (AbMBs) and duplex-specific nuclease (DSN)-mediated amplification cycles. Bivalent cholesterol (biChol)-modified RNA-DNA duplexes are designed to insert into the EV membrane, transforming EV signals into RNA signals and initiating the signal amplification. The membrane-based signal production pattern eliminates protein interference. By employing four antibodies specific to PCa-related membrane proteins, the AbMB-biChol platform enables the successful differentiation and monitoring of PCa-related EVs and distinguishes PCa patients from healthy donors with improved efficacy, exhibiting superior efficiency over the analyses based on clinically used biomarker CA19-9 and PCa-related proteins. As such, the developed system has great potential for clinical PCa diagnosis.

Interdigital electrodes of air@NiO porous nanoshells for high performance microsupercapacitors by thermally-assisted 3D printing.

Microsupercapacitors of air@NiO porous nanoshells are manufactured by a novel thermally-assisted 3D printing process. It entails the use of printing inks of the moderate solid content of CNT-PS@Ni-precursor-nanoparticle mixture, a real-time heating substrate to print 3D interdigital electrodes, and subsequent thermal annealing to convert PS@Ni-precursor particles into air@NiO porous nanoshells. The microstructure of 3D printed electrodes is characterized by air@NiO porous nanoshells being well dispersed in the CNT network. The CNT network provides a fast electronic migration path and meanwhile ensures the mechanical integrity of electrodes to prevent the fracture and/or collapsing of electrode structures during 3D printing manufacturing and charging/discharging cycles. The air@NiO porous nanoshells, manufactured in our labs, consist of randomly oriented nanosheets and offer superb charge storage via redox reactions. The metal layer is sputtered indiscriminately on the surface of interdigital electrodes and substrate before it is peeled off with electrolyte film and electrodes. The proposed tactic resolves problems connected with the tedious courses of traditional lithography and the delamination at the interface of active materials and collectors from mechanical stress. Experiments were conducted to study the performance of the microsupercapacitors (i.e. areal capacitances, energy and power densities) as a function of printing parameters, such as electrode heights, embedded amount of air@NiO porous nanoshells and the thickness of the metal layer on the electrochemical characteristics. The thickness of as-printed electrodes reaches up to 117 µm, which is vital in ensuring high energy density and is beyond the reach of any other technology. Moreover, the 3D printedmicrosupercapacitors of air@NiO porous nanoshells show excellent cycle stability and deliver an excellent areal capacitance of 56.7 mF cm-2, about a magnitude or two higher than that of C-based counterparts.

Epidemiology of Extrapulmonary Tuberculosis among Inpatients, China, 2008-2017.

We investigated the epidemiology of extrapulmonary tuberculosis (TB) among patients admitted to Beijing Chest Hospital, Beijing, China, during January 2008-December 2017. Of 19,279 hospitalized TB patients, 33.4% (6,433) had extrapulmonary TB and 66.6% (12,846) had pulmonary TB. The most frequent forms of extrapulmonary TB observed were skeletal TB (41.1%) and pleural TB (26.0%). Younger, female patients from rural areas were more likely to have extrapulmonary TB. However, patients with diabetes mellitus were less likely to have extrapulmonary TB compared with patients without diabetes. A higher proportion of multidrug-resistant (MDR) TB was observed among patients with extrapulmonary TB than among patients with pulmonary TB. We observed a large increase in MDR TB, from 17.3% to 35.7%, for pleural TB cases. The increasing rate of drug resistance among extrapulmonary TB cases highlights the need for drug susceptibility testing and the formulation of more effective regimens for extrapulmonary TB treatment.

Efficacy and safety of magnesium isoglycyrrhizinate injection in patients with acute drug-induced liver injury: A phase II trial.

BACKGROUND: Drug-induced liver injury (DILI) is the most common reason for a drug to be withdrawn from the market. Apart from stopping the offending drug, no regimens are available for treating idiosyncratic DILI in clinical practice. METHODS: We carried out a randomized, double-blind, multidoses, active drug controlled, multicentre phase II trial to assess the safety and efficacy of the study drug, magnesium isoglycyrrhizinate (MgIG), as compared to tiopronin, a standard therapy for DILI in China. The primary outcome was the proportion of alanine aminotransferase (ALT) normalization at week 4 after study drug administration. Logistic regression was used to examine the odds of ALT normalization between low dose (Group A) and high dose (Group B) vs active control (Group C). RESULTS: One hundred and seventy-four eligible subjects were randomized and enrolled into three groups: 59 in group A, 56 in group B and 59 in group C. It was shown that group A and group B lowered ALT level even at early stage of study drug administration; when compared with Group C (61.02%), the proportions of ALT normalization at week 4 were significantly greater in Group A (84.75%, P = .0029) and Group B (85.71%, P = .0037) respectively. The results from the univariate logistic model showed that the odds of ALT normalized among subjects in Group A were about 3.6 times greater (OR = 3.55, 95% CI: 1.47-8.57, P = .0049) than subjects in Group C. Similar effect was observed among subjects in Group B (OR = 3.83, 95% CI: 1.54-9.55, P = .0039). CONCLUSIONS: This trial provided preliminary evidence that MgIG is an effective and safe treatment for patients with acute DILI.

GeneXpert of stool versus gastric lavage fluid for the diagnosis of pulmonary tuberculosis in severely ill adults.

PURPOSE: Stool is an alternative specimen matrix for tuberculosis (TB) tests, because Mycobacterium tuberculosis (MTB) can be swallowed and detected in the samples from digestive tract. We aimed to assess the performance of GeneXpert on stool and gastric lavage fluid (GALF) in diagnosing TB among patients with severe pulmonary TB. METHODS: We enrolled adults with suspected pulmonary TB who were unable to produce sputum at visit between January 2016 and June 2018. Bacteriological samples consisted of one transtracheal aspirate sputum specimen, one stool specimen and/or one gastric lavage fluid specimen. Bacterial culture of transtracheal aspirate sputum provided the gold standard. RESULTS: Of 65 individuals recruited for analysis, MGIT culture identified the presence of MTB in 32 samples. Overall, 29 of 32 stool samples from culture-positive cases were detected by the GeneXpert test, demonstrating a sensitivity of 90.6%. For GALF, 13 patients were detected as infected with MTB by GeneXpert, yielding a sensitivity of 56.5%. The statistical analysis revealed that GeneXpert showed significantly better sensitivity in detecting MTB from stool samples than GALF samples (P = 0.003). Among individuals with GeneXpert-positive stool, the percentage of individuals with comorbid diabetes was significantly higher than among individuals with GeneXpert-negative stool (19.4% vs. 2.9%, P = 0.034). CONCLUSIONS: In conclusion, our data reveal that GeneXpert provides a higher detection rate on stool compared to GALF, indicating stool should be considered as an alternative for adult TB patients unable to produce sputum. Individuals with diabetes are more likely to have positive GeneXpert stool than nondiabetic individuals.

Implications of a school outbreak of multidrug-resistant tuberculosis in Northern China.

In this study, we identified a multidrug-resistant tuberculosis (MDR-TB) outbreak in a high school in northern China. The aim of this work was to describe TB transmission, drug resistance and treatment outcomes for this patient cluster. In January 2017, pulmonary TB was identified in a 17-year-old boy in northern China. Subsequently, a total of 11 TB cases were identified during 6-month follow-up of attendees of the same school. Of five students with latent TB infection (LTBI) receiving isoniazid preventive therapy (IPT), two pulmonary TB cases (40.0%) emerged in March and April, for an active case rate not significantly different from that of the non-IPT group (4/16, 25.0%, P = 0.598). All TB patients were first treated with a standardised first-line treatment regimen administered by the local TB hospital, with 11 of 12 active TB patients exhibiting poor treatment outcomes. Further data demonstrated that all nine patient isolates collected during this outbreak were MDR-TB and shared a common genotypic profile. In conclusion, our data demonstrate that diagnostic delay for the index MDR-TB case of this outbreak played a primary role in transmission of MDR-TB infection within a school setting. Importantly, IPT failed to prevent progression of MDR-TB from LTBI to active TB.

Efficacy of ultra-short course chemotherapy for new smear positive drug susceptible pulmonary tuberculosis: study protocol of a multicenter randomized controlled clinical trial.

BACKGROUND: Shortening the standard 6-month treatment for drug-susceptible pulmonary tuberculosis (DS-PTB) would be a major improvement for TB case management and disease control. METHODS: We are conducting a randomized, open-label, controlled, non-inferiority trial involving patients with smear-positive, newly diagnosed DS-PTB cases nationwide to assess the efficacy and safety of two 4.5- month regimens in comparison to the standard 6-month WHO recommended regimen. The regimen used in one experiment group is a 4.5-month fluoroquinolone-containing regimen, which consists of full course of levofloxacin, isoniazid (H), rifampin (R), parazinamid (Z) and ethambutol (E). Regimen used in the second experiment group includes 4.5-month full course of H, R, Z, E with levofloxacin removed. Patients in the control group, receive H, R, Z and E for 2 months, followed by 4 months of H and R. The primary endpoint is treatment failure or relapse within 24 month after treatment completion. DISCUSSION: Results from this trial along with other studies will contribute to the science of constructing a shorter, effective and safe regiment for TB patients. TRIAL REGISTRATION: The protocol has been registered on ClinicalTrials.gov on 2 September,2016 with identifier NCT02901288 .

Building clinical trial capacity to develop a new treatment for multidrug-resistant tuberculosis.

PROBLEM: New drugs for infectious diseases often need to be evaluated in low-resource settings. While people working in such settings often provide high-quality care and perform operational research activities, they generally have less experience in conducting clinical trials designed for drug approval by stringent regulatory authorities. APPROACH: We carried out a capacity-building programme during a multi-centre randomized controlled trial of delamanid, a new drug for the treatment of multidrug-resistant tuberculosis. The programme included: (i) site identification and needs assessment; (ii) achieving International Conference on Harmonization - Good Clinical Practice (ICH-GCP) standards; (iii) establishing trial management; and (iv) increasing knowledge of global and local regulatory issues. LOCAL SETTING: Trials were conducted at 17 sites in nine countries (China, Egypt, Estonia, Japan, Latvia, Peru, the Philippines, the Republic of Korea and the United States of America). Eight of the 10 sites in low-resource settings had no experience in conducting the requisite clinical trials. RELEVANT CHANGES: Extensive capacity-building was done in all 10 sites. The programme resulted in improved local capacity in key areas such as trial design, data safety and monitoring, trial conduct and laboratory services. LESSONS LEARNT: Clinical trials designed to generate data for regulatory approval require additional efforts beyond traditional research-capacity strengthening. Such capacity-building approaches provide an opportunity for product development partnerships to improve health systems beyond the direct conduct of the specific trial.

Delamanid for multidrug-resistant pulmonary tuberculosis.

BACKGROUND: Delamanid (OPC-67683), a nitro-dihydro-imidazooxazole derivative, is a new antituberculosis medication that inhibits mycolic acid synthesis and has shown potent in vitro and in vivo activity against drug-resistant strains of Mycobacterium tuberculosis. METHODS: In this randomized, placebo-controlled, multinational clinical trial, we assigned 481 patients (nearly all of whom were negative for the human immunodeficiency virus) with pulmonary multidrug-resistant tuberculosis to receive delamanid, at a dose of 100 mg twice daily (161 patients) or 200 mg twice daily (160 patients), or placebo (160 patients) for 2 months in combination with a background drug regimen developed according to World Health Organization guidelines. Sputum cultures were assessed weekly with the use of both liquid broth and solid medium; sputum-culture conversion was defined as a series of five or more consecutive cultures that were negative for growth of M. tuberculosis. The primary efficacy end point was the proportion of patients with sputum-culture conversion in liquid broth medium at 2 months. RESULTS: Among patients who received a background drug regimen plus 100 mg of delamanid twice daily, 45.4% had sputum-culture conversion in liquid broth at 2 months, as compared with 29.6% of patients who received a background drug regimen plus placebo (P=0.008). Likewise, as compared with the placebo group, the group that received the background drug regimen plus 200 mg of delamanid twice daily had a higher proportion of patients with sputum-culture conversion (41.9%, P=0.04). The findings were similar with assessment of sputum-culture conversion in solid medium. Most adverse events were mild to moderate in severity and were evenly distributed across groups. Although no clinical events due to QT prolongation on electrocardiography were observed, QT prolongation was reported significantly more frequently in the groups that received delamanid. CONCLUSIONS: Delamanid was associated with an increase in sputum-culture conversion at 2 months among patients with multidrug-resistant tuberculosis. This finding suggests that delamanid could enhance treatment options for multidrug-resistant tuberculosis. (Funded by Otsuka Pharmaceutical Development and Commercialization; ClinicalTrials.gov number, NCT00685360.).

[Evaluation of T-SPOT.TB assay in the diagnosis of pulmonary tuberculosis within different age groups].

OBJECTIVE: To evaluate the value of T-SPOT.TB assay in the diagnosis of pulmonary tuberculosis within different age groups. METHODS: We analyzed 1 518 suspected pulmonary tuberculosis (PTB) patients who were admitted to the Beijing Chest Hospital from November 2012 to February 2014 and had valid T-SPOT.TB tests before anti-tuberculosis therapy. The 599 microbiologically and/or histopathologically-confirmed PTB patients (16-89 years old, 388 males and 211 females) and 235 non-TB patients (14-85 years old, 144 males and 91 females) were enrolled for the analysis of diagnostic performance of T-SPOT.TB, while patients with uncertain diagnosis or diagnosis based on clinical impression (n=684) were excluded from the analysis. The sensitivity, specificity, positive predictive value, negative predictive value, positive likelihood ratio, and negative likelihood ratio of the T-SPOT.TB were analyzed according to the final diagnosis. Furthermore, the diagnostic performance of T-SPOT.TB assay in the younger patients (14-59 years old) and elderly patients (60-89 years old) were also analyzed respectively. Categorical variables were compared by Pearson's Chi-square test, while continuous variables were compared by the Mann-Whitney U-test. RESULTS: The sensitivity, specificity, positive predictive value, negative predictive value, positive likelihood ratio, and negative likelihood ratio of the T-SPOT.TB in diagnosis of PTB were 90.1% (540/599), 65.5% (154/235), 86.9% (540/621), 72.3% (154/213), 2.61, and 0.15, respectively. The sensitivity and specificity of T-SPOT.TB assay were 92.6% (375/405) and 75.6% (99/131), respectively in the younger patients, and 85.0% (165/194), 52.9% (55/104) respectively in the elderly patients. The sensitivity and specificity of T-SPOT.TB assay in the younger patients were significantly higher than those in the elderly patients (P<0.01), and the spot forming cells in the younger PTB patients were significantly higher than in the elderly PTB patients [300 (126, 666)/10(6) PBMCs vs. 258 (79, 621)/10(6) PBMCs, P=0.037]. CONCLUSION: T-SPOT.TB is a promising test in the diagnosis of younger patients (14-59 years old) with suspected PTB, but the diagnostic performance in elderly patients (60-89 years old) is relatively reduced.

Specific Cytokines Analysis Incorporating Latency-Associated Antigens Differentiates Mycobacterium tuberculosis Infection Status: An Exploratory Study.

Introduction: Current immunologic methods cannot distinguish Mycobacterium tuberculosis (Mtb) infection statuses, especially to discriminate active tuberculosis (ATB) from latent tuberculosis infection (LTBI). This study explored the potential of latency-associated antigens (Rv1733cSLP and Rv2028c) and multifactorial cytokine detection to distinguish tuberculosis infection states. Methods: ATB patients (20), LTBI healthcare workers (25), fever patients (11), and healthy controls (10) were enrolled. Cytokine levels (IFN-γ, TNF-α, IL-2, IL-6, IP-10, IL-1Ra, CXCL-1, and MCP-1) were measured using Luminex with/without MTB-specific virulence factor and latency-associated antigens stimulation. Results: Without antigen stimulation, IL-6, IP-10, MCP-1, and IL-1Ra were higher in the ATB group than in the LTBI group (p<0.05), but no significant differences between the ATB group and the fever group. Stimulated with the four antigens, respectively, the cytokines, including IP-10Esat-6, IP-10CFP-10, IFN-γRv1733cSLP, IFN-γRv2028c, IL-6Esat-6, IL-6Rv1733cSLP, IL-6Rv2028c, IL-2Rv1733cSLP, IL-2 Rv2028c, IL-1RaEsat-6, IL-1RaCFP-10, IL-1RaRv2028c, CXCL-1Esat-6, CXCL-1CFP-10, CXCL-1Rv1733cSLP, CXCL-1Rv2028c, MCP-1Esat-6 and MCP-1CFP-10, demonstrated accurate discrimination between ATB and LTBI (p<0.05). Additive concentrations demonstrated significant secretion differences of IFN-γ, IP-10 and IL-2, primarily by virulence factors in ATB and latency-associated antigens in LTBI. Latency-associated antigens synergized with virulence factors, enhancing TH1-type cytokine diagnostic efficacy for discriminating ATB from LTBI, the AUC for TNF-α increased from 0.696 to 0.820 (p=0.038), IFN-γ increased from 0.806 to 0.962 (p=0.025), and IL-2 increased from 0.565 to 0.868 (p=0.007). Model selected by forward likelihood method indicated combined detection of IFN-γCFP-10, IFN-γRv1733cSLP, IP-10Rv1733cSLP, and CXCL-1Rv1733cSLP achieved ATB diagnosis (AUC=0.996) and ATB-LTBI differentiation (AUC=0.992). Combined detection of IFN-γCFP-10 and IFN-γRv1733cSLP achieved tuberculosis infection diagnosis (AUC=0.943). Conclusion: Latency-associated antigens enhance multiple cytokine discriminatory ability, particularly TH1-type cytokines, for differentiating Mtb infection statuses.

A pragmatic randomized controlled trial to evaluate the efficacy and safety of an oral short-course regimen including bedaquiline for the treatment of patients with multidrug-resistant tuberculosis in China: study protocol for PROSPECT.

INTRODUCTION: The lack of safe, effective, and simple short-course regimens (SCRs) for multidrug-resistant/rifampicin-resistant tuberculosis (MDR/RR-TB) treatment has significantly impeded TB control efforts in China. METHODS: This phase 4, randomized, open-label, controlled, non-inferiority trial aims to assess the efficacy and safety of a 9-month all-oral SCR containing bedaquiline (BDQ) versus an all-oral SCR without BDQ for adult MDR-TB patients (18-65 years) in China. The trial design mainly mirrors that of the "Evaluation of a Standardized Treatment Regimen of Anti-Tuberculosis Drugs for Patients with MDR-TB" (STREAM) stage 2 study, while also incorporating programmatic data from South Africa and the 2019 consensus recommendations of Chinese MDR/RR-TB treatment experts. Experimental arm participants will receive a modified STREAM regimen C that replaces three group C drugs, ethambutol (EMB), pyrazinamide (PZA), and prothionamide (PTO), with two group B drugs, linezolid (LZD) and cycloserine (CS), while omitting high-dose isoniazid (INH) for confirmed INH-resistant cases. BDQ duration will be extended from 6 to 9 months for participants with Mycobacterium tuberculosis-positive sputum cultures at week 16. The control arm will receive a modified STREAM regimen B without high-dose INH and injectable kanamycin (KM) that incorporates experimental arm LZD and CS dosages, treatment durations, and administration methods. LZD (600 mg) will be given daily for ≥ 24 weeks as guided by observed benefits and harm. The primary outcome measures the proportion of participants with favorable treatment outcomes at treatment completion (week 40), while the same measurement taken at 48 weeks post-treatment completion is the secondary outcome. Assuming an α = 0.025 significance level (one-sided test), 80% power, 15% non-inferiority margin, and 10% lost to follow-up rate, each arm requires 106 participants (212 total) to demonstrate non-inferiority. DISCUSSION: PROSPECT aims to assess the safety and efficacy of a BDQ-containing SCR MDR-TB treatment at seventeen sites across China, while also providing high-quality data to guide SCRs administration under the direction of the China National Tuberculosis Program for MDR-TB. Additionally, PROSPECT will explore the potential benefits of extending the administration of the 9-month BDQ-containing SCR for participants without sputum conversion by week 16. TRIAL REGISTRATION: ClinicalTrials.gov NCT05306223. Prospectively registered on 16 March 2022 at https://clinicaltrials.gov/ct2/show/NCT05306223?term=NCT05306223&draw=1&rank=1 {2}.

Antigen-specific chemokine profiles as biomarkers for detecting Mycobacterium tuberculosis infection.

Background: Latent tuberculosis (TB) infection can progress to active TB, which perpetuates community transmission that undermines global TB control efforts. Clinically, interferon-γ release assays (IGRAs) are commonly used for active TB case detection. However, low IGRA sensitivity rates lead to false-negative results for a high proportion of active TB cases, thus highlighting IGRA ineffectiveness in differentiating MTB-infected individuals from healthy individuals. Methods: Participants enrolled at Beijing Chest Hospital from May 2020-April 2022 were assigned to healthy control (HC), LTBI, IGRA-positive TB, and IGRA-negative TB groups. Screening cohort MTB antigen-specific blood plasma chemokine concentrations were measured using Luminex xMAP assays then were verified via testing of validation cohort samples. Results: A total of 302 individuals meeting study inclusion criteria were assigned to screening and validation cohorts. Testing revealed significant differences in blood plasma levels of CXCL9, CXCL10, CXCL16, CXCL21, CCL1, CCL19, CCL27, TNF-α, and IL-4 between IGRA-negative TB and HC groups. Levels of CXCL9, CXCL10, IL-2, and CCL8 biomarkers were predictive for active TB, as reflected by AUC values of ≥0.9. CXCL9-based enzyme-linked immunosorbent assay sensitivity and specificity rates were 95.9% (95%CI: 91.7-98.3) and 100.0% (92.7-100.0), respectively. Statistically similar AUC values were obtained for CXCL9 and CXCL9-CXCL10 assays, thus demonstrating that combined analysis of CXCL10 and CXCL9 levels did not improve active TB diagnostic performance. Conclusion: The MTB antigen stimulation-based CXCL9 assay may compensate for low IGRA diagnostic accuracy when used to diagnose IGRA-negative active TB cases and thus is an accurate and sensitive alternative to IGRAs for detecting MTB infection.