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In this study, the goal was to develop a method for detecting and classifying organophosphorus pesticides (OPPs) in bodies of water. Sixty-five samples with different concentrations were prepared for each of the organophosphorus pesticides, namely chlorpyrifos, acephate, parathion-methyl, trichlorphon, dichlorvos, profenofos, malathion, dimethoate, fenthion, and phoxim, respectively. Firstly, the spectral data of all the samples was obtained using a UV–visible spectrometer. Secondly, five preprocessing methods, six manifold learning methods, and five machine learning algorithms were utilized to build detection models for identifying OPPs in water bodies. The findings indicate that the accuracy of machine learning models trained on data preprocessed using convolutional smoothing + first-order derivatives (SG + FD) outperforms that of models trained on data preprocessed using other methods. The backpropagation neural network (BPNN) model exhibited the highest accuracy rate at 99.95%, followed by the support vector machine (SVM) and convolutional neural network (CNN) models, both at 99.92%. The extreme learning machine (ELM) and K-nearest neighbors (KNN) models demonstrated accuracy rates of 99.84% and 99.81%, respectively. Following the application of a manifold learning algorithm to the full-wavelength data set for the purpose of dimensionality reduction, the data was then visualized in the first three dimensions. The results demonstrate that the t-distributed domain embedding (t-SNE) algorithm is superior, exhibiting dense clustering of similar clusters and clear classification of dissimilar ones. SG + FD-t-SNE-SVM ranks highest among the feature extraction models in terms of performance. The feature extraction dimension was set to 4, and the average classification accuracy was 99.98%, which slightly improved the prediction performance over the full-wavelength model. As shown in this study, the ultraviolet–visible (UV–visible) spectroscopy system combined with the t-SNE and SVM algorithms can effectively identify and classify OPPs in waterbodies. [ABSTRACT FROM AUTHOR]

Emerging infectious diseases pose a serious threat to human health and affect social stability. In recent years, the epidemic situation of emerging infectious diseases is very serious; among these infectious diseases, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has affected many countries and regions in a short time. The prevention and treatment of these diseases require rapid on-site detection methods. However, the common detection method, RT-PCR, requires expensive instruments, complex operations, and professional operators. Here, we developed a portable low-cost assay for rapid on-site detection of viral nucleic acid using reverse transcription-loop-mediated isothermal amplification (RT-LAMP). The SARS-CoV-2 RNA can be successfully amplified within 15 min in a thermos, and the detection result is read rapidly in a portable low-cost device with a sensitivity of 100 copies/µL. The portable low-cost device consists of a black box, a laser or LED and a filter, costing only a few cents. The rapid on-site detection method can provide strong support for the control of biological threats such as infectious diseases. It is also an emergency detection method for low-resource settings, relieving the huge pressure on health care. [ABSTRACT FROM AUTHOR]

As nucleic acid testing is playing a vital role in increasingly many research fields, the need for rapid on-site testing methods is also increasing. The test procedure often consists of three steps: Sample preparation, amplification, and detection. This review covers recent advances in on-chip methods for each of these three steps and explains the principles underlying related methods. The sample preparation process is further divided into cell lysis and nucleic acid purification, and methods for the integration of these two steps on a single chip are discussed. Under amplification, on-chip studies based on PCR and isothermal amplification are covered. Three isothermal amplification methods reported to have good resistance to PCR inhibitors are selected for discussion due to their potential for use in direct amplification. Chip designs and novel strategies employed to achieve rapid extraction/amplification with satisfactory efficiency are discussed. Four detection methods providing rapid responses (fluorescent, optical, and electrochemical detection methods, plus lateral flow assay) are evaluated for their potential in rapid on-site detection. In the final section, we discuss strategies to improve the speed of the entire procedure and to integrate all three steps onto a single chip; we also comment on recent advances, and on obstacles to reducing the cost of chip manufacture and achieving mass production. We conclude that future trends will focus on effective nucleic acid extraction via combined methods and direct amplification via isothermal methods.

Emerging infectious diseases pose a serious threat to human health and affect social stability. In recent years, the epidemic situation of emerging infectious diseases is very serious; among these infectious diseases, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has affected many countries and regions in a short time. The prevention and treatment of these diseases require rapid on-site detection methods. However, the common detection method, RT-PCR, requires expensive instruments, complex operations, and professional operators. Here, we developed a portable low-cost assay for rapid on-site detection of viral nucleic acid using reverse transcription-loop-mediated isothermal amplification (RT-LAMP). The SARS-CoV-2 RNA can be successfully amplified within 15 min in a thermos, and the detection result is read rapidly in a portable low-cost device with a sensitivity of 100 copies/µL. The portable low-cost device consists of a black box, a laser or LED and a filter, costing only a few cents. The rapid on-site detection method can provide strong support for the control of biological threats such as infectious diseases. It is also an emergency detection method for low-resource settings, relieving the huge pressure on health care.

The white spot syndrome virus is the most destructive virus threatening the shrimp industry worldwide, causing hundreds of millions of dollars in economic losses each year. There is currently no specific medicine to treat it. Therefore, rapid and accurate detection of WSSV is of great significance for controlling its spread and reducing economic losses. Traditional detection methods, such as polymerase chain reaction (PCR) and quantitative fluorescent PCR, rely on laboratory equipment and are not suitable for field testing. In this study, recombinase polymerase amplification (RPA) combined with a lateral flow strip (LFS) was developed. This method targets the entire genome and designs primers and probes accordingly. The detection can be completed in 30 min at 37°C, and the detection limit of each reaction is 20 copies, which is much more sensitive than other detection methods. The RPA-LFS method is highly specific to the white spot syndrome virus and has no cross-reactivity with other common shrimp viruses or pathogens. In total, 100 field samples were tested and compared to the real-time PCR method. Both methods detected 8 positive results, and the positive detection rate was 100%. The method was fast, simple, specific, and sensitive. It does not rely on laboratory equipment and has broad application prospects for in-field detection, especially in remote areas with underdeveloped medical equipment. [ABSTRACT FROM AUTHOR]

Haemophilus parasuis (H. parasuis) is a gram-negative bacterial pathogen that causes severe infections in swine, resulting in substantial economic losses. Currently, the majority of H. parasuis detection methods are impractical for on-site application due to their reliance on large instruments or complex procedures. Thus, there is an urgent need to develop a rapid, visually detectable, and highly sensitive detection method, especially under resource-limited environments and field conditions. In this study, we established a naked eye assay for highly sensitive detection by combining recombinase polymerase amplification (RPA) with CRISPR/Cas12a technology. Positive samples exhibited a clear red color visible to the naked eye, while negative samples appeared blue. We achieved a remarkable sensitivity, detecting H. parasuis down to a single copy, with no cross-reactivity with other bacteria. In a mouse model, our assay detected H. parasuis infection nearly 8 h earlier than traditional PCR. Compared to qPCR, our detection results were 100 % accurate. To enhance point-of-care applicability and mitigate the risk of aerosol contamination from uncapping, we consolidated RPA and CRISPR/Cas12a cleavage into a single-tube reaction system. This integrated approach was validated with 20 clinical lung samples, yielding results consistent with those obtained from qPCR. The entire procedure, from DNA extraction to detection, was completed in 35 min. We present an RPA-CRISPR/Cas12a assay suitable for the early and resource-efficient diagnosis of H. parasuis infections. Its simplicity and visual detection are advantageous for field diagnostics, representing a substantial develpoment in the diagnosis of H. parasuis. [Display omitted] • RPA-CRISPR/Cas12a for rapid Haemophilus parasuis detection. • One-pot assay reduces steps and contamination risk. • Naked-eye detection of H. parasuis for field diagnostics. [ABSTRACT FROM AUTHOR]

Species identification has become a significant concern due to the growing use of food alternatives that may cause allergies and reduce nutritional value. To address the issue of fraudulent adulteration of goat milk products with cow milk, we have developed an affordable, portable, and user-friendly platform called microfluidic-integrated nucleic acid lateral flow strips (LFS). This platform enables simultaneous detection of components derived from both goats and cows in goat milk. In this study, we have introduced an innovative nucleic acid labeling method. The loop primers of loop-mediated isothermal amplification (LAMP) have been modified with amplification terminator spacer C3 and an oligonucleotide sequence, thus eliminating the requirement for costly antibodies in traditional nucleic acid LFS. This modification not only lowers costs but also enables multiple detections. Additionally, we have integrated the LAMP and LFS assay steps into a microfluidic chip, allowing convenient on-site detection while effectively preventing aerosol contamination of LAMP products. The testing process includes rapid DNA extraction, followed by a short nucleic acid addition and incubation for visualized results in about 50 min. This platform is user-friendly, requiring no specialized equipment or extensive training, making it suitable for rapid on-site detection of dairy products by personnel in diverse fields. [Display omitted] • Development of a new nucleic acid labeling method to replace the use of antibody. • A novel microfluidic-assisted cassette integrated LAMP and LFS visual detection. • The complete testing procedure is instrument-free. • The entire testing procedure can be completed within 50min. • 5% of cow milk in goat milk could be detected. [ABSTRACT FROM AUTHOR]

The porcine epidemic diarrhea virus (PEDV) is a major pathogen of swine enteric diseases. Various etiology and serological methods have been employed for PEDV detection, but most of their applications are limited to laboratories. To extend PEDV detection to on-site applications, we design a homogeneous fluorescence resonance energy transfer (FRET)-based enzyme-linked immunosorbent assay (ELISA). Both donor and acceptor fluorescence microspheres modified PEDV antibodies can be linked only to the occurrence of PEDV antigen, thus generating FRET signals, which can be collected by our designed portable FRET immunoassay station (FRETIS). Verified by standard samples, FRET immunoassay reached high sensitivity with a detection limit as TCID 50 (median tissue culture infective dose) of 10/mL, which is 10 times more sensitive than colloidal gold test strips; and verified by clinical samples, it was also proved with high accuracy, good selectivity, and repeatability. More importantly, FRET immunoassay could detect PEDV in a 96-well plate in 35 min with only one step of incubation without any further washing steps using field-portable devices and field-operable procedures, well supporting on-site applications. Considering these advantages, this reported FRET immunoassay provides a promising way for multi-sample on-site PEDV detection and can be potentially used in the swine industry. [Display omitted] • Homogeneous FRET based ELISA for rapid on-site PEDV detection is developed. • It has rather high sensitivity of detection limit as TCID 50 of 10 /mL. • It has high accuracy, good selectivity and repeatability in clinical applications. • It has high detecting throughput only with single step incubation. • It only relies on field-portable devices and field-operable processes. [ABSTRACT FROM AUTHOR]

Florfenicol (FF), used popularly in prevention and treatment of virus infections in livestock and poultry, has widely been found in eggs and harmful to human health. In this work, a sensitive and quantitative on-site detecting solution, monoclonal antibody-based carboxylated fluorescent microsphere immunochromatographic test strip assay (FM-ICTS), is design and applied for FF detection. The proposed method can sensitively detect FF in low detection limit of 0.030 ng/g and quantitatively measure its concentration from 0.1 ng/mL to 8.1 ng/mL (R2 = 0.9991) with high repeatability (CV<8.0 %). In addition, the established FM-ICTS method exhibited high measurement accuracy in FF samples as compared with HPLC-MS analysis and demonstrated satisfied recoveries (99.1–101.3 %). More importantly, the quantitative FF test strip demonstrate ultra-high stability, which presents approximately equivalent detection ability to the fresh one after stored at 4 °C for more than one year or stored at 37 °C for 60 days. Therefore, the proposed method is a promising solution for rapidly and sensitively quantitative determination of FF in eggs. • Carboxylated fluorescent microsphere based immunochromatographic test strip is design and applied for florfenicol detection. • The method can sensitively and quantitatively detect FF in low detection limit with high repeatability. • The established FM-ICTS method exhibited high measurement accuracy and satisfied recovery in FF samples detection. • The quantitative FF test strip demonstrate ultra-high stability, which can be stored at 4°C for more than one year. [ABSTRACT FROM AUTHOR]

As nucleic acid testing is playing a vital role in increasingly many research fields, the need for rapid on-site testing methods is also increasing. The test procedure often consists of three steps: Sample preparation, amplification, and detection. This review covers recent advances in on-chip methods for each of these three steps and explains the principles underlying related methods. The sample preparation process is further divided into cell lysis and nucleic acid purification, and methods for the integration of these two steps on a single chip are discussed. Under amplification, on-chip studies based on PCR and isothermal amplification are covered. Three isothermal amplification methods reported to have good resistance to PCR inhibitors are selected for discussion due to their potential for use in direct amplification. Chip designs and novel strategies employed to achieve rapid extraction/amplification with satisfactory efficiency are discussed. Four detection methods providing rapid responses (fluorescent, optical, and electrochemical detection methods, plus lateral flow assay) are evaluated for their potential in rapid on-site detection. In the final section, we discuss strategies to improve the speed of the entire procedure and to integrate all three steps onto a single chip; we also comment on recent advances, and on obstacles to reducing the cost of chip manufacture and achieving mass production. We conclude that future trends will focus on effective nucleic acid extraction via combined methods and direct amplification via isothermal methods., Competing Interests: LP was employed by Zhejiang Hongzheng Testing Co., Ltd., XG was employed by Zhejiang Gongzheng Testing Center Co., Ltd., PL was employed by Key Laboratory of Biosafety detection for Zhejiang Market Regulation, Zhejiang Fangyuan Testing Group LO.T, and YT was employed by Hangzhou Tiannie Technology Co., Ltd. The remaining authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Wang, Jiang, Pan, Gu, Xiao, Liu, Tang, Fang, Li and Lu.)

The white spot syndrome virus is the most destructive virus threatening the shrimp industry worldwide, causing hundreds of millions of dollars in economic losses each year. There is currently no specific medicine to treat it. Therefore, rapid and accurate detection of WSSV is of great significance for controlling its spread and reducing economic losses. Traditional detection methods, such as polymerase chain reaction (PCR) and quantitative fluorescent PCR, rely on laboratory equipment and are not suitable for field testing. In this study, recombinase polymerase amplification (RPA) combined with a lateral flow strip (LFS) was developed. This method targets the entire genome and designs primers and probes accordingly. The detection can be completed in 30 min at 37°C, and the detection limit of each reaction is 20 copies, which is much more sensitive than other detection methods. The RPA-LFS method is highly specific to the white spot syndrome virus and has no cross-reactivity with other common shrimp viruses or pathogens. In total, 100 field samples were tested and compared to the real-time PCR method. Both methods detected 8 positive results, and the positive detection rate was 100%. The method was fast, simple, specific, and sensitive. It does not rely on laboratory equipment and has broad application prospects for in-field detection, especially in remote areas with underdeveloped medical equipment., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Zhang, Liu, Yang, Liu, Yang, Li, Feng, Wu, Jiang, Shen and Dong.)