RESUMO
We report a 23% asymptomatic severe acute respiratory syndrome coronavirus 2 (SARS CoV-2) Omicron carriage rate in participants being enrolled into a clinical trial in South Africa, 15-fold higher than in trials before Omicron. We also found lower CD4â +â T-cell counts in persons with human immunodeficiency virus (HIV) strongly correlated with increased odds of being SARS-CoV-2 polymerase chain reaction (PCR) positive.
Assuntos
COVID-19 , SARS-CoV-2 , Humanos , Reação em Cadeia da Polimerase , África do Sul/epidemiologiaRESUMO
The antileprosy drug clofazimine was recently repurposed as part of a newly endorsed short-course regimen for multidrug-resistant tuberculosis. It also enables significant treatment shortening when added to the first-line regimen for drug-susceptible tuberculosis in a mouse model. However, clofazimine causes dose- and duration-dependent skin discoloration in patients, and the optimal clofazimine dosing strategy in the context of the first-line regimen is unknown. We utilized a well-established mouse model to systematically address the impacts of duration, dose, and companion drugs on the treatment-shortening activity of clofazimine in the first-line regimen. In all studies, the primary outcome was relapse-free cure (culture-negative lungs) 6 months after stopping treatment, and the secondary outcome was bactericidal activity, i.e., the decline in the lung bacterial burden during treatment. Our findings indicate that clofazimine activity is most potent when coadministered with first-line drugs continuously throughout treatment and that equivalent treatment-shortening results are obtained with half the dose commonly used in mice. However, our studies also suggest that clofazimine at low exposures may have negative impacts on treatment outcomes, an effect that was evident only after the first 3 months of treatment. These data provide a sound evidence base to inform clofazimine dosing strategies to optimize the antituberculosis effect while minimizing skin discoloration. The results also underscore the importance of conducting long-term studies to allow the full evaluation of drugs administered in combination over long durations.
Assuntos
Antituberculosos/uso terapêutico , Clofazimina/uso terapêutico , Tuberculose/tratamento farmacológico , Animais , Modelos Animais de Doenças , Feminino , Camundongos , Camundongos Endogâmicos BALB C , Distribuição Aleatória , Tuberculose Resistente a Múltiplos Medicamentos/tratamento farmacológicoRESUMO
OBJECTIVES: The anti-leprosy drug clofazimine has been shown to have antimicrobial activity against Mycobacterium tuberculosis and has been associated with treatment-shortening activity in both clinical and preclinical studies of TB chemotherapy. However, a reported lack of early bactericidal activity (EBA) in TB patients has raised questions regarding the usefulness of clofazimine as an anti-TB drug. Our objective was to systematically evaluate the EBA of clofazimine in vitro and in vivo to provide insight into how and when this drug exerts its antimicrobial activity against M. tuberculosis. METHODS: We evaluated the 14 day EBA of clofazimine (i) in vitro at concentrations ranging from 4 times below to 4 times above the MIC for M. tuberculosis and (ii) in vivo in infected BALB/c mice at doses ranging from 1.5 to 100 mg/kg/day, and serum clofazimine levels were measured. In both experiments, isoniazid was used as the positive control. RESULTS: In vitro, clofazimine, at any concentration tested, did not exhibit bactericidal activity during the first week of exposure; however, in the second week, it exhibited concentration-dependent antimicrobial activity. In vivo, clofazimine, at any dose administered, did not exhibit bactericidal activity during the first week, and limited antimicrobial activity was observed during the second week of administration. While serum clofazimine levels were clearly dose dependent, the antimicrobial activity was not significantly related to the dose administered. CONCLUSIONS: Our data suggest that clofazimine's delayed antimicrobial activity may be due more to its mechanism of action rather than to host-related factors.
Assuntos
Antituberculosos/uso terapêutico , Carga Bacteriana/efeitos dos fármacos , Clofazimina/uso terapêutico , Mycobacterium tuberculosis/efeitos dos fármacos , Tuberculose Pulmonar/tratamento farmacológico , Animais , Antituberculosos/farmacocinética , Clofazimina/farmacocinética , Isoniazida/uso terapêutico , Pulmão/microbiologia , Camundongos , Camundongos Endogâmicos BALB C , Testes de Sensibilidade Microbiana , Tuberculose Pulmonar/microbiologiaRESUMO
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.
Assuntos
Biomarcadores/análise , Mycobacterium tuberculosis/química , Oxazóis/análise , Tuberculose/diagnóstico , Adenosina/análogos & derivados , Adenosina/análise , Animais , Técnicas de Tipagem Bacteriana , Cromatografia Líquida , Humanos , Pulmão/microbiologia , Camundongos , Mycobacterium tuberculosis/isolamento & purificação , Escarro/microbiologia , Espectrometria de Massas em TandemRESUMO
The antileprosy drug clofazimine has shown potential for shortening tuberculosis treatment; however, the current dosing of the drug is not evidence based, and the optimal dosing is unknown. Our objective was to conduct a preclinical evaluation of the pharmacokinetics and pharmacodynamics of clofazimine in the mouse model of tuberculosis, with the goal of providing useful information on dosing for future studies. Pharmacokinetic parameters were evaluated in infected and uninfected BALB/c mice. Pharmacodynamic parameters were evaluated in Mycobacterium tuberculosis-infected mice that were treated for 12 weeks with one of six different clofazimine dosing regimens, i.e., doses of 6.25, 12.5, and 25 mg/kg of body weight/day and 3 regimens with loading doses. Clofazimine progressively accumulated in the lungs, livers, and spleens of the mice, reaching levels of greater than 50 µg/g in all tissues by 4 weeks of administration, while serum drug levels remained low at 1 to 2 µg/ml. Elimination of clofazimine was extremely slow, and the half-life was dependent on the duration of drug administration. Clofazimine exhibited dose-dependent tissue and serum concentrations. At any dose, clofazimine did not have bactericidal activity during the first 2 weeks of administration but subsequently demonstrated potent, dose-independent bactericidal activity. The antituberculosis activity of clofazimine was dependent on neither the dose administered nor the drug concentrations in the tissues, suggesting that much lower doses could be effectively used for tuberculosis treatment.
Assuntos
Antituberculosos/farmacocinética , Clofazimina/farmacocinética , Tuberculose/sangue , Tuberculose/tratamento farmacológico , Animais , Antituberculosos/uso terapêutico , Cromatografia Líquida , Clofazimina/uso terapêutico , Feminino , Espectrometria de Massas , Camundongos , Camundongos Endogâmicos BALB C , Ligação ProteicaRESUMO
BACKGROUND: Studies of the quality of tuberculosis (TB) diagnostic evaluation of patients in high burden countries have generally shown poor adherence to international or national guidelines. Health worker perspectives on barriers to improving TB diagnostic evaluation are critical for developing clinic-level interventions to improve guideline implementation. METHODS: We conducted structured, in-depth interviews with staff at six district-level health centers in Uganda to elicit their perceptions regarding barriers to TB evaluation. Interviews were transcribed, coded with a standardized framework, and analyzed to identify emergent themes. We used thematic analysis to develop a logic model depicting health system and contextual barriers to recommended TB evaluation practices. To identify possible clinic-level interventions to improve TB evaluation, we categorized findings into predisposing, enabling, and reinforcing factors as described by the PRECEDE model, focusing on potentially modifiable behaviors at the clinic-level. RESULTS: We interviewed 22 health center staff between February 2010 and November 2011. Participants identified key health system barriers hindering TB evaluation, including: stock-outs of drugs/supplies, inadequate space and infrastructure, lack of training, high workload, low staff motivation, and poor coordination of health center services. Contextual barrier challenges to TB evaluation were also reported, including the time and costs borne by patients to seek and complete TB evaluation, poor health literacy, and stigma against patients with TB. These contextual barriers interacted with health system barriers to contribute to sub-standard TB evaluation. Examples of intervention strategies that could address these barriers and are related to PRECEDE model components include: assigned mentors/peer coaching for new staff (targets predisposing factor of low motivation and need for support to conduct job duties); facilitated workshops to implement same day microscopy (targets enabling factor of patient barriers to completing TB evaluation), and recognition/incentives for good TB screening practices (targets low motivation and self-efficacy). CONCLUSIONS: Our findings suggest that health system and contextual barriers work together to impede TB diagnosis at health centers and, if not addressed, could hinder TB case detection efforts. Qualitative research that improves understanding of the barriers facing TB providers is critical to developing targeted interventions to improve TB care.
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Atitude do Pessoal de Saúde , Atenção à Saúde , Tuberculose/diagnóstico , Assistência Ambulatorial/normas , Instituições de Assistência Ambulatorial/economia , Instituições de Assistência Ambulatorial/normas , Instituições de Assistência Ambulatorial/provisão & distribuição , Antituberculosos/provisão & distribuição , Custos e Análise de Custo , Feminino , Pessoal de Saúde/educação , Pessoal de Saúde/normas , Nível de Saúde , Humanos , Programas de Rastreamento/economia , Programas de Rastreamento/organização & administração , Programas de Rastreamento/normas , Quartos de Pacientes/provisão & distribuição , Opinião Pública , Pesquisa Qualitativa , Estigma Social , Tuberculose/economia , UgandaRESUMO
BACKGROUND: HIV type 1 (HIV-1) remains a global health concern, with the greatest burden in sub-Saharan Africa. Despite 40 years of research, no vaccine candidate has shown durable and protective efficacy against HIV-1 acquisition. Although pre-exposure prophylaxis in groups with high vulnerability can be very effective, barriers to its use, such as perceived low acquisition risk, fear of stigma, and concerns about side-effects, remain. Thus, a population-based approach, such as an HIV-1 vaccine, is needed. The current study aimed to evaluate the efficacy and safety of a heterologous HIV-1 vaccine regimen, consisting of a tetravalent mosaic adenovirus 26-based vaccine (Ad26.Mos4.HIV) and aluminium phosphate-adjuvanted clade C glycoprotein (gp) 140, in young women at risk of acquiring HIV-1 in southern Africa. METHODS: This randomised, double-blind, phase 2b study enrolled sexually active women without HIV-1 or HIV-2 aged 18-35 years at 23 clinical research sites in Malawi, Mozambique, South Africa, Zambia, and Zimbabwe. Participants were centrally randomly assigned (1:1) to receive intramuscular injections of vaccine or saline placebo in stratified permuted blocks via an interactive web response system. Study participants, study site personnel (except those with primary responsibility for study vaccine preparation and dispensing), and investigators were masked to treatment group allocation. The vaccine regimen consisted of Ad26.Mos4.HIV administered at months 0 and 3 followed by Ad26.Mos4.HIV administered concurrently with aluminium phosphate-adjuvanted clade C gp140 at months 6 and 12. The primary efficacy outcome was vaccine efficacy in preventing laboratory-confirmed HIV-1 acquisition diagnosed between visits at month 7 and month 24 after the first vaccination (VE[7-24]) in the per-protocol population, which included participants who had not acquired HIV-1 4 weeks after the third vaccination, received all planned vaccinations at the first three vaccination visits within the protocol-specified windows, and had no major protocol deviations that could affect vaccine efficacy. Primary safety outcomes were assessed in randomly assigned participants who received one study injection or more based on the actual injection received. The primary safety endpoints were the incidences of unsolicited adverse events (AEs), solicited local and systemic AEs, serious AEs, AEs of special interest, and AEs leading to discontinuation of vaccination. This trial is registered with ClinicalTrials.gov, NCT03060629, and is complete. FINDINGS: Between Nov 3, 2017, and June 30, 2019, 2654 women were randomly assigned, of whom 2636 women (median age of 23 years [IQR 20-25]) were enrolled and received at least one study injection (1313 assigned vaccine, 1323 placebo; 1317 received vaccine, 1319 placebo). Analysis of the primary efficacy outcome in the per-protocol cohort included 1080 women in the vaccine group and 1108 women in the placebo group; the incidence of HIV-1 acquisition per 100 person-years over months 7-24 after the first vaccination was 3·38 (95% CI 2·54-4·41) in the vaccine group and 3·94 (3·04-5·03) in the placebo group, with an estimated VE(7-24) of 14·10% (95% CI -22·00 to 39·51; p=0·40). There were no serious unsolicited AEs, AEs of special interest, or deaths related to the study vaccine. In the vaccine group, 663 (50·3%) of 1317 participants had grade 1 or 2 solicited local AEs and ten (0·8%) of 1317 participants had grade 3 or 4 solicited local AEs. In the placebo group, 305 (23·1%) of 1319 participants had grade 1 or 2 solicited local AEs and three (0·2%) of 1319 participants had grade 3 or 4 solicited local AEs. 863 (65·5%) of 1317 participants in the vaccine group had grade 1 or 2 solicited systemic AEs and 34 (2·6%) of 1317 participants had grade 3 or 4 solicited systemic AEs. 763 (57·8%) of 1319 participants in the placebo group had grade 1 or 2 solicited systemic AEs and 20 (1·5%) of 1319 participants had grade 3 or 4 solicited systemic AEs. Overall, three (0·2%) of 1317 participants in the vaccine group and three (0·2%) of 1319 participants in the placebo group discontinued vaccination due to an unsolicited AE, and three (0·2%) of 1317 participants in the vaccine group and one (0·1%) of 1319 participants in the placebo group discontinued vaccination due to a solicited AE. INTERPRETATION: The heterologous Ad26.Mos4.HIV and clade C gp140 vaccine regimen was safe and well tolerated but did not show efficacy in preventing HIV-1 acquisition in a population of young women in southern Africa at risk of HIV-1. FUNDING: Division of AIDS at the National Institute of Allergy and Infectious Diseases through the HIV Vaccine Trials Network, Bill & Melinda Gates Foundation, Janssen Vaccines & Prevention, US Army Medical Materiel Development Activity, and Ragon Institute.
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Access to sputum smear microscopy in high-tuberculosis (TB)-burden regions is limited by a scarcity of microscopes and experienced technicians. We evaluated the accuracy of CellScope, a novel digital fluorescence microscope that may expand access to microscopy. The study utilized smear microscopy slides prepared from sputum specimens submitted by consecutive adults with ≥ 2 weeks of cough who were admitted to Mulago Hospital (Kampala, Uganda). Conventional light-emitting diode (LED) fluorescence microscopy (FM) and mycobacterial culture were performed by experienced technicians. Two U.S.-based postgraduate researchers without prior microscopy experience restained, imaged, and interpreted the slides using CellScope. We assessed whether sensitivity and specificity of CellScope-based LED FM was noninferior to conventional LED FM by using a preselected margin of inferiority of 15%. Of 525 patients included, 72% were HIV seropositive and 39% had culture-confirmed TB. The proportions of positive results were similar with CellScope and conventional LED FM (34% versus 32%, respectively; P = 0.32), and agreement was substantial. CellScope accuracy was within the noninferiority margin for both sensitivity (63% versus 70%; difference, -7%; 95% confidence interval [CI], -13% to -1%) and specificity (85% versus 92%; difference, -7%; 95% CI, -12% to -3%). A subanalysis of 43 slides evaluated by each CellScope reader found substantial interreader reliability (custom-weighted kappa, 0.65) and variable intrareader reliability (custom-weighted kappa, 0.11 versus 0.48). CellScope offers promise for expanding microscopy services. Future studies should evaluate the device when operated by health workers in low-resource settings, the feasibility of image transmission and analysis by experienced microscopists, and the accuracy of automated image analysis algorithms.
Assuntos
Técnicas Bacteriológicas/métodos , Microscopia de Fluorescência/métodos , Sistemas Automatizados de Assistência Junto ao Leito , Tuberculose/diagnóstico , Adulto , Feminino , Humanos , Masculino , Sensibilidade e Especificidade , Manejo de Espécimes/métodos , Escarro/microbiologia , UgandaRESUMO
The early widespread dissemination of Omicron indicates the urgent need to better understand the transmission dynamics of this variant, including asymptomatic spread among immunocompetent and immunosuppressed populations. In early December 2021, the Ubuntu clinical trial, designed to evaluate efficacy of the mRNA-1273 vaccine (Moderna) among persons living with HIV (PLWH), began enrolling participants. Nasal swabs are routinely obtained at the initial vaccination visit, which requires participants to be clinically well to receive their initial jab. Of the initial 230 participants enrolled between December 2 and December 17, 2021, 71 (31%) were PCR positive for SARS-CoV-2: all of whom were subsequently confirmed by S gene dropout to be Omicron; 48% of the tested samples had cycle threshold (CT) values <25 and 18% less than 20, indicative of high titers of asymptomatic shedding. Asymptomatic carriage rates were similar in SARS-CoV-2 seropositive and seronegative persons (27% respectively). These data are in stark contrast to COVID-19 vaccine studies conducted pre-Omicron, where the SARS-CoV-2 PCR positivity rate at the first vaccination visit ranged from <1%-2.4%, including a cohort of over 1,200 PLWH largely enrolled in South Africa during the Beta outbreak. We also evaluated asymptomatic carriage in a sub study of the Sisonke vaccine trial conducted in South African health care workers, which indicated 2.6% asymptomatic carriage during the Beta and Delta outbreaks and subsequently rose to 16% in both PLWH and PHLWH during the Omicron period.
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Although small molecules shed from pathogens are widely used to diagnose infection, such tests have not been widely implemented for tuberculosis. Here we show that the recently identified compound, 1-tuberculosinyladenosine (1-TbAd), accumulates to comprise >1% of all Mycobacterium tuberculosis lipid. In vitro and in vivo, two isomers of TbAd were detected that might serve as infection markers. Using mass spectrometry and nuclear magnetic resonance, we established the structure of the previously unknown molecule, N(6)-tuberculosinyladenosine (N(6)-TbAd). Its biosynthesis involves enzymatic production of 1-TbAd by Rv3378c followed by conversion to N(6)-TbAd via the Dimroth rearrangement. Intact biosynthetic genes are observed only within M. tuberculosis complex bacteria, and TbAd was not detected among other medically important pathogens, environmental bacteria, and vaccine strains. With no substantially similar known molecules in nature, the discovery and in vivo detection of two abundant terpene nucleosides support their development as specific diagnostic markers of tuberculosis.
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Lipídeos/biossíntese , Mycobacterium tuberculosis/metabolismo , Nucleosídeos/análise , Terpenos/química , Tuberculose/diagnóstico , Animais , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Cromatografia Líquida de Alta Pressão , Isomerismo , Lipídeos/análise , Lipídeos/isolamento & purificação , Pulmão/metabolismo , Espectroscopia de Ressonância Magnética , Camundongos , Camundongos Endogâmicos BALB C , Mycobacterium tuberculosis/genética , Nucleosídeos/biossíntese , Nucleosídeos/química , Polimorfismo de Nucleotídeo Único , Espectrometria de Massas por Ionização por ElectrosprayRESUMO
In low-resource areas, the most common method of tuberculosis (TB) diagnosis is visual identification of rod-shaped TB bacilli in microscopic images of sputum smears. We present an algorithm for automated TB detection using images from digital microscopes such as CellScope, a novel, portable device capable of brightfield and fluorescence microscopy. Automated processing on such platforms could save lives by bringing healthcare to rural areas with limited access to laboratory-based diagnostics. Our algorithm applies morphological operations and template matching with a Gaussian kernel to identify candidate TB-objects. We characterize these objects using Hu moments, geometric and photometric features, and histograms of oriented gradients and then perform support vector machine classification. We test our algorithm on a large set of CellScope images (594 images corresponding to 290 patients) from sputum smears collected at clinics in Uganda. Our object-level classification performance is highly accurate, with average precision of 89.2% +/- 2.1%. For slide-level classification, our algorithm performs at the level of human readers, demonstrating the potential for making a significant impact on global healthcare.