On-site rapid detection of multiple pesticide residues in tea leaves by lateral flow immunoassay

The application of pesticides (mostly insecticides and fungicides) during the tea-planting process will undoubtedly increase the dietary risk associated with drinking tea. Thus, it is necessary to ascertain whether pesticide residues in tea products exceed the maximum residue limits. However, the complex matrices present in tea samples comprise a major challenge in the analytical detection of pesticide residues. In this study, nine types of lateral flow immunochromatographic strips (LFICSs) were developed to detect the pesticides of interest (fenpropathrin, chlorpyrifos, imidacloprid, thiamethoxam, acetamiprid, carbendazim, chlorothalonil, pyraclostrobin, and iprodione). To reduce the interference of tea substrates on the assay sensitivity, the pretreatment conditions for tea samples, including the extraction solvent, extraction time, and purification agent, were optimized for the simultaneous detection of these pesticides. The entire testing procedure (including pretreatment and detection) could be completed within 30 min. The detected results of authentic tea samples were confirmed by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS), which suggest that the LFICS coupled with sample rapid pretreatment can be used for on-site rapid screening of the target pesticide in tea products prior to their market release.

Rapid Detection of Somatic Cell Count Based on Hybrid Variable Selection Method

Somatic cell count detection is the daily work of dairy farms to monitor the health of cows.The feasibility of applying near-infrared spectroscopy to somatic cell count detection was researched in this paper.Milk samples with different somatic cell counts were collected and preprocessing methods were studied.Variable selection algorithm based on hybrid strategy and modelling method based on ensemble learning were explored for somatic cell count detection.Detection model was used to diagnose subclinical mastitis and the results showed that near-infrared spectroscopy could be a tool to realize rapid detection of somatic cell count in milk.

Recent Advances in the Rapid Detection and Performance Evaluation Methods of Detergent Additives for Gasoline

Although detergent additives for gasoline have been widely commercialized,their formulas are often kept confidential and there is still no standardized method for quickly detecting the main active ingredients and evaluating their effectiveness,which makes their regulation difficult.An overview of the current state of the development and application of detergent additives for gasoline in China and other regions,as well as a review of the rapid detection and performance evaluation methods available for analyzing detergent additives are given herein.The review focuses on the convenience,cost,efficiency,and feasibility of on-site detection and the evaluation of various methods,and also looks into future research directions,such as detecting and evaluating detergent additives in ethanol gasoline and with advanced engine technologies.

Establishment of a rapid detection method of Ureaplasma urealyticum based on recombinant polymerase amplification

Ureaplasma urealyticum(UU),is one of the most vital pathogens causing genitourinary tract infections of the body,and it can result in poor maternal and perinatal outcomes.The aim of this study was to establish a method to detect Ureaplasma urealyticum based on recombinant polymerase amplification(RPA)technique.Specific primers and probes were designed according to the 16sRNA gene sequence of Ureaplasma urealyticum.Six pathogens were detected for real-time fluorescence RPA specificity verification,including Mycoplasma hominis(MH),Chlamydia trachomatis(CT),Neisseria gonorrhoeae(NG),Staphylococcus aureus,Escherichia coli,and Lactobacillus vaginalis.The sensitivity of the method was performed by gradient dilution of the extracted template.A total of 60 clinical samples were detected by the established real-time fluorescence RPA.Detection of Ureaplasma urealyticum can be completed within 20 minutes at 39°C using established RPA method.The minimum detection limit of Ureaplasma urealyticum by real-time fluorescence RPA was 3 pg.The evaluation of 60 clinical samples proved that RPA method was feasible.A high specificity,sensitivity,simplicity and rapidity method for Ureaplasma urealyticum detection was successfully established based on the real-time fluorescence RPA method.

The Application of Reverse Transcription-loop-mediated Isothermal Amplification for the Rapid Detection of Maize Chlorotic Dwarf Virus

Maize chlorotic dwarf virus （MCDV） is a quarantine pest as approved by Chinese government. A rapid, sensitive and specific MCDV detection method using reverse transcription-loop-mediated isothermal amplification （RT-LAMP） was estab- lished in this study. Based on the sequence of MCDV coat protein coding gene, specific primers were designed and similar sensitivities were observed between RT- LAMP and RT-PCR, except that RT-LAMP was quicker, and the reaction could be finished within 1 h. In addition, the presence or absence of the fluorescent display in daylight allows naked easy detection of the amplification of MCDV genomic RNA using calcein. The RT-LAMP assay was applied successfully to detect MCDV in maize seeds, and the result by the addition of calcein was consistent with the result detected by the real time turbidimeter.

Deterioration mechanism and rapid detection of performances of an existing subgrade in southern China

To relieve the increasing traffic load, many early built highways need to be widened or reconstructed. The rapid performance detection to existing subgrades is important to their reasonable evaluation and maximized utilization. Based on five kinds of soils taken from an existing highway in southern China, three commonly detecting methods were used to determine their moisture contents, compaction degrees and resilient moduli. The results showed that the measured moisture contents were greater than the design value, and the compaction degrees decreased sharply compared to the original ones. The moisture and heat exchange produced a decrease in the resilient modulus of plate loading test(PLT) from the standard 60 MPa down to 40 MPa. Afterwards, the portable falling weight deflectometer(PFWD) and dynamic cone penetrometer(DCP) were used to evaluate the subgrade performances. The measured PFWD moduli and the DCP penetration rates were correlated with the resilient moduli of PLT, deflections of the Beckman beam test, compaction degrees and moisture contents. The correlation analysis indicates that both of two methods are suitable in rapid detecting subgrade performances, but PFWD method is more recommended for it has higher accuracy and efficiency.

Rapid detection of Pseudomonas aeruginosa by cross priming amplification

Pseudomonas aeruginosa(PA)is an opportunistic pathogen of humans and animals and a common source of nosocomial infections especially of the respiratory tract.Pseudomonas aeruginosa is also a major bacterial disease of poultry and in particular,eggs and newly hatched chicks.In this study,we developed a simple,accurate and rapid molecular detection method using cross priming amplification(CPA)with a nucleic acid test strip to detect P.aeruginosa.The assay efficiently amplified the target gene within 45 min at 62℃only using a simple water bath.The detection limit of the method was 1.18x 102 copiesμL^-1 for plasmid DNA and 4.4 CFU mL^-1 for bacteria in pure culture,and was 100 times more sensitive than conventional PCR.We screened 83 clinical samples from yellow-feather broiler breeder chickens and hospitalized/treated dogs and cats using CPA,PCR and traditional culture methods.The positive sample ratios were 15.3%(13/83)by CPA,13.3%(11/83)by PCR and 12.1%(10/83)by the culture method.The established CPA method has significant advantages for detecting P.aeruginosa.The method is easy to use and possesses high specificity and sensitivity without the requirements of complicated experimental equipment.The PA-CPA assay is especially fit for outdoor and primary medical units and is an ideal system for the rapid detection and monitoring of P.aeruginosa.

In vitro Selection of DNA Aptamers and Fluorescence-Based Recognition for Rapid Detection Listeria monocytogenes

Aptamers are specific nucleic acid sequences that can bind to a wide range of nucleic acid and non-nucleic acid targets with high affinity and specificity. Nucleic acid aptamers are selected in vitro from single stranded DNA or RNA ligands containing random sequences of up to a few hundred nucleotides. Systematic evolution of ligands by exponential enrichment （SELEX） was used to select and PCR amplify DNA sequences （aptamers） capable of binding to and detecting Listeria monocytogenes, one of the major food-borne pathogens. A simplified affinity separation approach was employed, in which L. monocytogenes in exponential （log） phase of growth was used as the separation target. A fluorescently-labeled aptamer assay scheme was devised for detecting L. monoeytogenes. This report described a novel approach to the detection of L. monocytogenes using DNA aptamers. Aptamers were developed by nine rounds of SELEX. A high affinity aptamer was successfully selected from the initial random DNA pool, and its secondary structure was also investigated. One of aptamers named e01 with the highest affinity was further tested in aptamer-peroxidase and aptamer-fluorescence staining protocols. This study has proved the principle that the whole-cell SELEX could be a promising technique to design aptamer-based molecular probes for dectection of pathogenic microorganisms without tedious isolation and purification of complex markers or targets.

Enzyme Inhibition Rate Method for Rapid Detection of Organophosphorus and Carbamate Pesticides in Cowpea

[Objectives ] The paper was to explore enzyme inhibition rate method for rapid detection of organophosphorus and carbamate pesticides in cowpea. [ Methods ] Acetylcholinesterase （ACHE） was added to cowpea extract, to determine the inhibition rate of extract against enzyme. The influences of different sampiing methods and sampling parts on detection results were compared. [ Results] The positive rate of standard sampling was 18.18% higher than that of non-stand- ard sampling, and the positive rate of samples collected from cowpea tail was 16.67% higher than that collected from other parts. [ Condmions] Enzyme inhibi- tion rate method is suitable for rapid detection of organophosphorus and carbamate pesticides in cowpea.

A Device for Rapid Detection of Dithiocarbamate Pesticide Residues in Fruits

Based on the chemical properties of dithiocarbamate pesticides,a device for rapid detection was developed in the paper,and the experimental conditions were optimized. Dithiocarbamate residues in fruits were successfully detected using molecular absorption spectro-photometry,and the recovery rate was over 80%.The rapid detection method was simple to operate with low cost,and was conducive to application in basic level and enterprise laboratories.

Dengue virus infection:A review of advances in the emerging rapid detection methods

Dengue virus infections are increasing worldwide generally and in Asia,Central and South America and Africa,particularly.It poses a serious threat to the children population.The rapid and accurate diagnostic systems are essentially required due to lack of effective vaccine against dengue virus and the progressive spread of the dengue virus infection.The recent progress in developing micro-and nano-fabrication techniques has led to low cost and scale down the biomedical point-of-care devices.Starting from the conventional and modern available methods for the diagnosis of dengue infection,this review examines several emerging rapid and point-of-care diagnostic devices that hold significant potential for the progress in smart diagnosis tools.The given review revealed that an effective vaccine is required urgently against all the dengue virus serotypes.However,the rapid detection methods of dengue virus help in early treatment and significantly reduce the dengue virus outbreak.

Rapid Detection of Fat Content in Chenopodium quinoa Willd by Near Infrared Spectroscopy

This study was conducted to find a method for rapid determination of fat content in complete quinoa （ Chenopodium quinoa Willd） seeds. The near infrared spectra of 100 quinoa samples were collected, and a mathematic model was built using the near infrared spectra, so as to perform prediction. The results showed that within the wavelength range of 1 0 000-4 000 cm ^-1 , the quantification model of fat content built by first derivative ＋vector normalization spectral pre-processing had better calibration and prediction effects, and showed a determination coefficient of cross validation （ r cv^ 2 ） of 0.939 3 and a determination coefficient of validation （ rval^2 ） of 0.923 5. The near infrared spectral model of fat could be used for rapid detection of fat contents in quinoa.

Establishment of Recombinase Polymerase Amplification for Rapid Detection of Spring Viremia of Carp Virus(SVCV)

[Objective]The paper was to develop a rapid method for the detection of spring Viremia of carp virus(SVCV).[Method]The specific primers were designed by targeting the G gene of SVCV.The recombinase polymerase amplification(RPA)assay for detecting SVCV was estab-lished by optimizing the reaction conditions.The optimal amplification temperature of RPA assay was 30℃,and the test could be finished within 20 min.[Result]The method was specific with no cross-reaction with other common fish infectious viruses.Sensitivity test showed that the lowest detection limit of the method was 89.2 copies/μL,higher than that of traditional RT-PCR.Moreover,a total of 80 clinical samples were detected by RPA and RT-PCR,respectively.The weak positive samples tested by RT-PCR could be detectable with RPA,indicating that RPA assay could be used in clinical detection.[Conclusion]The method established is rapid,simple,specific and sensitive for testing SVCV,and it will be widely used in grassroots laboratory and on-site inspection.

Synthetic Biology Construct of Ebola Virus in Bacteria Surrogate Is Stable and Safe for Rapid Detection Studies in a BSL-2 Laboratory Setting

Rapid detection of virulent pathogens during an outbreak is critical for public health advisories and control of the disease in a population. While many molecular techniques for point of care and clinical diagnosis abound, the US experience with the COVID-19 testing in the early stages of the pandemic underscores the critical importance of determining the appropriate target gene(s) with in-built controls that reliably detect pathogens with high sensitivity and specificity. Assays and research for diagnostics and therapy could be slowed during an epidemic because access to the required BSL-3 and BSL-4 laboratories are limited. So, during the 2014 West Africa Ebola outbreak, we tested the hypothesis that using synthetic cDNA of Ebolavirus in a bacteria surrogate (fit for all lab settings), would remain unmutated and safe after several generations, serving as an effective positive control in research settings, self test and point-of-care detection platforms. Primers were designed for the detection and quantification of the nucleoprotein (NP) gene of the 2014 Makona Ebola strain (KR781608.1, 733 - 1332 bp). To test the stability of artificially inserted translation arrest in the Orf of the model gene, it was edited to include three STOP codons in the RNA transcript using SNAP GENE. The segment was then spliced into a high copy number plasmid, cloned into One ShotTM TOP10 Escherichia coli (Invitrogen), and tested for stability and safety by periodic subculture, extraction and sequencing. Unlike COVID-19, rapid detection of blood-borne etiologies like Ebola requires optimized protocols for blood matrix. Using real-time PCR and newly designed primer pairs, the EBOV surrogate was detected and enumerated in human blood and regular broth and buffers. Based on aligned sequence analysis, the EBOV synthetic NP gene was stable (>99.9999% similarity coefficient) for at least 3 months. Detection sensitivity in broth and blood was at least 100 cells/ml or about 5.8 × 103 to 7.3 × 103 virion equivalents per ml. While the developments of transcription-and-replication-competent virus like particles (trVLP) have made it possible to study the infection and replication cycles of virulent pathogens in BSL-2 laboratories, the simplicity of our model and the reproducibility of detection and enumeration show the utility of synthetic bio-components as positive controls for point of care diagnostic tools. The inserted stop codons remained intact after many generations, suggesting that expressed virulent proteins can be easily silenced in synthetic biology models for research in BSL-1 and 2 and a wide range of pathogens. Synthetic bio-components can thereby aid further research by reducing costs and improving safety for workers and stakeholders.

Establishment of High-sensitivity Rapid Fluorescence Quantitative Detection Method for Antibody against Peste des Petits Ruminants Virus

[Objectives]This study was conducted to establish a rapid quantitative method for detecting antibody against Peste des Petits Ruminants Virus(PPR V)in sheep serum.[Methods]Soluble N protein and NH fusion protein were obtained in Escherichia coli prokaryotic expression system by optimizing codons and expression conditions of E.coli.Furthermore,based on the purified soluble N protein and NH fusion protein,a high-sensitivity fluorescence immunoassay kit for detecting the antibody against PPR V was established.[Results]The method could quickly and quantitatively detect PPR V antibody in sheep serum,with high sensitivity and specificity,without any cross reaction to other related sheep pathogens.The intra-batch and inter-batch coefficients of variation were less than 10%and 15%,respectively,and the method had good repeatability.Through detection on 292 clinical serum samples,it was compared with the French IDVET competitive ELISA kit,and the coincidence rate of the two methods reached 93.84%.Compared with the serum neutralization test,the detected titer value of the high-sensitivity rapid fluorescence quantitative detection method was basically consistent with the tilter value obtained by the neutralization test on the standard positive serum(provided by the WOAH Brucellosis Reference Laboratory of France).[Conclusions]This method can realize rapid quantitative detection of PPR V antibody on site,and has high practical value and popularization value.

ZnO/PANI nanoflake arrays sensor for ultra-low concentration and rapid detection of NO_(2)at room temperature

With the development of environmental monitoring,it is urgent to establish NO_(2)sensor with good sensing performance.Compared with the traditional NO_(2)sensors made of metal oxides,NO_(2)sensors made of n-p heterostructure nanocomposites have good sensing performance in detection limit and operating temperature.ZnO nanoflake arrays with polyaniline film grown on the surface were prepared on ceramic tubes by hydrothermal and vapor diffusion method.The gas-phase diffusion method can control the heterostructure by adjusting the diffusion time.At room temperature(25℃),the construction of rich n-p heterogeneous interface enables the sensor prepared by the nanocomposite to respond to NO_(2),showing the sensing performance with the response value of 28.00 to10.00×10^(-6)NO_(2);the detection limit improved to0.01×10^(-6)and the recovery time of 18 s.In this work,the sensing mechanism of NO_(2)at heterogeneous interface is analyzed,which provides a promising material for the detection of low concentration NO_(2)at room temperature.

Filtration assisted pretreatment for rapid enrichment and accurate detection of Salmonella in vegetables

Rapid detection of target foodborne pathogens plays more and more significant roles in food safety,which requires the efficiency,sensitivity,and accuracy.In this research,we proposed a new st rategy of isothermal-molecular-amplification integrated with lateral-flow-strip for rapid detection of Salmonella without traditional enrichment-culture.Th e designed syringe-assisted-filtration can contribute to simultaneous collection and concentration of target bacterium from vegetable samples in just 3 min,resolving the drawbacks of traditional random sampling protocols.After simple and convenient ultrasonication,samples can be directly amplified at 39℃ in 25 min and the amplicons are qualitatively and quantitatively analyzed with the designed lateral-flow-strip in 5 min.Finally,satisfied results have been achieved within 40 min,which greatly improve the efficiency while the accuracy is also guaranteed.Furthermore,all detection steps can be completed under instrument-free conditions.This method will hold great promise for target pathogen detection in the resource-limited district,or for emergency on-site identification.

Establishment and application of a rapid visualization method for detecting Vibrio parahaemolyticus nucleic acid

Background:Swift and accurate detection of Vibrio parahaemolyticus,which is a prominent causative pathogen associated with seafood contamination,is required to effectively combat foodborne disease and wound infections.The toxR gene is relatively conserved within V.parahaemolyticus and is primarily involved in the expression and regulation of virulence genes with a notable degree of specificity.The aim of this study was to develop a rapid,simple,and constant temperature detection method for V.parahaemolyticus in clinical and nonspecialized laboratory settings.Methods:In this study,specific primers and CRISPR RNA were used to target the toxR gene to construct a reaction system that combines recombinase polymerase amplification(RPA)with CRISPR‒Cas13a.The whole-genome DNA of the sample was extracted by self-prepared sodium dodecyl sulphate(SDS)nucleic acid rapid extraction reagent,and visual interpretation of the detection results was performed by lateral flow dipsticks(LFDs).Results:The specificity of the RPA-CRISPR/Cas13a-LFD method was validated using V.parahaemolyticus strain ATCC-17802 and six other non-parahaemolytic Vibrio species.The results demonstrated a specificity of 100%.Additionally,the genomic DNA of V.parahaemolyticus was serially diluted and analysed,with a minimum detectable limit of 1 copy/μL for this method,which was greater than that of the TaqMan-qPCR method(10^(2) copies/μL).The established methods were successfully applied to detect wild-type V.parahaemolyticus,yielding results consistent with those of TaqMan-qPCR and MALDI-TOF MS mass spectrometry identification.Finally,the established RPA-CRISPR/Cas13a-LFD method was applied to whole blood specimens from mice infected with V.parahaemolyticus,and the detection rate of V.parahaemolyticus by this method was consistent with that of the conventional PCR method.Conclusions:In this study,we describe an RPA-CRISPR/Cas13a detection method that specifically targets the toxR gene and offers advantages such as simplicity,rapidity,high specificity,and visual interpretation.This method serves as a valuable tool for the prompt detection of V.parahaemolyticus in nonspecialized laboratory settings.

A Rapid and Simple Method for the Detection of Rice Dwarf Virus by Aqueous Extract

Rice dwarf disease caused by rice dwarf virus （RDV） is one of the major rice virus diseases in China,which widely distributes in the rice area of China.A simple and rapid method for detection of RDV in rice plants and Nephotettix cincticeps was investigated in this study,and the whole detection only lasted for 20 min.After diseased leaves of rice （10 mg） and leaves of Nephotettix cincticeps （10 mg） both infected by RDV were ground by sterile water （100 μl）,the supernatant was analyzed by 1% agarose gel electrophoresis to steadily detect five specific electrophoretic bands with 1 000 bp to 5 000 bp.However,there was no electrophoretic band in the healthy rice leaves and aqueous extract of non-viruliferous N.cincticeps.Therefore,this method was more rapid and simple to detect RDV in rice plants and Nephotettix cincticeps,which avoided expensive reagents and tedious steps of conventional serological testing and molecular detection （RT-PCR and Western-blot）.

Safety issues and new rapid detection methods in traditional Chinese medicinal materials

The safety of traditional Chinese medicine(TCM) is a major strategic issue that involves human health. With the continuous improvement in disease prevention and treatment, the export of TCM and its related products has increased dramatically in China. However, the frequent safety issues of Chinese medicine have become the ‘bottleneck' impeding the modernization of TCM. It was proved that mycotoxins seriously affect TCM safety; the pesticide residues of TCM are a key problem in TCM international trade; adulterants have also been detected, which is related to market circulation. These three factors have greatly affected TCM safety. In this study, fast, highly effective, economically-feasible and accurate detection methods concerning TCM safety issues were reviewed, especially on the authenticity, mycotoxins and pesticide residues of medicinal materials.