Tuberculosis care models for children and adolescents: a scoping review.

Objective: To map which tuberculosis care models are best suited for children and adolescents. Methods: We conducted a scoping review to assess the impact of decentralized, integrated and family-centred care on child and adolescent tuberculosis-related outcomes, describe approaches for these care models and identify key knowledge gaps. We searched seven literature databases on 5 February 2021 (updated 16 February 2022), searched the references of 18 published reviews and requested data from ongoing studies. We included studies from countries with a high tuberculosis burden that used a care model of interest and reported tuberculosis diagnostic, treatment or prevention outcomes for an age group < 20 years old. Findings: We identified 28 studies with a comparator group for the impact assessment and added 19 non-comparative studies to a qualitative analysis of care delivery approaches. Approaches included strengthening capacity in primary-level facilities, providing services in communities, screening for tuberculosis in other health services, co-locating tuberculosis and human immunodeficiency virus treatment, offering a choice of treatment location and providing social or economic support. Strengthening both decentralized diagnostic services and community linkages led to one-to-sevenfold increases in case detection across nine studies and improved prevention outcomes. We identified only five comparative studies on integrated or family-centred care, but 11 non-comparative studies reported successful treatment outcomes for at least 71% of children and adolescents. Conclusion: Strengthening decentralized services in facilities and communities can improve tuberculosis outcomes for children and adolescents. Further research is needed to identify optimal integrated and family-centred care approaches.

Expanding Latent Tuberculosis Infection Testing and Treatment in Massachusetts Primary Care Clinics via the ECHO Model.

INTRODUCTION/OBJECTIVES: In the US, reactivation of latent tuberculosis infection (LTBI) accounts for 80% of new cases. In 2016, the US Preventive Services Task Force provided a new recommendation that primary care providers (PCPs) should conduct LTBI screening, whereas in the past, LTBI cases were evaluated and treated by specialty providers. This shift in care revealed knowledge gaps surrounding LTBI treatment among PCPs. This study assessed changes in PCPs' confidence for performing key aspects of LTBI care before and after participation in an LTBI Extension for Community Healthcare Outcomes (ECHO) course. METHODS: The ECHO Model™ is an evidence-based telementoring intervention. Participants were primary care team members from clinics throughout Massachusetts who voluntarily enrolled in the ECHO course. In this mixed-methods evaluation, primary outcomes were PCP self-reported confidence changes by pre- and post-course surveys and post-course semi-structured interviews. RESULTS: Twenty PCPs (43% of registered PCPs) attended at least 3 of the 6 sessions and 24 PCPs (31% of registered PCPs) completed at least one survey. Confidence increased in selecting a test (P = .004), interpreting tuberculosis infection test results (P = .03), and selecting a treatment regimen (P = .004). Qualitative interviews with 3 PCPs revealed practice changes including switching to interferon gamma release assays for testing and using rifampin for treatment. CONCLUSIONS: Use of the ECHO model to train PCPs in LTBI management is feasible and efficacious. For continuing medical education, ECHO courses can be leveraged to reduce health disparities in settings where PCPs' lack of familiarity about a treatment topic contributes to poor health outcomes.

Effective Host-Directed Therapy for Tuberculosis by Depletion of Myeloid-Derived Suppressor Cells and Related Cells Using a Diphtheria Toxin Fusion Protein.

Myeloid-derived suppressor cells (MDSCs) are present in elevated numbers in tuberculosis patients and have been found to be permissive for Mycobacterium tuberculosis proliferation. To determine whether depletion of MDSCs may improve host control of tuberculosis, we used a novel diphtheria toxin-based fusion protein DABIL-4 that targets and depletes interleukin 4 (IL-4) receptor-positive cells. We show that DABIL-4 depletes both polymorphonuclear MDSCs and monocytic MDSCs, increases interferon-γ + T cells, and reduces the lung bacillary burden in a mouse tuberculosis model. These results indicate that MDSC-depleting therapies targeting the IL-4 receptor are beneficial in tuberculosis and offer an avenue towards host-directed tuberculosis therapy.

Dual mTORC1/mTORC2 Inhibition as a Host-Directed Therapeutic Target in Pathologically Distinct Mouse Models of Tuberculosis.

Efforts to develop more effective and shorter-course therapies for tuberculosis have included a focus on host-directed therapy (HDT). The goal of HDT is to modulate the host response to infection, thereby improving immune defenses to reduce the duration of antibacterial therapy and/or the amount of lung damage. As a mediator of innate and adaptive immune responses involved in eliminating intracellular pathogens, autophagy is a potential target for HDT in tuberculosis. Because Mycobacterium tuberculosis modulates mammalian target of rapamycin (mTOR) signaling to impede autophagy, pharmacologic mTOR inhibition could provide effective HDT. mTOR exists within two distinct multiprotein complexes, mTOR complex-1 (mTORC1) and mTOR complex-2 (mTORC2). Rapamycin and its analogs only partially inhibit mTORC1. We hypothesized that novel mTOR kinase inhibitors blocking both complexes would have expanded therapeutic potential. We compared the effects of two mTOR inhibitors, rapamycin and the orally available mTOR kinase domain inhibitor CC214-2, which blocks both mTORC1 and mTORC2, as adjunctive therapies against murine TB when added to the first-line regimen (isoniazid, rifampin, pyrazinamide, and ethambutol [RHZE]) or the novel bedaquiline-pretomanid-linezolid (BPaL) regimen. Neither mTOR inhibitor affected lung CFU counts after 4 to 8 weeks of treatment when combined with BPaL or RHZE. However, addition of CC214-2 to BPaL and RHZE was associated with significantly fewer relapses in C3HeB/FeJ mice compared to addition of rapamycin and, in RHZE-treated mice, resulted in fewer relapses than RHZE alone. Therefore, CC214-2 and related mTOR kinase inhibitors may be more effective candidates for HDT than rapamycin analogs and may have the potential to shorten the duration of TB treatment.

Short Communication: Genetic Variation in Human IL10 Proximal Promoter and Susceptibility to HIV-1 Infection in Mali, West Africa.

It is now recognized that to fully understand the role of host genetic variation on susceptibility to HIV-1 infection, investigations must be extended to African populations. We sought to determine if genetic variation in IL10 are associated with HIV-1 infection in a West African cohort in Mali. HIV-infected and -uninfected individuals were genotyped for three common single nucleotide polymorphisms (SNPs) located at positions -592 (C/A), -819 (C/T), and -1082 (G/A) of the IL10 promoter. We found that the ATA haplotype, which has been previously associated with low IL-10 expression, was the most represented in the cohort. Although we observed a trend toward an increased frequency of ATA/ATA carriage in HIV-infected compared with -uninfected individuals, the difference was not statistically significant. Similarly, individual IL10 SNPs were not significantly enriched in the HIV-infected group, suggesting that IL10 genetic variants are not associated with HIV-1 in this West African cohort from Mali.

The integrated stress response mediates necrosis in murine Mycobacterium tuberculosis granulomas.

The mechanism by which only some individuals infected with Mycobacterium tuberculosis develop necrotic granulomas with progressive disease while others form controlled granulomas that contain the infection remains poorly defined. Mice carrying the sst1-suscepible (sst1S) genotype develop necrotic inflammatory lung lesions, similar to human tuberculosis (TB) granulomas, which are linked to macrophage dysfunction, while their congenic counterpart (B6) mice do not. In this study we report that (a) sst1S macrophages developed aberrant, biphasic responses to TNF characterized by superinduction of stress and type I interferon pathways after prolonged TNF stimulation; (b) the late-stage TNF response was driven via a JNK/IFN-ß/protein kinase R (PKR) circuit; and (c) induced the integrated stress response (ISR) via PKR-mediated eIF2α phosphorylation and the subsequent hyperinduction of ATF3 and ISR-target genes Chac1, Trib3, and Ddit4. The administration of ISRIB, a small-molecule inhibitor of the ISR, blocked the development of necrosis in lung granulomas of M. tuberculosis-infected sst1S mice and concomitantly reduced the bacterial burden. Hence, induction of the ISR and the locked-in state of escalating stress driven by the type I IFN pathway in sst1S macrophages play a causal role in the development of necrosis in TB granulomas. Interruption of the aberrant stress response with inhibitors such as ISRIB may offer novel host-directed therapy strategies.

Cavitary tuberculosis: the gateway of disease transmission.

Tuberculosis continues to be a major threat to global health. Cavitation is a dangerous consequence of pulmonary tuberculosis associated with poor outcomes, treatment relapse, higher transmission rates, and development of drug resistance. However, in the antibiotic era, cavities are often identified as the most extreme outcome of treatment failure and are one of the least-studied aspects of tuberculosis. We review the epidemiology, clinical features, and concurrent standards of care for individuals with cavitary tuberculosis. We also discuss developments in the understanding of tuberculosis cavities as dynamic physical and biochemical structures that interface the host response with a unique mycobacterial niche to drive tuberculosis-associated morbidity and transmission. Advances in preclinical models and non-invasive imaging can provide valuable insights into the drivers of cavitation. These insights will guide the development of specific pharmacological interventions to prevent cavitation and improve lung function for individuals with tuberculosis.

Dynamic imaging in patients with tuberculosis reveals heterogeneous drug exposures in pulmonary lesions.

Tuberculosis (TB) is the leading cause of death from a single infectious agent, requiring at least 6 months of multidrug treatment to achieve cure1. However, the lack of reliable data on antimicrobial pharmacokinetics (PK) at infection sites hinders efforts to optimize antimicrobial dosing and shorten TB treatments2. In this study, we applied a new tool to perform unbiased, noninvasive and multicompartment measurements of antimicrobial concentration-time profiles in humans3. Newly identified patients with rifampin-susceptible pulmonary TB were enrolled in a first-in-human study4 using dynamic [11C]rifampin (administered as a microdose) positron emission tomography (PET) and computed tomography (CT). [11C]rifampin PET-CT was safe and demonstrated spatially compartmentalized rifampin exposures in pathologically distinct TB lesions within the same patients, with low cavity wall rifampin exposures. Repeat PET-CT measurements demonstrated independent temporal evolution of rifampin exposure trajectories in different lesions within the same patients. Similar findings were recapitulated by PET-CT in experimentally infected rabbits with cavitary TB and confirmed using postmortem mass spectrometry. Integrated modeling of the PET-captured concentration-time profiles in hollow-fiber bacterial kill curve experiments provided estimates on the rifampin dosing required to achieve cure in 4 months. These data, capturing the spatial and temporal heterogeneity of intralesional drug PK, have major implications for antimicrobial drug development.

Assessing whether isoniazid is essential during the first 14 days of tuberculosis therapy: a phase 2a, open-label, randomised controlled trial.

BACKGROUND: Clinical studies suggest that isoniazid contributes rapid bacterial killing during the initial two days of tuberculosis treatment but that isoniazid's activity declines significantly after day three. We conducted a 14-day phase IIa open label, randomized trial to assess the essentiality of isoniazid in standard tuberculosis therapy. METHODS: A total of 69 adults with newly diagnosed sputum-positive tuberculosis from the South African Western Cape region were enrolled and randomized to a four-arm parallel assignment model. Participants were followed for 14 days as inpatients at either the University of Cape Town Lung Institute or at the TASK Applied Science clinical research organization. All arms received standard daily rifampicin, ethambutol, and pyrazinamide but differed as follows: isoniazid only on days one and two (n=17), isoniazid on days one and two then moxifloxacin on days three through 14 (n=16), no isoniazid (n=18), and a control group that received isoniazid for all 14 days (standard therapy, n=18). The primary endpoint was the rate of colony forming unit (CFU) decline during the first 14 days of treatment. RESULTS: For 62 participants analyzed, the initial 14-day mean daily fall in log10 CFU (95% CI) was 0·14 (0·11, 0·18) for participants receiving isoniazid for two days only; 0·13 (0·09, 0·17) for participants receiving isoniazid for two days followed by moxifloxacin; 0·12 (0·08, 0·15) for those not receiving isoniazid; and 0·13 (0·09, 0·16) for the standard therapy group. CONCLUSIONS: The 14 day EBA for the combination rifampicin, ethambutol, and pyrazinamide was not significantly changed by the addition of isoniazid for the first two days or for the first 14 days of treatment. In a post hoc analysis, significantly higher day-two EBAs were observed for all groups among participants with higher baseline sputum CFUs. Our finding that INH does not contribute to EBA suggests that INH could be replaced with another drug during standard treatment to improve efficacy and decrease rates of resistance to first-line drugs. (Funded by the NIH AIDS Clinical Trial Groups and NIH; A5307 ClinicalTrials.gov number, NCT01589497).

Second-generation IL-2 receptor-targeted diphtheria fusion toxin exhibits antitumor activity and synergy with anti-PD-1 in melanoma.

Denileukin diftitox (DAB-IL-2, Ontak) is a diphtheria-toxin-based fusion protein that depletes CD25-positive cells including regulatory T cells and has been approved for the treatment of persistent or recurrent cutaneous T cell lymphoma. However, the clinical use of denileukin diftitox was limited by vascular leak toxicity and production issues related to drug aggregation and purity. We found that a single amino acid substitution (V6A) in a motif associated with vascular leak induction yields a fully active, second-generation biologic, s-DAB-IL-2(V6A), which elicits 50-fold less human umbilical vein endothelial cell monolayer permeation and is 3.7-fold less lethal to mice by LD50 analysis than s-DAB-IL-2. Additionally, to overcome aggregation problems, we developed a production method for the fusion toxin using Corynebacterium diphtheriae that secretes fully folded, biologically active, monomeric s-DAB-IL-2 into the culture medium. Using the poorly immunogenic mouse B16F10 melanoma model, we initiated treatment 7 days after tumor challenge and observed that, while both s-DAB-IL-2(V6A) and s-DAB-IL-2 are inhibitors of tumor growth, the capacity to treat with higher doses of s-DAB-IL-2(V6A) could provide a superior activity window. In a sequential dual-therapy study in tumors that have progressed for 10 days, both s-DAB-IL-2(V6A) and s-DAB-IL-2 given before checkpoint inhibition with anti-programmed cell death-1 (anti-PD-1) antibodies inhibited tumor growth, while either drug given as monotherapy had less effect. s-DAB-IL-2(V6A), a fully monomeric protein with reduced vascular leak, is a second-generation diphtheria-toxin-based fusion protein with promise as a cancer immunotherapeutic both alone and in conjunction with PD-1 blockade.

Matrix Metalloproteinase Inhibition in a Murine Model of Cavitary Tuberculosis Paradoxically Worsens Pathology.

Matrix metalloproteinases (MMPs) degrade extracellular matrix and are implicated in tuberculosis pathogenesis and cavitation. In particular, MMP-7 is induced by hypoxia and highly expressed around pulmonary cavities of Mycobacterium tuberculosis-infected C3HeB/FeJ mice. In this study, we evaluated whether administration of cipemastat, an orally available potent inhibitor of MMP-7, could reduce pulmonary cavitation in M. tuberculosis-infected C3HeB/FeJ mice. We demonstrate that, compared with untreated controls, cipemastat treatment paradoxically increases the frequency of cavitation (32% vs 7%; P = .029), immunopathology, and mortality. Further studies are needed to understand the role of MMP inhibitors as adjunctive treatments for pulmonary tuberculosis.

Diverse Cavity Types and Evidence that Mechanical Action on the Necrotic Granuloma Drives Tuberculous Cavitation.

Effacement of normal lung parenchyma by cavities is an important sequela of pulmonary tuberculosis. Despite its clinical significance, the pathogenesis of tuberculous cavitation is poorly understood, with controversy as to whether the fundamental mechanism involves matrix depletion, lipid pneumonia, or mechanical factors. In this study, a repetitive aerosol infection model using Mycobacterium tuberculosis was used to generate cavities in 20 New Zealand white rabbits. Serial computed tomography was performed to monitor cavity progression over 14 weeks. Three-dimensional reconstructions were compiled for each time point, allowing comprehensive four-dimensional cavity mapping. Terminally, cavities were processed for histopathology. Cavities progressed rapidly from areas of consolidation, and often showed a pattern of explosive growth followed by gradual contraction. Cavities formed preferentially in the caudodorsal lung fields, and frequently were subpleural. Cavitation was associated invariably with necrosis. Histomorphology showed four distinct cavity types that provide mechanistic clues and insight on early cavity development. Our study shows that cavitation is a highly dynamic process with preferential formation at sites of high mechanical stress. These findings suggest a model for the pathogenesis of tuberculous cavitation in which mechanical stress acts on the necrotic granuloma to produce acute tears in structurally weakened tissue, with subsequent air trapping and cavity expansion.

Pharmacokinetics of rifapentine and rifampin in a rabbit model of tuberculosis and correlation with clinical trial data.

In clinical trials of two rifamycin antibiotics (rifampin and rifapentine) for treating tuberculosis (TB), patients with cavitary lung lesions did not appear to derive benefit from rifapentine. Rifapentine was found not to outperform rifampin, despite a lower minimum inhibitory concentration against Mycobacterium tuberculosis in mouse models of TB. To understand these findings, we have developed a rabbit model of TB that reliably develops lung cavities with features similar to those of patients with pulmonary cavitary TB. After single or multiple doses of rifampin or rifapentine that produced human-equivalent plasma exposures, rabbits were sacrificed at different time points after dosing. We measured site-of-disease drug pharmacokinetics and tissue drug distribution. We used pharmacokinetic-pharmacodynamic (PK/PD) modeling to estimate drug penetration into different types of tubercular lesions. Both drugs penetrated rabbit lung cellular lesions, as well as the fibrotic cavity wall of cavitary lesions (penetration coefficients ≥1 compared to plasma). For the necrotic liquefied material inside cavitary lesions known as caseum (which contains high numbers of bacteria), the penetration coefficient was 1.0 for rifampin but only 0.25 for rifapentine. When estimates of site-of-disease drug PK were substituted into clinical PK/PD models, the relationship between site-of-action exposure and sputum culture conversion was significant (P < 10-7). We propose that poor penetration of rifapentine into lung cavitary lesions explains, in part, why rifapentine doses required to improve treatment outcomes in two phase 2 clinical trials were four times higher in TB patients with large cavities compared to TB patients without cavitary lung disease.

Repetitive Aerosol Exposure Promotes Cavitary Tuberculosis and Enables Screening for Targeted Inhibitors of Extensive Lung Destruction.

Background: Cavitation is a serious consequence of tuberculosis. We tested the hypothesis that repetitive exposure to the same total bacterial burden of Mycobacterium tuberculosis drives greater lung destruction than a single exposure. We also tested whether inhibition of endogenous matrix metalloproteinase-1 (MMP-1) may inhibit cavitation during tuberculosis. Methods: Over a 3-week interval, we infected rabbits with either 5 aerosols of 500 colony-forming units (CFU) of M. tuberculosis or a single aerosol of 2500 CFU plus 4 sham aerosols. We administered the MMP-1 inhibitor cipemastat (100 mg/kg daily) during weeks 5-10 to a subset of the animals. Results: Repetitive aerosol infection produced greater lung inflammation and more cavities than a single aerosol infection of the same bacterial burden (75% of animals vs 25%). Necropsies confirmed greater lung pathology in repetitively exposed animals. For cipemastat-treated animals, there was no significant difference in cavity counts, cavity volume, or disease severity compared to controls. Conclusions: Our data show that repetitive aerosol exposure with M. tuberculosis drives greater lung damage and cavitation than a single exposure. This suggests that human lung destruction due to tuberculosis may be exacerbated in settings where individuals are repeatedly exposed. MMP-1 inhibition with cipemastat did not prevent the development of cavitation in our model.

Pathogenesis and Animal Models of Post-Primary (Bronchogenic) Tuberculosis, A Review.

Primary and post-primary tuberculosis (TB) are different diseases caused by the same organism. Primary TB produces systemic immunity. Post-primary TB produces cavities to support massive proliferation of organisms for transmission of infection to new hosts from a person with sufficient immunity to prevent systemic infection. Post-primary, also known as bronchogenic, TB begins in humans as asymptomatic bronchial spread of obstructive lobular pneumonia, not as expanding granulomas. Most lesions regress spontaneously. However, some undergo caseation necrosis that is coughed out through the necrotic bronchi to form cavities. Caseous pneumonia that is not expelled through the bronchi is retained to become the focus of fibrocaseous disease. No animal reproduces this entire process. However, it appears that many mammals utilize similar mechanisms, but fail to coordinate them as do humans. Understanding this makes it possible to use human tuberculous lung sections to guide manipulation of animals to produce models of particular human lesions. For example, slowly progressive and reactivation TB in mice resemble developing human bronchogenic TB. Similarly, bronchogenic TB and cavities resembling those in humans can be induced by bronchial infection of sensitized rabbits. Granulomas in guinea pigs have characteristics of both primary and post primary TB. Mice can be induced to produce a spectrum of human like caseating granulomas. There is evidence that primates can develop bronchogenic TB. We are optimistic that such models developed by coordinated study of human and animal tissues can be used with modern technologies to finally address long-standing questions about host/parasite relationships in TB, and support development of targeted therapeutics and vaccines.

Genetics of human susceptibility to active and latent tuberculosis: present knowledge and future perspectives.

Tuberculosis is an ancient human disease, estimated to have originated and evolved over thousands of years alongside modern human populations. Despite considerable advances in disease control, tuberculosis remains one of the world's deadliest communicable diseases with 10 million incident cases and 1·8 million deaths in 2015 alone based on the annual WHO report, due to inadequate health service resources in less-developed regions of the world, and exacerbated by the HIV/AIDS pandemic and emergence of multidrug-resistant strains of Mycobacterium tuberculosis. Recent findings from studies of tuberculosis infection and of patients with Mendelian predisposition to severe tuberculosis have started to reveal human loci influencing tuberculosis outcomes. In this Review, we assess the current understanding of the contribution of host genetics to disease susceptibility and to drug treatment. Despite remarkable progress in technology, only a few associated genetic variants have so far been identified, strongly indicating the need for larger global studies that investigate both common and under-represented rare variants to develop new approaches to combat the disease. Pharmacogenomic discoveries are also likely to lead to more efficient drug design and development, and ultimately safer and more effective therapies for tuberculosis.

A Systematic Approach for Developing Bacteria-Specific Imaging Tracers.

The modern patient is increasingly susceptible to bacterial infections including those due to multidrug-resistant organisms (MDROs). Noninvasive whole-body analysis with pathogen-specific imaging technologies can significantly improve patient outcomes by rapidly identifying a source of infection and monitoring the response to treatment, but no such technology exists clinically. METHODS: We systematically screened 961 random radiolabeled molecules in silico as substrates for essential metabolic pathways in bacteria, followed by in vitro uptake in representative bacteria-Staphylococcus aureus, Escherichia coli, Pseudomonas aeruginosa, and mycobacteria. Fluorine-labeled analogs, that could be developed as PET-based imaging tracers, were evaluated in a murine myositis model. RESULTS: We identified 3 novel, nontoxic molecules demonstrating selective bacterial uptake: para-aminobenzoic acid (PABA), with uptake in all representative bacteria including Mycobacterium tuberculosis; mannitol, with selective uptake in S. aureus and E. coli; and sorbitol, accumulating only in E. coli None accumulated in mammalian cells or heat-killed bacteria, suggesting metabolism-derived specificity. In addition to an extended bacterial panel of laboratory strains, all 3 molecules rapidly accumulated in respective clinical isolates of interest including MDROs such as methicillin-resistant S. aureus, extended-spectrum ß-lactamase-producing, and carbapenem-resistant Enterobacteriaceae. In a murine myositis model, fluorine-labeled analogs of all 3 molecules could rapidly detect and differentiate infection sites from sterile inflammation in mice (P = 0.03). Finally, 2-deoxy-2-[F-18]fluoro-d-sorbitol (18F-FDS) can be easily synthesized from 18F-FDG. PET, with 18F-FDS synthesized using current good manufacturing practice, could rapidly differentiate true infection from sterile inflammation to selectively localize E. coli infection in mice. CONCLUSION: We have developed a systematic approach that exploits unique biochemical pathways in bacteria to develop novel pathogen-specific imaging tracers. These tracers have significant potential for clinical translation to specifically detect and localize a broad range of bacteria, including MDROs.

Cathepsin K Contributes to Cavitation and Collagen Turnover in Pulmonary Tuberculosis.

Cavitation in tuberculosis enables highly efficient person-to-person aerosol transmission. We performed transcriptomics in the rabbit cavitary tuberculosis model. Among 17 318 transcripts, we identified 22 upregulated proteases. Five type I collagenases were overrepresented: cathepsin K (CTSK), mast cell chymase-1 (CMA1), matrix metalloproteinase 1 (MMP-1), MMP-13, and MMP-14. Studies of collagen turnover markers, specifically, collagen type I C-terminal propeptide (CICP), urinary deoxypyridinoline (DPD), and urinary helical peptide, revealed that cavitation in tuberculosis leads to both type I collagen destruction and synthesis and that proteases other than MMP-1, MMP-13, and MMP-14 are involved, suggesting a key role for CTSK. We confirmed the importance of CTSK upregulation in human lung specimens, using immunohistochemical analysis, which revealed perigranulomatous staining for CTSK, and we showed that CTSK levels were increased in the serum of patients with tuberculosis, compared with those in controls (3.3 vs 0.3 ng/mL; P = .005).

Biomarkers for Tuberculosis Based on Secreted, Species-Specific, Bacterial Small Molecules.

Improved biomarkers are needed for tuberculosis. To develop tests based on products secreted by tubercle bacilli that are strictly associated with viability, we evaluated 3 bacterial-derived, species-specific, small molecules as biomarkers: 2 mycobactin siderophores and tuberculosinyladenosine. Using liquid chromatography-tandem mass spectrometry, we demonstrated the presence of 1 or both mycobactins and/or tuberculosinyladenosine in serum and whole lung tissues from infected mice and sputum, cerebrospinal fluid (CSF), or lymph nodes from infected patients but not uninfected controls. Detection of the target molecules distinguished host infection status in 100% of mice with both serum and lung as the target sample. In human subjects, we evaluated detection of the bacterial small molecules (BSMs) in multiple body compartments in 3 patient cohorts corresponding to different forms of tuberculosis. We detected at least 1 of the 3 molecules in 90%, 71%, and 40% of tuberculosis patients' sputum, CSF, and lymph node samples, respectively. In paucibacillary forms of human tuberculosis, which are difficult to diagnose even with culture, detection of 1 or more BSM was rapid and compared favorably to polymerase chain reaction-based detection. Secreted BSMs, detectable in serum, warrant further investigation as a means for diagnosis and therapeutic monitoring in patients with tuberculosis.