Method For Rapid Identification Research Articles

Comparison of MALDI-TOF mass spectrometry and 16S rDNA sequencing for identification of environmental bacteria: a case study of cave mussel-associated culturable microorganisms.

Matrix-assisted laser desorption ionization-time of flight mass spectrometry (MALDI-TOF MS) is routinely used as a rapid and cost-effective method for pathogen identification in clinical settings. In comparison, its performance in other microbiological fields, such as environmental microbiology, is still being tested, although isolates of environmental microbes are essential for in-depth in vivo studies of their biology, including biotechnological applications. We investigated the applicability of MALDI-TOF MS for the identification of bacterial isolates from a highly oligotrophic environment-Dinaric Karst caves, which likely harbor specific microorganisms. We cultured bacteria from the shell surface of the endemic mussel Congeria jalzici, one of the three known cave mussels in the world that lives in the Dinaric karst underground. The bacterial isolates were obtained by swabbing the shell surface of mussels living in microhabitats with different amounts of water: 10 air-exposed mussels, 10 submerged mussels, and 10 mussels in the hygropetric zone. A collection of 87 pure culture isolates was obtained, mostly belonging to the phylum Bacillota (72%), followed by Pseudomonadota (16%), Actinomycetota (11%), and Bacteroidota (1%). We compared the results of MALDI-TOF MS identification (Bruker databases DB-5989 and version 11, v11) with the results of 16S rDNA-based phylogenetic analysis, a standard procedure for bacterial identification. Identification to the genus level based on 16S rDNA was possible for all isolates and clearly outperformed the results from MALDI-TOF MS, although the updated MALDI-TOF MS database v11 gave better results than the DB-5989 version (85% versus 62%). However, identification to the species-level by 16S rDNA sequencing was achieved for only 17% of isolates, compared with 14% and 40% for the MALDI-TOF MS databases DB-5989 and v11 database, respectively. In conclusion, our results suggest that continued enrichment of MALDI-TOF MS libraries will result with this method soon becoming a rapid, accurate, and efficient tool for assessing the diversity of culturable bacteria from different environmental niches.

A rapid method for identification of Lanxangia tsaoko origin and fruit shape: FT-NIR combined with chemometrics and image recognition.

Lanxangia tsaoko's accurate classifications of different origins and fruit shapes are significant for research in L. tsaoko difference between origin and species as well as for variety breeding, cultivation, and market management. In this work, Fourier transform-near infrared (FT-NIR) spectroscopy was transformed into two-dimensional and three-dimensional correlation spectroscopies to further investigate the spectral characteristics of L. tsaoko. Before building the classification model, the raw FT-NIR spectra were preprocessed using multiplicative scatter correction and second derivative, whereas principal component analysis, successive projections algorithm, and competitive adaptive reweighted sampling were used for spectral feature variable extraction. Then combined with partial least squares-discriminant analysis (PLS-DA), support vector machine (SVM), decision tree, and residual network (ResNet) models for origin and fruit shape discriminated in L. tsaoko. The PLS-DA and SVM models can achieve 100% classification in origin classification, but what is difficult to avoid is the complex process of model optimization. The ResNet image recognition model classifies the origin and shape of L. tsaoko with 100% accuracy, and without the need for complex preprocessing and feature extraction, the model facilitates the realization of fast, accurate, and efficient identification.

Rapid classification and identification of chemical components in three different Zanthoxylum species by ultra-high-performance-liquid chromatography quadrupole-orbitrap-mass spectrometry.

Zanthoxylum, as a medicinal and edible herbal medicine, has a long history and complex chemical composition. There are many varieties of Zanthoxylum, and there are differences in composition between varieties. In this study, a rapid classification and identification method for the main components of Zanthoxylum was established using ultra-high-performance-liquid chromatography quadrupole-orbitrap-mass spectrometry. The components of Shandong Zanthoxylum bungeanum, Wudu Zanthoxylum bungeanum, and Zanthoxylum schinifolium were identified by studying the characteristic fragmentations and neutral losses of characteristic components. A total of 48 common components and 24 different components were identified and the fragmentation patterns of the main components, such as flavonoids, alkaloids, and organic acids were summarized. These findings provided a reference for the study of pharmacodynamic substance basis and quality control of different varieties of Zanthoxylum.

Rapid method for preliminary identification of subthreshold strongly lensed counterparts to superthreshold gravitational-wave events

Rapid method for preliminary identification of subthreshold strongly lensed counterparts to superthreshold gravitational-wave events

A Novel Method for Rapid Identification of Acinetobacter baumannii Among ICU Patients

A Novel Method for Rapid Identification of Acinetobacter baumannii Among ICU Patients

An aptamer-based SERS method for rapid screening and identification of pathogens assisted by machine learning technique with robustness evaluation

Pathogen testing is one of the compulsory items for the quality control of pharmaceutical products. Up to now, rapid screening and identification of low-concentration pathogen contaminations is still full of challenge. Herein, an aptamer-coated magnetic beads (Apt-MBs)-based isolation strategy combined with culture medium turbidity observing and label-free SERS detection technique has been developed for rapid screening and detection of Escherichia coli, Salmonella Typhimurium and Staphylococcus aureus. Apt-MBs were employed for capturing and isolation of target pathogens. Then, with a simple enrichment process, the potential pathogens could be screened out by observing the turbidity of liquid culture medium. The presumptive bacteria species was furtherly identified via a silicon-based SERS chip couple with an optimized machine learning model, which was selected by directly evaluating the model’s generalization ability on test data. The results demonstrated that SoftMax classifier had the optimal performance with the recognition accuracy of 100 % for all samples. Finally, the method was successfully applied for identification of pathogens at ∼1 CFU spiked level in drug samples within 24 h, which is much shorter than that of the standard culture-based method (3–5 d). Therefore, the developed method showed a promising application in monitoring pathogen levels in pharmaceutical industries.

Comparison of three rapid diagnostic tests for bloodstream infections using Benefit-risk Evaluation Framework (BED-FRAME).

Rapid diagnostic tests (RDTs) for bloodstream infections have the potential to reduce time to appropriate antimicrobial therapy and improve patient outcomes. Previously, an in-house, lipid-based, matrix-assisted laser desorption/ionization-time of flight mass spectrometry (MALDI-TOF MS) method, Fast Lipid Analysis Technique (FLAT MS), has shown promise as a rapid pathogen identification method. In this study, FLAT MS for direct from blood culture identification was evaluated and compared to FDA-cleared identification methods using the Benefit-risk Evaluation Framework (BED-FRAME) analysis. FLAT MS was evaluated and compared to Bruker Sepsityper and bioMérieux BioFire FilmArray BCID2 using results from a previous study. For this study, 301 positive blood cultures were collected from the University of Maryland Medical Center. The RDTs were compared by their sensitivities, time-to-results, hands-on time, and BED-FRAME analysis. The overall sensitivity of all platforms compared to culture results from monomicrobial-positive blood cultures was 88.3%. However, the three RDTs differed in their accuracy for identifying Gram-positive bacteria, Gram-negative bacteria, and yeast. Time-to-results for FLAT MS, Sepsityper, and BioFire BCID2 were all approximately one hour. Hands-on times for FLAT MS, Sepsityper, and BioFire BCID2 were 10 (±1.3), 40 (±2.8), and 5 (±0.25) minutes, respectively. BED-FRAME demonstrated that each RDT had utility at different pathogen prevalence and relative importance. BED-FRAME is a useful tool that can used to determine which RDT is best for a healthcare center.

Rapid identification of highly glass-forming alloys in Cu–Zr binary system by laser glazing

The high-efficient experimental identification of novel metallic glasses with high glass-forming ability (GFA) is crucial for breaking through the traditional trial-and-error approach, which is time-consuming and costly. A rapid detection method for highly glass-forming alloys using laser glazing was introduced and validated in a classic binary Cu–Zr system with well-known GFA across a wide compositional range. The vitrification degree induced by laser glazing was assessed for each selected alloy and shows a strong correlation with GFA determined by traditional rapid solidification methods. The combination of experimental kinetic analysis and theoretical finite element simulation revealed possible thermodynamic mechanisms of glass formation and crystallization in the heat-affected zone (HAZ) formed by laser glazing. The results offer insights into not only the rapid identification method of highly glass-forming alloys using laser glazing but also the mechanism of glass formation under fast heating and cooling processes, providing guidance on the construction of bulk metallic glasses through laser additive manufacturing.

Rapid and reliable method for identification of three medium-sized horseshoe bat species in Europe

Rapid and reliable method for identification of three medium-sized horseshoe bat species in Europe

Ultrasensitive Electrochemiluminescence Biosensor to Detect Ampicillin Resistance Gene (ARGAMP) Based on a Novel Near-Infrared Ruthenium Carbene Complex/TPrA/PEI Ternary ECL System.

The establishment of rapid target identification and analysis methods for antibiotic resistance genes (ARGs) is urgently needed. In this study, we unprecedently designed a target-catalyzed hairpin assembly (CHA) electrochemiluminescent (ECL) biosensor for the ultrasensitive detection of ampicillin resistance genes (ARGAMP) based on a novel, efficient near-infrared ruthenium carbene complex/TPrA/PEI ternary ECL system with low oxidation potential. The ternary NIR-ECL system illustrated in this work displayed double ECL intensity in comparison with their corresponding traditional binary ECL system. The as-prepared ECL biosensor illustrated in this work demonstrates highly selective and sensitive determination of ARGAMP from 1 fM to 1 nM and a low detection limit of 0.23 fM. Importantly, it also exhibits good accuracy and stabilities to identify ARGAMP in plasmid and bacterial genome DNA, which demonstrates its excellent reliability and great potential in detecting ARGAMP in real environmental samples.

Detection of carbofuran in fruits and vegetables by Raman spectroscopy combined with immunochromatography.

It has long been desired to develop rapid methods for the rapid identification and quantification of pesticides and their metabolites. Carbofuran, a representative pesticide of the carbamate group, is highly systemic and is used on vegetables, fruits and grains, which has led many countries to test for residues in food and the environment. In this study, gold and silver composite core-shell (Au@Ag) nanoparticles were used to label the carbofuran antibody and Raman molecule 5,5-dithiobis-2-nitrobenzoic acid (DTNB) to synthesize Raman immune probes. The signal value of DTNB was read using a Raman spectrometer, and the quantitative detection technology of carbofuran was established based on lateral flow immunochromatographic assay (ICA) combined with surface-enhanced Raman spectroscopy (SERS). SERS-ICA is a rapid, quantitative and ultrasensitive test for the determination of carbofuran in fruits and vegetables with a sensitivity of 0.1 pg mL-1. Consequently, the results demonstrate that the SERS-based lateral flow immunosensor developed in this study has the advantages of excellent assay sensitivity and remarkable multiplexing capability, and thus it will have great application potential in food safety monitoring.

Label-free electrochemical immunosensor employing new redox probes/porous organic polymers/graphene oxide nanocomposite towards multiplex detection of three SARS-COV2-induced storming proteins for severe COVID-19 diagnosis

Since an outbreak of severe acute respiratory disease caused by coronavirus 2 (SARS-CoV-2) in the province of Wuhan, China, coronavirus disease 2019 (COVID-19) has tremendously evolved into a global pandemic. COVID-19 causing serious failures in organ functions is found to be involved in many groups or kinds of “cytokine storm effect” protein biomarkers resulting from the human immunological responses to the virus. The in-body abnormalities and clinical features relevant to COVID-19 exhibit a wide range and it is difficult to forecast the severity of infection in patients. Consequently, the development of a fast and highly accurate method for screening, diagnosis, monitoring, rapid identification, and isolation of COVID-19-infected patients, and monitoring its severity is urgently needed. This work aims to develop a highly sensitive multiplex label-free electrochemical immunosensor based on a porous organic polymer (POP-AQ)/graphene oxide (GO) electrode to screen and diagnose COVID-19 severity. The screen-printed carbon electrode (SPCE) consisting of three working electrodes is modified by a new POP-AQ/GO nanocomposite to enhance its specific surface area and electrochemical reactivity. The resultant platform is functionalized with three different chitosan-encapsulated redox probes (m-cresol purple (mCP), thymol blue (TMB), and polydopamine (PDA)), followed by three different antibodies, which are employed to quantitatively detect the corresponding proteins from the immunological response. This novel platform allows for the simultaneous measurement of critical illness biomarkers (IL-6, D-dimer, and ferritin), with high sensitivities and low detection limits (LODs). The biosensor displays an excellent analytical performance with wide linear dynamic ranges of 0.005–5, 2.5−100, and 2.5−100 ng mL−1, and LODs of 0.0040, 0.844, and 0.806 ng mL−1 for IL-6, D-dimer, and ferritin, respectively. Moreover, this platform reveals high reproducibility and stability, while maintaining low instrumentation complexity, and cost-effectiveness. This proposed multiplex immunosensor has excellent potential for fast COVID-19 diagnosis and severity screening in infected patients. Additionally, its versatility suggests applicability in diagnosing other diseases and emerging pathogens.

Point-of-care diagnostic devices for periodontitis - current trends and urgent need.

Point of care (POC) diagnostic devices provide a method for rapid accurate identification of disease through analysis of biologically relevant substances. This review focuses on the utility of POC testing for early detection of periodontitis, a critical factor in treating the disease. Accessing the oral cavity for biological sampling is less invasive when compared to other internal test sites, and oral fluids contain biomarkers indicative of periodontitis. The ease of access makes the mouth an excellent target location for the development of POC devices. In this review, accepted standards in industry by which these devices must adhere, provided by the World Health Organization such as REASSURED and CLIA, are discussed. An overview is provided for many periodontal biomarkers currently being investigated as a means of predicting periodontal disease and its progression. POC devices currently being investigated for the identification and monitoring of periodontal disease such as paper-based and lab-on-a-chip based devices are outlined. Limitations of current POC devices on the market are provided and future directions in leveraging biomarkers as an adjunctive method for oral diagnosis along with AI-based analysis systems are discussed. Here, we present the ESSENCE sensor platform, which combines a porous non-planar electrode with enhanced shear flow to achieve unprecedented sensitivity and selectivity. The combination of the ESENCE chip with an automated platform allows us to meet the WHO's ASSURED criteria. This platform promises to be an exciting POC candidate for early detection of periodontitis.

Proteomic Analysis of Dictyostelium discoideum by Mass Spectrometry.

The large-scale proteomic analysis of Dictyostelium discoideum has contributed to our understanding of intracellular as well as secreted proteins in this versatile model eukaryote. Mass spectrometry-based proteomic analysis is a robust, sensitive, and rapid analytical method for identification and characterization of proteins extracted from tissues, cells, cell fractions, or pull-down assays. The availability of core facilities which make proteomics inexpensive and easy to do has facilitated a wide range of research projects. In this chapter, we present a simple standard methodology to extract proteins and prepare samples from D. discoideum for mass spectrometry and methods to analyze the identified proteins.

Analysis of Targeted Components in the Combination Extract of Hovenia dulcis and Phyllanthus emblica by UPLC-MS/MS

This study aimed to establish a UPLC-MS/MS method for rapid analysis and identification of target components in the combination extract of Hovenia dulcis and Phyllanthus emblica. The gradient elution was carried out with methanol (A) −0.2% formic acid solution (A) with low rate of 1.0 mL · min−1, column temperature of 30 °C, detection wavelength of 285 nm. The ESI ion source was used to collect data under the negative ion mode. Through analysis and identification, five target components were inferred, namely gallic acid, dihydromyricetin, corilagine, taxifolin, and ellagic acid in the combination extract. Quantitative analysis was conducted on the combination extract. By utilizing the theory of modern supramolecular technology in traditional Chinese medicine, the functional composite product was obtained by selecting the compound extraction of Hovenia dulcis and Phyllanthus emblica as raw materials, and quantitative analysis was conducted on on the combination extract. The results showed that the flavonoid extraction rate and the content of target components were both higher when the mass ratio of Hovenia dulcis to Phyllanthus emblica was 1:1 and 1:2. This indicates that the co-extraction of Hovenia dulcis and Phyllanthus emblica not only simplified the extraction process but also enhances the extraction rate of the flavonoids in Hovenia dulcis. The developed UPLC-MS/MS method is rapid, sensitive, which provides a technical basis for the selection of raw material mass ratios and product development of Hovenia dulcis and Phyllanthus emblica combination.

Development and evaluation of a triplex droplet digital PCR method for differentiation of M. tuberculosis, M. bovis and BCG.

Tuberculosis, caused by Mycobacterium tuberculosis complex (MTBC), remains a global health concern in both human and animals. However, the absence of rapid, accurate, and highly sensitive detection methods to differentiate the major pathogens of MTBC, including M. tuberculosis, M. bovis, and BCG, poses a potential challenge. In this study, we have established a triplex droplet digital polymerase chain reaction (ddPCR) method employing three types of probe fluorophores, with targets M. tuberculosis (targeting CFP-10-ESAT-6 gene of RD1 and Rv0222 genes of RD4), M. bovis (targeting CFP-10-ESATs-6 gene of RD1), and BCG (targeting Rv3871 and Rv3879c genes of ΔRD1), respectively. Based on optimization of annealing temperature, sensitivity and repeatability, this method demonstrates a lower limit of detection (LOD) as 3.08 copies/reaction for M. tuberculosis, 4.47 copies/reaction for M. bovis and 3.59 copies/reaction for BCG, without cross-reaction to Mannheimia haemolytica, Mycoplasma bovis, Haemophilus parasuis, Escherichia coli, Pasteurella multocida, Ochrobactrum anthropi, Salmonella choleraesuis, Brucella melitensis, and Staphylococcus aureus, and showed repeatability with coefficients of variation (CV) lower than 10%. The method exhibits strong milk sample tolerance, the LOD of detecting in spike milk was 5 × 103 CFU/mL, which sensitivity is ten times higher than the triplex qPCR. 60 clinical DNA samples, including 20 milk, 20 tissue and 20 swab samples, were kept in China Animal Health and Epidemiology Center were tested by the triplex ddPCR and triplex qPCR. The triplex ddPCR presented a higher sensitivity (11.67%, 7/60) than that of the triplex qPCR method (8.33%, 5/60). The positive rates of M. tuberculosis, M. bovis, and BCG were 1.67, 10, and 0% by triplex ddPCR, and 1.67, 6.67, and 0% by triplex qPCR, with coincidence rates of 100, 96.7, and 100%, respectively. Our data demonstrate that the established triplex ddPCR method is a sensitive, specific and rapid method for differentiation and identification of M. tuberculosis, M. bovis, and BCG.

A point-of-care testing platform for on-site identification of genetically modified crops.

Genetically modified (GM) food is still highly controversial nowadays. Due to the disparate policies and attitudes worldwide, demands for a rapid, cost-effective and user-friendly GM crop identification method are increasingly significant for import administration, market supervision, etc. However, as the most-recognized methods, nucleic acid-based identification approaches require bulky instruments, long turn-around times and trained personnel, which are only suitable in laboratories. To fulfil the urgent needs of on-site testing, we develop a point-of-care testing platform that is able to identify 12 types of GM crops in less than 40 minutes without using laboratory settings. Our system integrates sample pre-treatment modules in a microfluidic chip, performs DNA amplification via a battery-powered portable kit, and presents results via eye-recognized colorimetric change. A paraffin-based reflow method and a slip plate-based fluid switch are developed to encapsulate and release amplification primers in individual microwells on demand, thus enabling identification of varied targets simultaneously. Our system offers an efficient, affordable and convenient tool for GM crop identification, thus it will not only benefit customs and market administration bureaus, but also satisfy demands of numerous consumers.

A multi-channel electrochemical biosensor based on polyadenine tetrahedra for the detection of multiple drug resistance genes.

Antimicrobial resistance poses a serious threat to human health due to the high morbidity and mortality caused by drug-resistant microbial infections. Therefore, the development of rapid, sensitive and selective identification methods is key to improving the survival rate of patients. In this paper, a sandwich-type electrochemical DNA biosensor based on a polyadenine-DNA tetrahedron probe was constructed. The key experimental conditions were optimized, including the length of polyadenine, the concentration of the polyadenine DNA tetrahedron, the concentration of the signal probe and the hybridization time. At the same time, poly-avidin-HRP80 was used to enhance the electrochemical detection signal. Finally, excellent biosensor performance was achieved, and the detection limit for the synthetic DNA target was as low as 1 fM. In addition, we verified the practicability of the system by analyzing E. coli with the MCR-1 plasmid and realized multi-channel detection of the drug resistance genes MCR-1, blaNDM, blaKPC and blaOXA. With the ideal electrochemical interface, the polyA-based biosensor exhibits excellent stability, which provides powerful technical support for the rapid detection of antibiotic-resistant strains in the field.

Development of a Genus Identification Method for Poisonous Plants Using Real-Time PCR

We have developed a rapid genus identification method for poisonous plants. The real-time PCR using the TaqMan® probe method was employed for detection, with the amplified targets being the "trnL (UAA)-intron" or "trnL-trnF intergenic spacer" regions of chloroplast DNA. The targeted plants were selected six genera (Aconitum, Colchicum, Veratrum, Brugmansia, Scopolia and Narcissus), which have been implicated in many instances of food poisoning in Japan. A tissue lysis solution was used for DNA extraction, which can be completed within approximate 30 min. A master mix corresponding to the tissue lysis solution was used for real-time PCR reagents. As a result, we were able to complete the entire process from DNA extraction to genus identification in 4 to 5 hr. The detection sensitivity was estimated at approximately 1 pg of DNA for all six plant genera. Remarkably, an amplification plot was discerned even with the crude cell lysates of all samples. It was also possible to obtain amplification curves for three plant samples that had been subjected to simulated cooking (boiling). This study suggests that the developed method can rapidly identify six genera of poisonous plants.

A Rapid Circuit Phase Error Identification and Compensation Method for MEMS QMG Achieving 99.7% Reduction in ZRO Drift

A Rapid Circuit Phase Error Identification and Compensation Method for MEMS QMG Achieving 99.7% Reduction in ZRO Drift