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1.
Food Res Int ; 193: 114767, 2024 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-39160035

RESUMO

In recent years, foodborne diseases have posed a serious threat to human health, and rapid detection of foodborne pathogens is particularly crucial for the prevention and control of such diseases. This article offers a detailed overview of the development of detection techniques for foodborne pathogens, transitioning from traditional microbiological culture methods to the current array of techniques, including immunological, molecular biological, and biosensor-based methods. It summarizes the technical principles, advantages, disadvantages, and research progress of these diverse methods. Furthermore, the article demonstrates that the combination of different methods enhances the efficiency and accuracy of pathogens detection. Specifically, the article focuses on the application and advantages of combining CRISPR/Cas systems with other detection methods in the detection of foodborne pathogens. CRISPR/Cas systems, with their high specificity, sensitivity, and ease of operation, show great potential in the field of foodborne pathogens detection. When integrated with other detection techniques such as immunological detection techniques, molecular biology detection techniques, and biosensors, the accuracy and efficiency of detection can be further improved. By fully utilizing these tools, early detection and control of foodborne diseases can be achieved, enhancing public health and preventing disease outbreaks. This article serves as a valuable reference for exploring more convenient, accurate, and sensitive field detection methods for foodborne pathogens, promoting the application of rapid detection techniques, and ensuring food safety and human health.


Assuntos
Técnicas Biossensoriais , Microbiologia de Alimentos , Inocuidade dos Alimentos , Doenças Transmitidas por Alimentos , Doenças Transmitidas por Alimentos/microbiologia , Doenças Transmitidas por Alimentos/prevenção & controle , Microbiologia de Alimentos/métodos , Inocuidade dos Alimentos/métodos , Humanos , Técnicas Biossensoriais/métodos , Sistemas CRISPR-Cas , Contaminação de Alimentos/análise
2.
Diagn Microbiol Infect Dis ; 110(2): 116446, 2024 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-39096664

RESUMO

COVID-19 has afflicted millions of lives worldwide. Although there are many rapid methods to detect it based on colorimetric loop-mediated isothermal amplification, there remains room for improvement. This study aims to 1) integrate multiple primers into a singleplex assay to enhance the diagnostic sensitivity, and 2) utilize a high-throughput smartphone-operatable AI-driven color reading tool to enable a rapid result analysis. This setup can improve the sensitivity by 10-100 times and can analyze approximately 6700 samples per minute. The assay is simpler than RT-qPCR, with a turnaround time of less than 75 min. It can detect various types of SARS-CoV-2 by targeting 3 genes, increasing the likelihood that it will remain effective even if the virus undergoes mutations in any single target gene. In summary, it affords potential for adaptation to detection of new/re-emerging diseases with the visual readout for maximum assay simplicity and AI-operated mode for large-scale testing.


Assuntos
COVID-19 , Colorimetria , Técnicas de Diagnóstico Molecular , Técnicas de Amplificação de Ácido Nucleico , SARS-CoV-2 , Sensibilidade e Especificidade , Colorimetria/métodos , Humanos , COVID-19/diagnóstico , COVID-19/virologia , SARS-CoV-2/genética , SARS-CoV-2/isolamento & purificação , Técnicas de Amplificação de Ácido Nucleico/métodos , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Diagnóstico Molecular/normas , Primers do DNA/genética , Teste de Ácido Nucleico para COVID-19/métodos , Smartphone , Teste para COVID-19/métodos
3.
Front Cell Infect Microbiol ; 14: 1424212, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39165916

RESUMO

Introduction: Goose astrovirus (GAstV) is a newly emerging pathogen that is currently widespread among geese, causing visceral gout and leading to substantial gosling mortalities, posing a severe threat to the waterfowl industry. GAstV II is the predominant epidemic strain, characterized by its high morbidity and mortality rate. Consequently, there is an urgent necessity to develop an effective diagnostic approach to control the dissemination of GAstV II, particularly in clinical farms with limited laboratory resources. Methods: In this study, a novel multi-enzyme isothermal rapid amplification (MIRA) and lateral flow dipstick (LFD) combined assay was developed. Different primers designed specific targeting a highly conserved region within the viral RdRp gene for the detection of GAstV II. Primers optimized and MIRA-LFD assay analyzed its performance regarding limits of detection, specificity, and efficiency of detection. Results: The developed MIRA amplification is conducted at a constant temperature and accomplished within 10 minutes. Subsequent naked-eye observation of the LFD strips merely takes 5 minutes. The established MIRA-LFD method exhibits high specificity, with no cross-reaction with other pathogens and attains a detection sensitivity of 1 copy/µl, which is consistent with the reverse transcription quantitative PCR (RT-qPCR) assay. Further evaluation with clinical samples indicates that the accuracy of this MIRA-LFD method correlates well with RT-qPCR for the detection of GAstV II. Conclusion: In summary, the convenience, sensitivity, and rapidity of this newly developed detection method offer a significant advantage for on-site diagnosis of GAstV II.


Assuntos
Infecções por Astroviridae , Gansos , Técnicas de Diagnóstico Molecular , Técnicas de Amplificação de Ácido Nucleico , Doenças das Aves Domésticas , Sensibilidade e Especificidade , Animais , Infecções por Astroviridae/diagnóstico , Infecções por Astroviridae/veterinária , Infecções por Astroviridae/virologia , Gansos/virologia , Técnicas de Amplificação de Ácido Nucleico/métodos , Técnicas de Diagnóstico Molecular/métodos , Doenças das Aves Domésticas/diagnóstico , Doenças das Aves Domésticas/virologia , Avastrovirus/genética , Avastrovirus/isolamento & purificação , Primers do DNA/genética , RNA Viral/genética
4.
Exp Parasitol ; 265: 108813, 2024 Aug 06.
Artigo em Inglês | MEDLINE | ID: mdl-39117169

RESUMO

Babesia duncani, responsible for human babesiosis, is one of the most important tick-borne intraerythrocytic pathogens. Traditionally, babesiosis is definitively diagnosed by detecting parasite DNA in blood samples and examining Babesia parasites in Giemsa-stained peripheral blood smears. Although these techniques are valuable for determining Babesia duncani, they are often time-consuming and laborious. Therefore, developing rapid and reliable B. duncani identification assays is essential for subsequent epidemiological investigations and prevention and control. In this study, a cross-priming amplification (CPA) assay was developed, combined with a vertical flow visualization strip, to rapidly and accurately detect B. duncani infection. The detection limit of this method was as low as 0.98 pg/µl of genomic DNA from B. duncani merozoites per reaction at 59 °C for 60 min. There were no cross-reactions between B. duncani and other piroplasms infective to humans and mammals. A total of 592 blood samples from patients bitten by ticks and experimental infected hamsters were accurately assessed using CPA assay. The average cost of the CPA assay is as low as approximately $ 0.2 per person. These findings indicate that the CPA assay may therefore be a rapid screening tool for detection B. duncani infection, based on its accuracy, speed, and cost-effectiveness, particularly in resource-limited regions with a high prevalence of human babesiosis.

5.
Sci Total Environ ; : 175527, 2024 Aug 14.
Artigo em Inglês | MEDLINE | ID: mdl-39153617

RESUMO

The Yangtze River Estuary (YRE) is one of the areas in China most severely affected by harmful algal blooms (HABs). This study explored the distributive patterns of HABs in the YRE and how they are influenced by the El Niño-Southern Oscillation (ENSO) and other environmental factors. Quantitative real-time PCR (qPCR) was employed to detect and quantify the four predominant HAB species in the YRE, Karenia mikimotoi, Margalefidinium polykrikoides, Prorocentrum donghaiense, and Heterosigma akashiwo. Additionally, the study analyzed how turbidity, pH, salinity, and temperature influence these algae. Distribution of the four HAB species in the YRE area shows clear geographical variations: K. mikimotoi is predominantly found in the northwest and central sea areas, M. polykrikoides (East Asian Ribotype, EAR) is mainly distributed in the southeastern part, P. donghaiense is abundant in the northern regions, and H. akashiwo is especially prevalent at stations S26 and S27 in the northeastern part of the study area. HABs dominated by H. akashiwo and P. donghaiense were observed in the northeastern sea area of the YRE on July 22, 2020. Our study reveals that K. mikimotoi, M. polykrikoides (EAR), and P. donghaiense are mainly affected by turbidity, pH, and salinity, while temperature predominantly influences the blooms of H. akashiwo. Moreover, runoff in the YRE has a certain correlation with ENSO events, which may also impact the nutrient content of the region. The findings of this study illustrate the distributive patterns of the four HAB species under various ecological conditions in the YRE and emphasize the importance of establishing practical cases for future warning systems. To better understand how climate change affects HABs, exploring the link between ENSO and HABs is essential.

6.
Front Plant Sci ; 15: 1442225, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39148615

RESUMO

Rapid detection of plant phenotypic traits is crucial for plant breeding and cultivation. Traditional measurement methods are carried out by rich-experienced agronomists, which are time-consuming and labor-intensive. However, with the increasing demand for rapid and high-throughput testing in tea plants traits, digital breeding and smart cultivation of tea plants rely heavily on precise plant phenotypic trait measurement techniques, among which hyperspectral imaging (HSI) technology stands out for its ability to provide real-time and rich-information. In this paper, we provide a comprehensive overview of the principles of hyperspectral imaging technology, the processing methods of cubic data, and relevant algorithms in tea plant phenomics, reviewing the progress of applying hyperspectral imaging technology to obtain information on tea plant phenotypes, growth conditions, and quality indicators under environmental stress. Lastly, we discuss the challenges faced by HSI technology in the detection of tea plant phenotypic traits from different perspectives, propose possible solutions, and envision the potential development prospects of HSI technology in the digital breeding and smart cultivation of tea plants. This review aims to provide theoretical and technical support for the application of HSI technology in detecting tea plant phenotypic information, further promoting the trend of developing high quality and high yield tea leaves.

7.
Microbiol Spectr ; : e0104424, 2024 Aug 20.
Artigo em Inglês | MEDLINE | ID: mdl-39162506

RESUMO

Carbapenem-resistant Acinetobacter baumannii (CRAB) are increasingly reported worldwide and a leading cause of mortality associated with antimicrobial resistance. Their early detection, particularly in the cases of bloodstream infections, is crucial in attempting to initiate effective antibiotic treatment. The immunochromatographic assay RESIST ACINETO (Coris BioConcept) is a new test developed for the detection of OXA-23, OXA-40/58, and New-Delhi Metallo-beta-lactamase (NDM) carbapenemases in Acinetobacter spp. We evaluated this test on a collection of 121 Acinetobacter spp. clinical isolates, including 104 carbapenemase producers (97 carbapenemases targeted by the test) and 17 non-carbapenemase producers. The performance of the RESIST ACINETO test was evaluated according to the manufacturer's recommendations from bacterial and blood cultures. The strains producing the carbapenemases OXA-23, -40, -58, or/and NDM were accurately detected from bacterial cultures and directly from blood cultures, with the exception of one OXA-23/NDM-1-positive Acinetobacter radioresistens isolate (only detected through standard culture). None of the non-carbapenemase producers tested positive. The RESIST ACINETO test demonstrated sensitivity/specificity of 100%/100% and 99%/100% on bacterial and blood cultures, respectively. IMPORTANCE: The incidence of bloodstream infections with carbapenem-resistant Acinetobacter baumannii (CRAB) could be very high in some countries such as the Balkans or Southeast Asia. In case of positive blood cultures with Gram-negative bacteria, the use of the RESIST ACINETO test could prove highly beneficial for the rapid identification of these imipenem-resistant bacteria and their antibiotic resistance mechanisms. In addition, it is now well established that New-Delhi Metallo-beta-lactamase (NDM) carbapenemase-producing isolates can have increased MICs of cefiderocol, which is an alternative treatment for these infections. This test may also allow the optimization of treatment based on the type of carbapenemase present. Finally, the RESIST ACINETO test is a rapid, easy-to-use, and cost-effective assay that demonstrates excellent performance in detecting the major acquired carbapenemases present in the Acinetobacter species.

8.
Poult Sci ; 103(10): 104141, 2024 Aug 03.
Artigo em Inglês | MEDLINE | ID: mdl-39137501

RESUMO

Rapid and accurate detection of goose parvovirus (GPV) is crucial for controlling outbreaks and mitigating their economic impact on the poultry industry. This study introduces recombinase polymerase amplification combined with the Pyrococcus furiosus argonaute (RPA-PfAgo) system, a novel diagnostic platform designed to address the limitations of traditional GPV detection methods. Capitalizing on the rapid DNA amplification of RPA and stringent nucleic acid cleavage by the PfAgo protein, the RPA-PfAgo system offers high specificity and sensitivity in detecting GPV. Our optimization efforts included primer and probe configurations, reaction parameters, and guided DNA selection, culminating in a detection threshold of 102 GPV DNA copies per microlitre. The specificity of the proposed method was rigorously validated against a spectrum of avian pathogens. Clinical application to lung tissues from GPV-infected geese yielded a detection concordance of 100%, surpassing that of qPCR and PCR in both rapidity and operational simplicity. The RPA-PfAgo system has emerged as a revolutionary diagnostic modality for managing this disease, as it is a promising rapid, economical, and onsite GPV detection method amenable to integration into broad-scale disease surveillance frameworks. Future explorations will extend the applicability of this method to diverse avian diseases and assess its field utility across various epidemiological landscapes.

9.
J Food Sci ; 2024 Aug 13.
Artigo em Inglês | MEDLINE | ID: mdl-39138629

RESUMO

Tomato is sweet and sour with high nutritional value, and soluble solids content (SSC) is an important indicator of tomato flavor. Due to the different mechanisms of nitrogen uptake and assimilation in plants, exogenous supply of different forms of nitrogen will have different effects on the growth, development, and physiological metabolic processes of tomato, thus affecting the tomato flavor. In this paper, hyperspectral imaging (HSI) technique combined with neural network prediction model was used to predict SSC of tomato under different nitrogen treatments. Competitive adaptive reweighed sampling (CARS) and iterative retained information variable (IRIV) were used to extract the feature wavelengths. Based on the characteristic wavelength, the prediction models of tomato SSC are established by custom convolutional neural network (CNN) model that was constructed and optimized. The results showed that the SSC of tomato was negatively correlated with nitrogen fertilizer concentration. For tomatoes treated with different nitrogen concentrations, the residual predictive deviation (RPD) of CARS-CNN and IRIV-parallel convolutional neural networks (PCNN) reached 1.64 and 1.66, both more than 1.6, indicating good model prediction. This study provides technical support for future online nondestructive testing of tomato quality. PRACTICAL APPLICATION: The CARS-CNN and IRIV-PCNN were the best data processing model. Four customized convolutional neural networks were used for predictive modeling. The CNN model provides more accurate results than conventional methods.

10.
Spectrochim Acta A Mol Biomol Spectrosc ; 323: 124889, 2024 Jul 31.
Artigo em Inglês | MEDLINE | ID: mdl-39116595

RESUMO

Pesticide residues are currently a prominent concern for food safety, and the development of a rapid, convenient, and accurate method for detecting pesticide residues is crucial to ensure the quality of agricultural products. In this study, a small molecule fluorescent probe based on biphenyl disulfonic acid (BDSA) was designed and prepared, and a sensitive, specific, and rapid detection method for diquat (DQ) and paraquat (PQ) was developed. The fluorescent molecule (BDSA-NDA) was synthesized through amide reaction between BDSA and 1,8-naphthalic anhydride, which exhibited cyan fluorescence (480 nm) when excited at 305 nm in aqueous solution with a large Stokes shift (>150 nm). Diquat and paraquat were found to quench the fluorescence of the probe through internal filtration effect (IFE) and photoelectron transfer (PET). Moreover, diquat possessed a large conjugated structure that emitted fluorescence at 340 nm which was assembled into a pair of ratio fluorescence with BDSA-NDA. Under optimized experimental conditions, the developed method achieved detection limits of 0.003 mg/L for diquat and 0.202 mg/L for paraquat. Furthermore, it could identify paraquat doped in diquat formulations. Additionally, when applied to environmental water samples as well as rice and urine, this detection method demonstrated good recovery rates (water: 96.2-100.6 %, rice: 93.5-101.9 %, urine: 96-103.7 %), meeting actual sample detection requirements effectively. This work presents a novel approach for rapidly detecting diquat and paraquat residues which holds practical application value in areas such as pesticide residue analysis in foods, environmental or clinical samples.

11.
Molecules ; 29(15)2024 Jul 27.
Artigo em Inglês | MEDLINE | ID: mdl-39124939

RESUMO

The detection of pathogens in medical wastewater is crucial due to the high content of pathogenic microorganisms that pose significant risks to public health and the environment. Medical wastewater, which includes waste from infectious disease and tuberculosis facilities, as well as comprehensive medical institutions, contains a variety of pathogens such as bacteria, viruses, fungi, and parasites. Traditional detection methods like nucleic acid detection and immunological assays, while effective, are often time-consuming, expensive, and not suitable for rapid detection in underdeveloped areas. Electrochemical biosensors offer a promising alternative with advantages including simplicity, rapid response, portability, and low cost. This paper reviews the sources of pathogens in medical wastewater, highlighting specific bacteria (e.g., E. coli, Salmonella, Staphylococcus aureus), viruses (e.g., enterovirus, respiratory viruses, hepatitis virus), parasites, and fungi. It also discusses various electrochemical biosensing techniques such as voltammetry, conductometry, impedance, photoelectrochemical, and electrochemiluminescent biosensors. These technologies facilitate the rapid, sensitive, and specific detection of pathogens, thereby supporting public health and environmental safety. Future research may should pay more attention on enhancing sensor sensitivity and specificity, developing portable and cost-effective devices, and innovating detection methods for diverse pathogens to improve public health protection and environmental monitoring.


Assuntos
Técnicas Biossensoriais , Técnicas Eletroquímicas , Águas Residuárias , Técnicas Biossensoriais/métodos , Técnicas Eletroquímicas/métodos , Águas Residuárias/virologia , Águas Residuárias/microbiologia , Águas Residuárias/análise , Vírus/isolamento & purificação , Bactérias/isolamento & purificação , Humanos , Monitoramento Ambiental/métodos , Fungos/isolamento & purificação
12.
Front Microbiol ; 15: 1429486, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39119142

RESUMO

Background: Porcine deltacoronavirus (PDCoV) is a newly discovered porcine intestinal pathogenic coronavirus with a single-stranded positive-sense RNA genome and an envelope. PDCoV infects pigs of different ages and causes acute diarrhea and vomiting in newborn piglets. In severe cases, infection leads to dehydration, exhaustion, and death in sick piglets, entailing great economic losses on pig farms. The clinical symptoms of PDCoV infection are very similar to those of other porcine enteroviruses. Although it is difficult to distinguish these viral infections without testing, monitoring PDCoV is very important because it can spread in populations. The most commonly used methods for the detection of PDCoV is qPCR, which is time-consuming and require skilled personnel and equipment. Many farms cannot meet the conditions required for detection. Therefore, it is necessary to establish a faster and more convenient method for detecting PDCoV. Aims: To establish a rapid and convenient detection method for PDCoV by combining RPA (Recombinase Polymerase Isothermal Amplification) with CRISPR/Cas13a. Methods: Specific RPA primers and crRNA for PDCoV were designed, and the nucleic acids in the samples were amplified with RPA. Fluorescent CRISPR/Cas13a detection was performed. We evaluated the sensitivity and specificity of the RPA-CRISPR/Cas13a assay using qPCR as the control method. Results: CRISPR/Cas13a-assisted detection was completed within 90 min. The minimum detection limit of PDCoV was 5.7 × 101 copies/µL. A specificity analysis showed that the assay did not cross-react with three other porcine enteroviruses. Conclusion: The RPA-CRISPR/Cas13a method has the advantages of high sensitivity, strong specificity, fast response, and readily accessible results, and can be used for the detection of PDCoV.

13.
Int J Biol Macromol ; 277(Pt 3): 134485, 2024 Aug 03.
Artigo em Inglês | MEDLINE | ID: mdl-39102926

RESUMO

The blood glucose concentration in aquatic organisms, a crucial indicator reflecting their health status, holds significant importance for detecting glucose levels in serum in terms of processing and quality monitoring. In this study, a novel POD biomimetic enzyme (p-BEs) with horseradish peroxidase catalytic properties was designed, optimized, and its mechanism was discussed in detail. Based on this, a portable system has been developed capable of determining glucose levels in three ways: quantitatively analyzed through UV-Vis/MD, quantitatively analyzed on-site using a mobile phone RGB, and semi-quantitatively analyzed through a drip plate. Meanwhile, compared with other catalytic methods for detecting glucose, we achieved a lower limit of detection (0.03 µM) and shorter detection time (12 min), with high catalytic activity. This study provides new insights into the design of efficient and reliable cascade catalytic systems responsive to glucose, offering a low-cost, simplicity of operation method for glucose detection.

14.
Front Cell Infect Microbiol ; 14: 1430302, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39099883

RESUMO

Introduction: Human cytomegalovirus (HCMV) is the most common viral infection seen in newborns. The major route of transmission for acquired human cytomegalovirus infection is breast milk from mothers who are HCMV seropositive to the infants. Thus, a rapid, economical, and simple method to perform HCMV test in breast milk is crucial and necessary for preventing acquired HCMV infection, especially in underdeveloped regions with limited laboratory resources. Methods: In this study, an effective technique for the detection of HCMV was constructed by combining multienzyme isothermal rapid amplification (MIRA) and lateral flow chromatography strip (LFD). Primers for the conserved HCMV sequence UL83 were utilized for MIRA-LFD testing. Results: Our results showed that the entire MIRA reaction could be completed in 12 minutes at 37°C, and LFD outcomes could be observed visibly after 10 minutes. The detection sensitivity of this method reached 50 copy/µl. Samples of breast milk were examined to compare MIRA-LFD and conventional qPCR. The accuracy of MIRA-LFD was 100%. Discussion: The straightforward, rapid, economic features of the test can provide the significant advantages for the prevention of breast milk-acquired cytomegalovirus infection, particularly in resource-limited locations with high seroprevalence of cytomegalovirus.


Assuntos
Infecções por Citomegalovirus , Citomegalovirus , Leite Humano , Técnicas de Diagnóstico Molecular , Técnicas de Amplificação de Ácido Nucleico , Sensibilidade e Especificidade , Humanos , Citomegalovirus/genética , Citomegalovirus/isolamento & purificação , Leite Humano/virologia , Infecções por Citomegalovirus/diagnóstico , Infecções por Citomegalovirus/virologia , Técnicas de Amplificação de Ácido Nucleico/métodos , Técnicas de Diagnóstico Molecular/métodos , Feminino , Recém-Nascido , Fatores de Tempo
15.
Spectrochim Acta A Mol Biomol Spectrosc ; 323: 124886, 2024 Jul 28.
Artigo em Inglês | MEDLINE | ID: mdl-39089069

RESUMO

Mercury, one of the various harmful metals, is particularly significant in affecting aquatic organisms, currently gaining more attentions and sparking discussions. In response to the limitations of traditional detections, fluorescent probes have emerged as a promising solution with some advantages, such as weaker background interference, shorter processing time, higher accuracy. Thus, a novel fluorescent probe, FS-Hg-1, has been developed for assessing mercury ion (Hg2+) concentrations in aquatic products. This probe displays specific recognition of mercury ions in fluorescence spectra. Notably, FS-Hg-1 exhibits a distinct color change to pink when combined with Hg2+ (with a 948-fold increase in absorption at 568 nm) and a substantial fluorescence change towards Hg2+ (361-fold increase, excitation at 562 nm, emission at 594 nm) in N, N-dimethylformamide. The probe boasts a detection limit of 0.14 µM and rapid reaction with Hg2+ within 10 s, showing an excellent linear correlation with [Hg2+] in the range of 0 to 10 µM. Through thorough analysis using FS-Hg-1, the results align with those from the standard method (P > 0.05), with spiked recovery rates ranging from 108.4% to 113.2%. With its precise recognition, low detection limit, and remarkable sensitivity, this fluorescent assay proves effective in mercury concentration determination in aquatic samples without interference. The potential of FS-Hg-1 is promising for speedy detection of residual Hg2+ and holds significance in ensuring food safety.

16.
Infect Med (Beijing) ; 3(2): 100111, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38948389

RESUMO

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 (102 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.

17.
Anal Chim Acta ; 1317: 342872, 2024 Aug 15.
Artigo em Inglês | MEDLINE | ID: mdl-39030016

RESUMO

Fluorescently labeled DNA oligonucleotides and gold nanospheres have been frequently utilized in biosensors, providing efficient nucleic acid detection. Nevertheless, the restricted loading capacity of gold nanospheres undermines overall sensitivity. In this study, we employed four-atom-thick ultrathin gold nanosheets (AuNSs), utilizing a "pre-mix model" for rapid target nucleic acid detection. In this approach, fluorescently labeled DNA probes were pre-incubated with the target nucleic acid, followed by the addition of AuNSs for probe adsorption and fluorescence quenching. With the developed method, we efficiently and rapidly detected the SARS-CoV-2 N gene sequence within 30 min, involving a brief 15-min target pre-incubation and a subsequent 15-min adsorption of free probes and fluorescence quenching by AuNSs. This method exhibited heightened sensitivity compared to gold nanospheres, boasting a limit of detection (LOD) of 0.808 nM. Furthermore, exceptional recovery was achieved in simulated biological samples. The study introduces an effective strategy for nucleic acid sensing characterized by rapidity, heightened sensitivity, ease of operation, and robustness. These findings encourage further development of rapid biomarker sensing methods employing 2D nanomaterials.


Assuntos
Técnicas Biossensoriais , Corantes Fluorescentes , Ouro , Limite de Detecção , Nanopartículas Metálicas , SARS-CoV-2 , Ouro/química , SARS-CoV-2/isolamento & purificação , Corantes Fluorescentes/química , Técnicas Biossensoriais/métodos , Nanopartículas Metálicas/química , Humanos , Espectrometria de Fluorescência , Nanoestruturas/química , Sondas de DNA/química , COVID-19/diagnóstico , COVID-19/virologia
18.
Artigo em Inglês | MEDLINE | ID: mdl-39053111

RESUMO

An imidacloprid colloidal gold immunochromatographic strip was developed in this work, and systematic analytical conditions were deeply investigated. The test strips were used for rapid screening of imidacloprid residues in Chinese herbal medicines. The performance of the colloidal gold test strips was investigated by using five selected Chinese herbal medicines (malt, Coix seed, lotus seed, dried ginger and honeysuckle). As a result, the developed imidacloprid colloidal gold immunochromatographic test strips could be used for rapid screening of imidacloprid residues in 60 kinds of different herbs (including 26 kinds of root/rhizome medicines, 20 kinds of seed/fruit/pericarp medicines, 11 kinds of flower/leaf/whole herb medicines, and 3 kinds of bark/aboveground issues of herb medicines), and the cut-off value was 50 µg/kg. The development of this method can achieve the goal of on-site, rapid and low-cost screening of imidacloprid residues in different herbs, which is of great significance for the quality assurance of herbs.


Assuntos
Cromatografia de Afinidade , Medicamentos de Ervas Chinesas , Coloide de Ouro , Limite de Detecção , Neonicotinoides , Nitrocompostos , Neonicotinoides/análise , Nitrocompostos/análise , Medicamentos de Ervas Chinesas/química , Medicamentos de Ervas Chinesas/análise , Coloide de Ouro/química , Cromatografia de Afinidade/métodos , Reprodutibilidade dos Testes , Resíduos de Praguicidas/análise , Modelos Lineares
19.
Curr Drug Deliv ; 21(10): 1285-1299, 2024.
Artigo em Inglês | MEDLINE | ID: mdl-39034714

RESUMO

The field of microfluidics encompasses the study of fluid behavior within micro-channels and the development of miniature systems featuring internal compartments or passageways tailored for fluid control and manipulation. Microfluidic devices capitalize on the unique chemical and physical properties exhibited by fluids at the microscopic scale. In contrast to their larger counterparts, microfluidic systems offer a multitude of advantages. Their implementation facilitates the investigation and utilization of reduced sample, solvent, and reagent volumes, thus yielding decreased operational expenses. Owing to their compact dimensions, these devices allow for the concurrent execution of multiple procedures, leading to expedited experimental timelines. Over the past two decades, microfluidics has undergone remarkable advancements, evolving into a multifaceted discipline. Subfields such as organ-on-a-chip and paper-based microfluidics have matured into distinct fields of study. Nonetheless, while scientific progress within the microfluidics realm has been notable, its translation into autonomous end-user applications remains a frontier to be fully explored. This paper sets forth the central objective of scrutinizing the present research paradigm, prevailing limitations, and potential prospects of customizable microfluidic devices. Our inquiry revolves around the latest strides achieved, prevailing constraints, and conceivable trajectories for adaptable microfluidic technologies. We meticulously delineate existing iterations of microfluidic systems, elucidate their operational principles, deliberate upon encountered limitations, and provide a visionary outlook toward the future trajectory of microfluidic advancements. In summation, this work endeavors to shed light on the current state of microfluidic systems, underscore their operative intricacies, address incumbent challenges, and unveil promising pathways that chart the course toward the next frontier of microfluidic innovation.


Assuntos
Dispositivos Lab-On-A-Chip , Humanos , Microfluídica/instrumentação , Microfluídica/tendências , Técnicas Analíticas Microfluídicas/instrumentação , Desenho de Equipamento/tendências
20.
Int J Food Microbiol ; 422: 110822, 2024 Sep 16.
Artigo em Inglês | MEDLINE | ID: mdl-39013210

RESUMO

Foodborne illnesses, caused by harmful microorganisms in food, are a significant global health issue. Current methods for identifying these pathogens are both labor-intensive and time-consuming. In this research, we devised a swift and precise detection technique using recombinase polymerase amplification combined with a lateral flow dipstick (RPA-LFD) for three foodborne pathogens found in meat. By employing a dedicated detection device, RPA-LFD allows for the rapid analysis of DNA from Escherichia coli O157 (E. coli O157), Salmonella, and Shigella-pathogens that are prohibited in food. The detection thresholds for E. coli O157, Salmonella, and Shigella are 0.168 fg/µl (1.04 CFU/ml), 0.72 fg/µl (27.49 CFU/ml), and 1.25 fg/µl (48.84 CFU/ml), respectively. This method provides a short detection window, operates at low temperatures, follows simple procedures, and exhibits high sensitivity. Our study establishes the RPA-LFD method for simultaneously identifying the nucleic acid of three foodborne pathogens, offering an efficient solution for quickly identifying multiple contaminants.


Assuntos
Escherichia coli O157 , Contaminação de Alimentos , Microbiologia de Alimentos , Técnicas de Amplificação de Ácido Nucleico , Recombinases , Salmonella , Shigella , Escherichia coli O157/isolamento & purificação , Escherichia coli O157/genética , Salmonella/isolamento & purificação , Salmonella/genética , Técnicas de Amplificação de Ácido Nucleico/métodos , Microbiologia de Alimentos/métodos , Recombinases/metabolismo , Shigella/isolamento & purificação , Shigella/genética , Contaminação de Alimentos/análise , Carne/microbiologia , DNA Bacteriano/genética , Animais , Sensibilidade e Especificidade , Doenças Transmitidas por Alimentos/microbiologia
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