RESUMEN
Lower Respiratory Tract Infections (LRTIs) represent the leading cause of death due to infectious diseases. Current diagnostic modalities primarily depend on clinical symptoms and lack specificity, especially in light of common colonization without overt infection. To address this, we developed a noninvasive diagnostic approach that employs BreathBiomics™, an advanced human breath sampling system, to detect protease activities induced by bacterial infection in the lower respiratory tract. Specifically, we engineered a high-sensitivity and high-specificity molecular sensor for human neutrophil elastase (HNE). The sensor undergoes cleavage in the presence of HNE, an event that is subsequently detected via Matrix-Assisted Laser Desorption/Ionization Time of Flight Mass Spectrometry (MALDI-TOF MS). Application of this methodology to clinical samples, breath specimens collected from intubated patients with LRTIs, demonstrated the detection of the cleaved sensor by MALDI-TOF MS. Our findings indicate that this novel approach offers a noninvasive and specific diagnostic strategy for people with LRTIs.
RESUMEN
Diagnosing respiratory tract infections (RTIs) in critical care settings is essential for appropriate antibiotic treatment and lowering mortality. The current diagnostic method, which primarily relies on clinical symptoms, lacks sensitivity and specificity, resulting in incorrect or delayed diagnoses, putting patients at a heightened risk. In this study we developed a noninvasive diagnosis method based on collecting non-volatile compounds in human exhaled air. We hypothesized that non-volatile compound profiles could be effectively used for bacterial RTI diagnosis. Exhaled air samples were collected from subjects receiving mechanical ventilation diagnosed with or without bacterial RTI in intensive care units at the Johns Hopkins Hospital. Truncated proteoforms, a class of non-volatile compounds, were characterized by top-down proteomics, and significant features associated with RTI were identified using feature selection algorithms. The results showed that three truncated proteoforms, collagen type VI alpha three chain protein, matrix metalloproteinase-9, and putative homeodomain transcription factor II were independently associated with RTI with thep-values of 2.0 × 10-5, 1.1 × 10-4, and 1.7 × 10-3, respectively, using multiple logistic regression. Furthermore, a score system named 'TrunScore' was constructed by combining the three truncated proteoforms, and the diagnostic accuracy was significantly improved compared to that of individual truncated proteoforms, with an area under the receiver operator characteristic curve of 96.9%. This study supports the ability of this noninvasive breath analysis method to provide an accurate diagnosis for RTIs in subjects receiving mechanical ventilation. The results of this study open the doors to be able to potentially diagnose a broad range of diseases using this non-volatile breath analysis technique.
Asunto(s)
Infecciones del Sistema Respiratorio , Compuestos Orgánicos Volátiles , Humanos , Respiración Artificial , Pruebas Respiratorias/métodos , Infecciones del Sistema Respiratorio/diagnóstico , Espiración , Compuestos Orgánicos Volátiles/análisisRESUMEN
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes the ongoing coronavirus disease 2019 (COVID-19) pandemic. Here we report a novel strategy for the rapid detection of SARS-CoV-2 based on an enrichment approach exploiting the affinity between the virus and cellulose sulfate ester functional groups, hot acid hydrolysis, and matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry (MALDI-TOF MS). Virus samples were enriched using cellulose sulfate ester microcolumns. Virus peptides were prepared using the hot acid aspartate-selective hydrolysis and characterized by MALDI-TOF MS. Collected spectra were processed with a peptide fingerprint algorithm, and searching parameters were optimized for the detection of SARS-CoV-2. These peptides provide high sequence coverage for nucleocapsid (N protein) and allow confident identification of SARS-CoV-2. Peptide markers contributing to the detection were rigorously identified using bottom-up proteomics. The approach demonstrated in this study holds the potential for developing a rapid assay for COVID-19 diagnosis and detecting virus variants from a variety of sources, such as sewage and nasal swabs.
Asunto(s)
COVID-19 , SARS-CoV-2 , COVID-19/diagnóstico , Prueba de COVID-19 , Celulosa/análogos & derivados , Ésteres , Humanos , Péptidos/química , Espectrometría de Masa por Láser de Matriz Asistida de Ionización Desorción/métodosRESUMEN
Human breath contains trace amounts of non-volatile organic compounds (NOCs) which might provide non-invasive methods for evaluating individual health. In previous work, we demonstrated that lipids detected in exhaled breath aerosol (EBA) could be used as markers of active tuberculosis (TB). Here, we advanced our analytical platform for characterizing small metabolites and lipids in EBA samples collected from participants enrolled in clinical trials designed to identify molecular signatures of active TB. EBA samples from 26 participants with active TB and 73 healthy participants were processed using a dual-phase extraction method, and metabolites and lipids were identified via mass spectrometry database matching. In total, 13 metabolite and 9 lipid markers were identified with statistically different optimized relative standard deviation values between individuals diagnosed with active TB and the healthy controls. Importantly, EBA lipid profiles can be used to separate the two sample types, indicating the diagnostic potential of the identified molecules. A feature ranking algorithm reduced this number to 10 molecules, with the membrane glycerophospholipid, phosphatidylinositol 24:4, emerging as the top driver of segregation between the two groups. These results support the use of this approach to identify consistent NOC signatures from EBA samples in active TB cases. This suggests the potential to apply this method to other human diseases which alter respiratory NOC release.
Asunto(s)
Líquidos Corporales , Tuberculosis , Compuestos Orgánicos Volátiles , Aerosoles/análisis , Biomarcadores/análisis , Líquidos Corporales/química , Pruebas Respiratorias/métodos , Espiración , Humanos , Lípidos/análisis , Tuberculosis/diagnóstico , Compuestos Orgánicos Volátiles/análisisRESUMEN
Antibacterial drugs are an important component of malaria therapy. We studied the interactions of clindamycin, tetracycline, chloramphenicol, and ciprofloxacin against Plasmodium falciparum under static and dynamic conditions. In microtiter plate assays (static conditions), and as expected, parasites displayed the delayed death response characteristic for apicoplast-targeting drugs. However, rescue by isopentenyl pyrophosphate was variable, ranging from 2,700-fold for clindamycin to just 1.7-fold for ciprofloxacin, suggesting that ciprofloxacin has targets other than the apicoplast. We also examined the pharmacokinetic-pharmacodynamic relationships of these antibacterials in an in vitro glass hollow-fiber system that exposes parasites to dynamically changing drug concentrations. The same total dose and area under the concentration-time curve (AUC) of the drug was deployed either as a single short-lived high peak (bolus) or as a constant low concentration (infusion). All four antibacterials were unambiguously time-driven against malaria parasites: infusions had twice the efficacy of bolus regimens, for the same AUC. The time-dependent efficacy of ciprofloxacin against malaria is in contrast to its concentration-driven action against bacteria. In silico simulations of clinical dosing regimens and resulting pharmacokinetics revealed that current regimens do not maximize time above the MICs of these drugs. Our findings suggest that simple and rational changes to dosing may improve the efficacy of antibacterials against falciparum malaria.
Asunto(s)
Antibacterianos/farmacología , Malaria Falciparum/tratamiento farmacológico , Plasmodium falciparum/efectos de los fármacos , Antibacterianos/farmacocinética , Área Bajo la Curva , Humanos , Cinética , Malaria Falciparum/metabolismo , Pruebas de Sensibilidad Microbiana/métodosRESUMEN
Hsp90 inhibitors, well studied in the laboratory and clinic for antitumor indications, have promising activity against protozoan pathogens, including Trypanosoma brucei which causes African sleeping sickness, and the malaria parasite, Plasmodium falciparum To progress these experimental drugs toward clinical use, we adapted an in vitro dynamic hollow-fiber system and deployed artificial pharmacokinetics to discover the driver of their activity: either concentration or time. The activities of compounds from three major classes of Hsp90 inhibitors in development were evaluated against trypanosomes. In all circumstances, the activities of the tested Hsp90 inhibitors were concentration driven. By optimally deploying the drug to match its kinetic driver, the efficacy of a given dose was improved up to 5-fold, and maximal efficacy was achieved with a significantly lower drug exposure. The superiority of concentration-driven regimens was evident in vitro over several logs of drug exposure and was predictive of efficacy in a mouse model of African trypanosomiasis. In studies with P. falciparum, antimalarial activity was similarly concentration driven. This experimental strategy offers an expedient and versatile translational tool to assess the impact of pharmacokinetics on antiprotozoal activity. Knowing kinetic governance early in drug development provides an additional metric for judging lead compounds and allows the incisive design of animal efficacy studies.
Asunto(s)
Antiprotozoarios/farmacocinética , Proteínas HSP90 de Choque Térmico/antagonistas & inhibidores , Malaria Falciparum/tratamiento farmacológico , Plasmodium falciparum/efectos de los fármacos , Proteínas Protozoarias/antagonistas & inhibidores , Trypanosoma brucei brucei/efectos de los fármacos , Tripanosomiasis Africana/tratamiento farmacológico , Animales , Antineoplásicos/sangre , Antineoplásicos/farmacocinética , Antineoplásicos/farmacología , Antiprotozoarios/sangre , Antiprotozoarios/farmacología , Área Bajo la Curva , Benzodioxoles/sangre , Benzodioxoles/farmacocinética , Benzodioxoles/farmacología , Benzoquinonas/sangre , Benzoquinonas/farmacocinética , Benzoquinonas/farmacología , Bioensayo , Modelos Animales de Enfermedad , Reposicionamiento de Medicamentos , Femenino , Expresión Génica , Proteínas HSP90 de Choque Térmico/genética , Proteínas HSP90 de Choque Térmico/metabolismo , Imidazoles/sangre , Imidazoles/farmacocinética , Imidazoles/farmacología , Isoxazoles/sangre , Isoxazoles/farmacocinética , Isoxazoles/farmacología , Lactamas Macrocíclicas/sangre , Lactamas Macrocíclicas/farmacocinética , Lactamas Macrocíclicas/farmacología , Malaria Falciparum/parasitología , Ratones , Modelos Biológicos , Plasmodium falciparum/crecimiento & desarrollo , Proteínas Protozoarias/genética , Proteínas Protozoarias/metabolismo , Resorcinoles/sangre , Resorcinoles/farmacocinética , Resorcinoles/farmacología , Trypanosoma brucei brucei/crecimiento & desarrollo , Tripanosomiasis Africana/parasitologíaRESUMEN
BACKGROUND: Operation Installation (OI), a community-based smoke alarm installation programme in Dallas, Texas, targets houses in high-risk urban census tracts. Residents of houses that received OI installation (or programme houses) had 68% fewer medically treated house fire injuries (non-fatal and fatal) compared with residents of non-programme houses over an average of 5.2â years of follow-up during an effectiveness evaluation conducted from 2001 to 2011. OBJECTIVE: To estimate the cost-benefit of OI. METHODS: A mathematical model incorporated programme cost and effectiveness data as directly observed in OI. The estimated cost per smoke alarm installed was based on a retrospective analysis of OI expenditures from administrative records, 2006-2011. Injury incidence assumptions for a population that had the OI programme compared with the same population without the OI programme was based on the previous OI effectiveness study, 2001-2011. Unit costs for medical care and lost productivity associated with fire injuries were from a national public database. RESULTS: From a combined payers' perspective limited to direct programme and medical costs, the estimated incremental cost per fire injury averted through the OI installation programme was $128,800 (2013 US$). When a conservative estimate of lost productivity among victims was included, the incremental cost per fire injury averted was negative, suggesting long-term cost savings from the programme. The OI programme from 2001 to 2011 resulted in an estimated net savings of $3.8 million, or a $3.21 return on investment for every dollar spent on the programme using a societal cost perspective. CONCLUSIONS: Community smoke alarm installation programmes could be cost-beneficial in high-fire-risk neighbourhoods.
Asunto(s)
Prevención de Accidentes/economía , Prevención de Accidentes/instrumentación , Accidentes Domésticos/prevención & control , Planificación en Salud Comunitaria , Incendios/economía , Incendios/prevención & control , Equipos de Seguridad/economía , Accidentes Domésticos/economía , Análisis Costo-Beneficio , Incendios/estadística & datos numéricos , Estudios de Seguimiento , Vivienda , Humanos , Modelos Teóricos , Desarrollo de Programa , Evaluación de Programas y Proyectos de Salud , Texas , Población UrbanaAsunto(s)
Prevención de Accidentes/métodos , Planificación en Salud Comunitaria/métodos , Incendios/prevención & control , Equipos de Seguridad/estadística & datos numéricos , Lesión por Inhalación de Humo/epidemiología , Estudios de Seguimiento , Humanos , Incidencia , Evaluación de Programas y Proyectos de Salud , Humo , Texas/epidemiología , Factores de TiempoRESUMEN
Knowledge of pharmacokinetic/pharmacodynamic (PK/PD) relationships can enhance the speed and economy of drug development by enabling informed and rational decisions at every step, from lead selection to clinical dosing. For anti-infective agents in particular, dynamic in vitro hollow-fiber cartridge experiments permit exquisite control of kinetic parameters and the study of their consequent impact on pharmacodynamic efficacy. Such information is of great interest for the cost-restricted but much-needed development of new antimalarial drugs, especially since the major human pathogen Plasmodium falciparum can be cultivated in vitro but is not readily available in animal models. This protocol describes the materials and procedures for determining the PK/PD relationships of antimalarial compounds.