Blood-based diagnosis of pediatric tuberculosis: A prospective cohort study in South Africa and Dominican Republic.

Objectives: Pediatric tuberculosis (TB) diagnosis is complicated by challenges in obtaining invasive respiratory specimens that frequently contain few Mycobacterium tuberculosis (Mtb) bacilli. We report the diagnostic performance of an Mtb antigen-derived peptide (MAP-TB) assay and its ability to monitor TB treatment response.

Methods: Study cohorts enrolled children who presented with presumptive TB at two hospitals in South Africa from 2012 to 2017 (157 children aged <13 years) and at community-based clinics in the Dominican Republic from 2019 to 2023 (101 children aged <18 years).

Label-Free Raman Probing of the Intrinsic Electric Field for High-Efficiency Screening of Electricity-Producing Bacteria at the Single-Cell Level.

The fabrication of high-performance microbial fuel cells requires the evaluation of the activity of electrochemically active bacteria. However, this is challenging because of the time-consuming nature of biofilm formation and the invasive nature of labeling. To address this issue, we developed a fast, label-free, single-cell Raman spectroscopic method.

CRISPR for companion diagnostics in low-resource settings.

New point-of-care tests (POCTs), which are especially useful in low-resource settings, are needed to expand screening capacity for diseases that cause significant mortality: tuberculosis, multiple cancers, and emerging infectious diseases. Recently, clustered regularly interspaced short palindromic repeats (CRISPR)-based diagnostic (CRISPR-Dx) assays have emerged as powerful and versatile alternatives to traditional nucleic acid tests, revealing a strong potential to meet this need for new POCTs. In this review, we discuss CRISPR-Dx assay techniques that have been or could be applied to develop POCTs, including techniques for sample processing, target amplification, multiplex assay design, and signal readout.

Valence-Change MnO-Coated Arsenene Nanosheets as a Pin1 Inhibitor for Hepatocellular Carcinoma Treatment.

The heterogeneity of hepatocellular carcinoma (HCC) can prevent effective treatment, emphasizing the need for more effective therapies. Herein, we employed arsenene nanosheets coated with manganese dioxide and polyethylene glycol (AMPNs) for the degradation of Pin1, which is universally overexpressed in HCC. By employing an "AND gate", AMPNs exhibited responsiveness toward excessive glutathione and hydrogen peroxide within the tumor microenvironment, thereby selectively releasing AsO to mitigate potential side effects of AsO.

Serum Cell-Free DNA-based Detection of Complex Infection.

complex (MAC) is the most common cause of nontuberculous mycobacterial (NTM) pulmonary disease (PD), which exhibits increasing global incidence. Current microbiologic methods routinely used in clinical practice lack sensitivity and have long latencies, leading to delays in diagnosis and treatment initiation and evaluation. A clustered regularly interspaced short palindromic repeats (CRISPR)-based assay that measures MAC cell-free DNA (cfDNA) concentrations in serum could provide a rapid means to detect MAC infection and monitor response to antimicrobial treatment.

Sensitive Blood-Based Detection of HIV-1 and Mycobacterium tuberculosis Peptides for Disease Diagnosis by Immuno-Affinity Liquid Chromatography-Tandem Mass Spectrometry: A Method Development and Proof-of-Concept Study.

Background: Novel approaches that allow early diagnosis and treatment monitoring of both human immunodeficiency virus-1 (HIV-1) and tuberculosis disease (TB) are essential to improve patient outcomes.

Methods: We developed and validated an immuno-affinity liquid chromatography-tandem mass spectrometry (ILM) assay that simultaneously quantifies single peptides derived from HIV-1 p24 and Mycobacterium tuberculosis (Mtb) 10-kDa culture filtrate protein (CFP10) in trypsin-digested serum derived from cryopreserved serum archives of cohorts of adults and children with/without HIV and TB.

Results: ILM p24 and CFP10 results demonstrated good intra-laboratory precision and accuracy, with recovery values of 96.

Clinical Peptidomics: Advances in Instrumentation, Analyses, and Applications.

Extensive effort has been devoted to the discovery, development, and validation of biomarkers for early disease diagnosis and prognosis as well as rapid evaluation of the response to therapeutic interventions. Genomic and transcriptomic profiling are well-established means to identify disease-associated biomarkers. However, analysis of disease-associated peptidomes can also identify novel peptide biomarkers or signatures that provide sensitive and specific diagnostic and prognostic information for specific malignant, chronic, and infectious diseases.

Outlook for CRISPR-based tuberculosis assays now in their infancy.

Tuberculosis (TB) remains a major underdiagnosed public health threat worldwide, being responsible for more than 10 million cases and one million deaths annually. TB diagnosis has become more rapid with the development and adoption of molecular tests, but remains challenging with traditional TB diagnosis, but there has not been a critical review of this area. Here, we systematically review these approaches to assess their diagnostic potential and issues with the development and clinical evaluation of proposed CRISPR-based TB assays.

CRISPR Assays for Disease Diagnosis: Progress to and Barriers Remaining for Clinical Applications.

Numerous groups have employed the special properties of CRISPR/Cas systems to develop platforms that have broad potential applications for sensitive and specific detection of nucleic acid (NA) targets. However, few of these approaches have progressed to commercial or clinical applications. This review summarizes the properties of known CRISPR/Cas systems and their applications, challenges associated with the development of such assays, and opportunities to improve their performance or address unmet assay needs using nano-/micro-technology platforms.

CRISPR-based assays for low-resource settings.

CRISPR-based assays can be adopted as ultrasensitive molecular diagnostics in resource-limited settings, but point-of-care applications must address additional requirements. Here, we discuss the major obstacles for developing these assays and offer insights into how to surmount them.

Nanopore-based disease diagnosis using pathogen-derived tryptic peptides from serum.

Nanopore sensors have shown great utility in nucleic acid detection and sequencing approaches. Recent studies also indicate that current signatures produced by peptide-nanopore interactions can distinguish high purity peptide mixtures, but the utility of nanopore sensors in clinical applications still needs to be explored due to the inherent complexity of clinical specimens. To fill this gap between research and clinical nanopore applications, we describe a methodology to select peptide biomarkers suitable for use in an immunoprecipitation-coupled nanopore (IP-NP) assay, based on their pathogen specificity, antigenicity, charge, water solubility and ability to produce a characteristic nanopore interaction signature.

Advanced technologies for molecular diagnosis of cancer: State of pre-clinical tumor-derived exosome liquid biopsies.

Exosomes are membrane-defined extracellular vesicles (EVs) approximately 40-160 ​nm in diameter that are found in all body fluids including blood, urine, and saliva. They act as important vehicles for intercellular communication between both local and distant cells and can serve as circulating biomarkers for disease diagnosis and prognosis. Exosomes play a key role in tumor metastasis, are abundant in biofluids, and stabilize biomarkers they carry, and thus can improve cancer detection, treatment monitoring, and cancer staging/prognosis.

Identifying the Phenotypes of Tumor-Derived Extracellular Vesicles Using Size-Coded Affinity Microbeads.

Tumor-derived extracellular vesicle (tEV) biomarkers can reflect cancer cell phenotypes and have great potential for cancer diagnosis and treatment. However, tEVs display high heterogeneity, and rapid and sensitive identification of EV biomarkers remains challenging due to their low expression. Spectral overlap also significantly limits the multiplex analysis of EV biomarkers by fluorescent probes.

Monocrystalline Labeling Enables Stable Plasmonic Enhancement for Isolation-Free Extracellular Vesicle Analysis.

Sensitive detection of extracellular vesicles (EVs) as emerging biomarkers has shown great promises for disease diagnosis. Plasmonic metal nanostructures conjugated with molecules that bind specific biomarker targets are widely used for EVs sensing but involve tradeoffs between particle-size-dependent signal intensity and conjugation efficiency. One solution to this problem would be to induce nucleation on nanoparticles that have successfully bound a target biomarker to permit in situ nanoparticle growth for signal amplification, but approaches that are evaluated to date require harsh conditions or lack nucleation specificity, prohibiting their effective use with most biological specimens.

Extracellular vesicles: Emerging tools as therapeutic agent carriers.

Extracellular vesicles (EVs) are secreted by both eukaryotes and prokaryotes, and are present in all biological fluids of vertebrates, where they transfer DNA, RNA, proteins, lipids, and metabolites from donor to recipient cells in cell-to-cell communication. Some EV components can also indicate the type and biological status of their parent cells and serve as diagnostic targets for liquid biopsy. EVs can also natively carry or be modified to contain therapeutic agents (, nucleic acids, proteins, polysaccharides, and small molecules) by physical, chemical, or bioengineering strategies.

Diagnosis of paediatric tuberculosis by optically detecting two virulence factors on extracellular vesicles in blood samples.

Sensitive and specific blood-based assays for the detection of pulmonary and extrapulmonary tuberculosis would reduce mortality associated with missed diagnoses, particularly in children. Here we report a nanoparticle-enhanced immunoassay read by dark-field microscopy that detects two Mycobacterium tuberculosis virulence factors (the glycolipid lipoarabinomannan and its carrier protein) on the surface of circulating extracellular vesicles. In a cohort study of 147 hospitalized and severely immunosuppressed children living with HIV, the assay detected 58 of the 78 (74%) cases of paediatric tuberculosis, 48 of the 66 (73%) cases that were missed by microbiological assays, and 8 out of 10 (80%) cases undiagnosed during the study.

CRISPR detection of circulating cell-free Mycobacterium tuberculosis DNA in adults and children, including children with HIV: a molecular diagnostics study.

Background: Tuberculosis remains a leading cause of global mortality, especially for adults and children living with HIV (CLHIV) underdiagnosed by sputum-based assays. Non-sputum-based assays are needed to improve tuberculosis diagnosis and tuberculosis treatment monitoring. Our aim in this study was to determine whether ultrasensitive detection of Mycobacterium tuberculosis cell-free DNA (Mtb-cfDNA) in blood can diagnose tuberculosis and evaluate tuberculosis treatment responses.

Silicon Nanodisk Huygens Metasurfaces for Portable and Low-Cost Refractive Index and Biomarker Sensing.

Biomarker detection and bulk refractive index sensing are important across multiple industries ranging from early medical diagnosis to chemical process quality control. The bulky size, high cost, and complex architecture of existing refractive index and biomarker sensing technologies limit their use to highly skilled environments like hospitals, large food processing plants, and research labs. Here, we demonstrate a compact and inexpensive refractive index sensor based on resonant dielectric photonic nanoantenna arrays or metasurfaces.