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International Journal of... Apr 2024On a global scale, India holds the distinction of having the greatest number of tuberculosis (TB) cases caused by Mycobacterium tuberculosis (MTB) complex. The study... (Comparative Study)
Comparative Study
BACKGROUND
On a global scale, India holds the distinction of having the greatest number of tuberculosis (TB) cases caused by Mycobacterium tuberculosis (MTB) complex. The study aimed at evaluating the sensitivity, specificity, accuracy, cost, rapidity, and feasibility of the performance of the colorimetric nitrate reductase-based antibiotic susceptibility (CONRAS) test against the indirect proportion method (IPM) on Lowenstein-Jensen media as the gold standard.
METHODS
A comparative cross-sectional study was performed on 51 MTB isolates. Fresh subcultures were used for drug susceptibility testing by IPM on the Lowenstein-Jensen medium and the CONRAS method in liquid medium. Quality control for drug susceptibility testing was done using a known sensitive strain of MTB (H37Rv) and strains resistant to both isoniazid (INH) and rifampicin (RIF) - multidrug-resistant (MDR), mono-resistant to RIF, streptomycin (STM), and ethambutol (EMB). Statistical analysis was performed using MedCalc software (Version 20.027).
RESULTS
CONRAS, carried out in microfuge tubes, was cost-efficient and easy to perform/interpret with most results being available in 10 days compared to 42 days in the case of IPM. The sensitivity, specificity, and accuracy of RIF and INH were 100%, 97.37%, and 98.04 and 93.33%, 97.59%, and 96.08%, respectively, which translates into an almost perfect agreement between the two methods as indicated by κ value of 0.905 and 0.949, respectively, for the two drugs. The performance of CONRAS was less satisfactory for STM and EMB when compared to IPM.
CONCLUSIONS
CONRAS may serve as a useful test for the detection of MDR-TB because of its accuracy, low cost, ease of performance/interpretation, and rapidity when compared to IPM on LJ medium. It does not involve the use of expensive reagents and equipment, as is the case with molecular methods like GeneXpert and line probe assay, making it a suitable option for the detection of MDR-TB in resource-poor settings.
Topics: Nitrate Reductase; Mycobacterium tuberculosis; Colorimetry; Microbial Sensitivity Tests; Antitubercular Agents; Humans; Cross-Sectional Studies; Sensitivity and Specificity; Culture Media; India; Tuberculosis, Multidrug-Resistant
PubMed: 38916391
DOI: 10.4103/ijmy.ijmy_69_24 -
BioRxiv : the Preprint Server For... Jun 2024Cellular functional pathways have evolved through selection based on fitness benefits conferred through protein intra- and inter-molecular interactions that comprise all...
Cellular functional pathways have evolved through selection based on fitness benefits conferred through protein intra- and inter-molecular interactions that comprise all protein conformational features and protein-protein interactions, collectively referred to as the interactome. While the interactome is regulated by proteome levels, it is also regulated independently by, post translational modification, co-factor, and ligand levels, as well as local protein environmental factors, such as osmolyte concentration, pH, ionic strength, temperature and others. In modern biomedical research, cultivatable cell lines have become an indispensable tool, with selection of optimal cell lines that exhibit specific functional profiles being critical for success in many cases. While it is clear that cell lines derived from different cell types have differential proteome levels, increased understanding of large-scale functional differences requires additional information beyond abundance level measurements, including how protein conformations and interactions are altered in certain cell types to shape functional landscapes. Here, we employed quantitative protein cross-linking coupled to mass spectrometry to probe large-scale protein conformational and interaction changes among three commonly employed human cell lines, HEK293, MCF-7, and HeLa cells. Isobaric quantitative Protein Interaction Reporter (iqPIR) technologies were used to obtain quantitative values of cross-linked peptides across three cell lines. These data illustrated highly reproducible (R values larger than 0.8 for all biological replicates) quantitative interactome levels across multiple biological replicates. We also measured protein abundance levels in these cells using data independent acquisition quantitative proteomics methods. Combining quantitative interactome and proteomics information allowed visualization of cell type- specific interactome changes mediated by proteome level adaptations as well as independently regulated interactome changes to gain deeper insight into possible drivers of these changes. Among the biggest detected alterations in protein interactions and conformations are changes in cytoskeletal proteins, RNA-binding proteins, chromatin remodeling complexes, mitochondrial proteins, and others. Overall, these data demonstrate the utility and reproducibility of quantitative cross-linking to study systems-level interactome variations. Moreover, these results illustrate how combined quantitative interactomics and proteomics can provide unique insight on cellular functional landscapes.
PubMed: 38915502
DOI: 10.1101/2024.06.12.598691 -
Health Science Reports Jun 2024The current study aimed to evaluate the efficiency of Enzyme-linked immunosorbent assay (ELISA) assay and monoplex and multiplex real-time reverse-transcription PCR...
Evaluating the efficiency of ELISA, monoplex and multiplex probe-based real-time reverse-transcription PCR assays in the detection of SARS-CoV-2 (COVID-19) and influenza A and B viruses: A cross-sectional study.
BACKGROUND AND AIMS
The current study aimed to evaluate the efficiency of Enzyme-linked immunosorbent assay (ELISA) assay and monoplex and multiplex real-time reverse-transcription PCR (rRT-PCR) in the detection of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) and influenza A and B viruses (Flu A and Flu B).
METHODS
The SARS-CoV-2 -specific IgG and IgM antibodies, as well as, Flu A (H1N1 and H3N2 serotypes) and Flu B virus antibodies were determined by ELISA assay. The one-step qRT-PCR method was used to detect the SARS-CoV-2 in nasopharyngeal swab samples. Furthermore, the presence of Flu A and B viruses was evaluated using probe-based RT-PCR. Simultaneous detection of SARS-CoV-2, Flu A and B viruses was performed by multiplex rRT-PCR assay.
RESULTS
SARS CoV-2 IgM and IgG antibodies were detected in 33.3% and 58.3% of patients, respectively. In contrast, the SARS CoV-2 genome was detected in 50% of patients using the one-step monoplex RT-PCR assay. Flu A serotypes H1N1 and H3N2 were found in 16.7% and 8.3% of patients. Probe-based RT-PCR revealed that 39.3% of patients were positive for the Flu A virus. Multiplex rRT-PCR detect the SARS-CoV-2, Flu A, and Flu B in 50%, 39.3%, and 19% of samples, respectively. The sensitivity and specificity of multiplex rRT-PCR assay in comparison to monoplex RT-PCR were 100% and 55%, respectively. Coinfection with SARS-CoV-2, Flu A, and Flu B viruses was found in 9.5% of patients.
CONCLUSION
Multiplex rRT-PCR can be used as a repaid, cost-effective and suitable tool for molecular surveillance of SARS-CoV-2 and Flu A/B viruses.
PubMed: 38915351
DOI: 10.1002/hsr2.2140 -
Communications Biology Jun 2024Chromatin organization and dynamics play important roles in governing the regulation of nuclear processes of biological cells. However, due to the constant diffusive...
Chromatin organization and dynamics play important roles in governing the regulation of nuclear processes of biological cells. However, due to the constant diffusive motion of chromatin, examining chromatin nanostructures in living cells has been challenging. In this study, we introduce interferometric scattering correlation spectroscopy (iSCORS) to spatially map nanoscopic chromatin configurations within unlabeled live cell nuclei. This label-free technique captures time-varying linear scattering signals generated by the motion of native chromatin on a millisecond timescale, allowing us to deduce chromatin condensation states. Using iSCORS imaging, we quantitatively examine chromatin dynamics over extended periods, revealing spontaneous fluctuations in chromatin condensation and heterogeneous compaction levels in interphase cells, independent of cell phases. Moreover, we observe changes in iSCORS signals of chromatin upon transcription inhibition, indicating that iSCORS can probe nanoscopic chromatin structures and dynamics associated with transcriptional activities. Our scattering-based optical microscopy, which does not require labeling, serves as a powerful tool for visualizing dynamic chromatin nano-arrangements in live cells. This advancement holds promise for studying chromatin remodeling in various crucial cellular processes, such as stem cell differentiation, mechanotransduction, and DNA repair.
Topics: Chromatin; Humans; Spectrum Analysis; Interferometry; Chromatin Assembly and Disassembly; Cell Nucleus
PubMed: 38914653
DOI: 10.1038/s42003-024-06457-2 -
PeerJ 2024Breast carcinoma (BRCA) is a life-threatening malignancy in women and shows a poor prognosis. Cuproptosis is a novel mode of cell death but its relationship with BRCA is...
BACKGROUND
Breast carcinoma (BRCA) is a life-threatening malignancy in women and shows a poor prognosis. Cuproptosis is a novel mode of cell death but its relationship with BRCA is unclear. This study attempted to develop a cuproptosis-relevant prognostic gene signature for BRCA.
METHODS
Cuproptosis-relevant subtypes of BRCA were obtained by consensus clustering. Differential expression analysis was implemented using the 'limma' package. Univariate Cox and multivariate Cox analyses were performed to determine a cuproptosis-relevant prognostic gene signature. The signature was constructed and validated in distinct datasets. Gene set variation analysis (GSVA) and gene set enrichment analysis (GSEA) were also conducted using the prognostic signature to uncover the underlying molecular mechanisms. ESTIMATE and CIBERSORT algorithms were applied to probe the linkage between the gene signature and tumor microenvironment (TME). Immunotherapy responsiveness was assessed using the Tumor Immune Dysfunction and Exclusion (TIDE) web tool. Real-time quantitative PCR (RT-qPCR) was performed to detect the expressions of cuproptosis-relevant prognostic genes in breast cancer cell lines.
RESULTS
Thirty-eight cuproptosis-associated differentially expressed genes (DEGs) in BRCA were mined by consensus clustering and differential expression analysis. Based on univariate Cox and multivariate Cox analyses, six cuproptosis-relevant prognostic genes, namely SAA1, KRT17, VAV3, IGHG1, TFF1, and CLEC3A, were mined to establish a corresponding signature. The signature was validated using external validation sets. GSVA and GSEA showed that multiple cell cycle-linked and immune-related pathways along with biological processes were associated with the signature. The results ESTIMATE and CIBERSORT analyses revealed significantly different TMEs between the two Cusig score subgroups. Finally, RT-qPCR analysis of cell lines further confirmed the expressional trends of SAA1, KRT17, IGHG1, and CLEC3A.
CONCLUSION
Taken together, we constructed a signature for projecting the overall survival of BRCA patients and our findings authenticated the cuproptosis-relevant prognostic genes, which are expected to provide a basis for developing prognostic molecular biomarkers and an in-depth understanding of the relationship between cuproptosis and BRCA.
Topics: Humans; Breast Neoplasms; Female; Prognosis; Computational Biology; Biomarkers, Tumor; Gene Expression Regulation, Neoplastic; Gene Expression Profiling; Tumor Microenvironment; Cell Line, Tumor
PubMed: 38912044
DOI: 10.7717/peerj.17419 -
ACS Bio & Med Chem Au Jun 2024Synthetic modification of oligodeoxynucleotides (ODNs) via conjugation to nucleic acid binding small molecules can improve hybridization and pharmacokinetic properties....
Synthetic modification of oligodeoxynucleotides (ODNs) via conjugation to nucleic acid binding small molecules can improve hybridization and pharmacokinetic properties. In the present study, five Hoechst 33258 derived benzimidazoles were conjugated to T rich ODNs and their hybridization effectiveness was tested. Thermal denaturation studies revealed significant stabilization of complementary duplexes by ODN-benzimidazole conjugates, with the extent of stabilization being highly dependent on the length of the linker between DNA and benzimidazole. The increases in thermal stability were determined to be due to the binding of the benzimidazole moiety to the duplex. Circular dichroism and molecular modeling studies provided insights toward the influence of conjugation on duplex structure and how linker length impacts placement of the benzimidazole moiety in the minor groove. Furthermore, thermal denaturation studies with the complementary strand containing a single base mismatch or being RNA revealed that covalent conjugation of benzimidazoles to an ODN also enhances the sequence specificity. The fundamental studies reported herein provide a strategy to improve the stability and specificity properties of the ODN probes, which can be of use for targeting and diagnostics applications.
PubMed: 38911908
DOI: 10.1021/acsbiomedchemau.3c00074 -
ACS Photonics Jun 2024Electrically connected and plasmonically enhanced molecular junctions combine the optical functionalities of high field confinement and enhancement (cavity function),...
Electrically connected and plasmonically enhanced molecular junctions combine the optical functionalities of high field confinement and enhancement (cavity function), and of high radiative efficiency (antenna function) with the electrical functionalities of molecular transport. Such combined optical and electrical probes have proven useful for the fundamental understanding of metal-molecule contacts and contribute to the development of nanoscale optoelectronic devices including ultrafast electronics and nanosensors. Here, we employ a self-assembled metal-molecule-metal junction with a nanoparticle bridge to investigate correlated fluctuations in conductance and tunneling-induced light emission at room temperature. Despite the presence of hundreds of molecules in the junction, the electrical conductance and light emission are both highly sensitive to atomic-scale fluctuations-a phenomenology reminiscent of picocavities observed in Raman scattering and of luminescence blinking from photoexcited plasmonic junctions. Discrete steps in conductance associated with fluctuating emission intensities through the multiple plasmonic modes of the junction are consistent with a finite number of randomly localized, point-like sources dominating the optoelectronic response. Contrasting with these microscopic fluctuations, the overall plasmonic and electronic functionalities of our devices feature long-term survival at room temperature and under an electrical bias of a few volts, allowing for measurements over several months.
PubMed: 38911841
DOI: 10.1021/acsphotonics.4c00291 -
ACS Energy Letters Jun 2024Understanding photophysical processes in lead halide perovskites is an important aspect of optimizing the performance of optoelectronic devices. The determination of... (Review)
Review
Understanding photophysical processes in lead halide perovskites is an important aspect of optimizing the performance of optoelectronic devices. The determination of exact charge carrier extraction rate constants remains elusive, as there is a large and persistent discrepancy in the reported absolute values. In this review, we concentrate on experimental procedures adopted in the literature to obtain kinetic estimates of charge transfer processes and limitations imposed by the spectroscopy technique employed. Time-resolved techniques (e.g., transient absorption-reflection and time-resolved photoluminescence spectroscopy) are commonly employed to probe charge transfer at perovskite/transport layer interfaces. The variation in sample preparation and measurement conditions can produce a wide dispersion of the measured kinetic parameters. The selected time window and the kinetic fitting model employed introduce additional uncertainty. We discuss here evaluation strategies that rely on multiexponential fitting protocols (regular or stretched) and show how the dispersion in the reported values for carrier transfer rate constants can be resolved.
PubMed: 38911533
DOI: 10.1021/acsenergylett.4c00736 -
Talanta Jun 2024The certification of cosmetic products has always been a prominent concern. Here, we have developed a pH sensor and applied it in the field of cosmetic safety....
The certification of cosmetic products has always been a prominent concern. Here, we have developed a pH sensor and applied it in the field of cosmetic safety. Initially, we designed two probes, CH with aggregation-induced emission (AIE) effect and the near-infrared fluorophore derivative CYTYR. By encapsulating them with DSPE-PEG2000-NH, we obtained the CHCY-lipo nano-micelles with fluorescence resonance energy transfer (FRET) response. By combining them into a sensor array called pC, we achieved sensitive detection of a wide pH range, ranging from 4.69 to 9.25. To validate the performance of the pC sensor array, we employed a multi-channel mode and applied it to differentiate commercial anti-aging creams. Through linear discriminant analysis and 3D fingerprint analysis, the pC sensor array successfully distinguished anti-aging creams from different countries, providing a rapid and accurate method for cosmetic safety identification. The results of this study demonstrate the potential of the pC sensor array for quick authentication of cosmetic products, offering significant support and application prospects in safeguarding consumer health.
PubMed: 38909596
DOI: 10.1016/j.talanta.2024.126447 -
Time-resolved fluorescence of tryptophan characterizes membrane perturbation by cyclic lipopeptides.Biophysical Journal Jun 2024Viscosin is a membrane-permeabilizing, cyclic lipopeptide (CLiP) produced by Pseudomonas species. Here, we have studied four synthetic analogs (L1W, V4W, L5W, L7W), each...
Viscosin is a membrane-permeabilizing, cyclic lipopeptide (CLiP) produced by Pseudomonas species. Here, we have studied four synthetic analogs (L1W, V4W, L5W, L7W), each with one leucine (Leu; L) or valine residue exchanged for tryptophan (Trp; W) by means of time-resolved fluorescence spectroscopy of Trp. To this end, we recorded the average fluorescence lifetime, rotational correlation time and limiting anisotropy, dipolar relaxation time and limiting extent of relaxation, rate constant of acrylamide quenching, effect of HO-DO exchange, and time-resolved halfwidth of the spectrum in the absence and presence of POPC liposomes. Structure, localization, and hydration of the peptides were described by molecular dynamics simulations. The combination of the parameters provides a good description of the molecular environments of the Trp positions and the behavior of viscosin as a whole. Of particular value for characterizing the impact of viscosin on the membrane is the dipolar relaxation of Trp4 in V4W, which is deeply embedded in the hydrophobic core. The limiting relaxation level represents the membrane perturbation - unlike typical membrane probes - at the site of the perturbant. Fractions of Trp4 relax at different rates; the one not in contact with water upon excitation relaxes via recruitment of a water molecule on the 10 ns time scale. This rate is sensitive to the concerted membrane perturbation by more than one lipopeptide, which appears at high lipopeptide concentration and is assumed a prerequisite for the final formation of a membrane-permeabilizing defect. Trp7 relaxes primarily with respect to neighboring Ser residues. Trp5 flips between a membrane-inserted and surface-exposed orientation.
PubMed: 38909278
DOI: 10.1016/j.bpj.2024.06.022