scholarly journals Recombinase-Aided Amplification Coupled with Lateral Flow Dipstick for Efficient and Accurate Detection of Porcine Parvovirus

Life ◽  
2021 ◽  
Vol 11 (8) ◽  
pp. 762
Author(s):  
Yihong He ◽  
Wenxian Chen ◽  
Jindai Fan ◽  
Shuangqi Fan ◽  
Hongxing Ding ◽  
...  
Keyword(s):  
Detection Method ◽  
Cross Reactivity ◽  
Lateral Flow ◽  
Porcine Parvovirus ◽  
Swine Industry ◽  
Resource Limited ◽  
Efficient Detection ◽  
Lateral Flow Dipstick ◽  
Polymerase Chain ◽  
Embryonic Death

Porcine parvovirus (PPV) infection is the primary cause of SMEDI (stillbirth; mummification; embryonic death; infertility) syndrome, which is a global burden for the swine industry. Thus, it is crucial to establish a rapid and efficient detection method against PPV infection. In the present work, we developed a recombinase-aided amplification (RAA) assay, coupled with a lateral flow dipstick (LFD), to achieve an amplification of PPV DNA at 37 °C within 15 min. The detection limits of PPV RAA-LFD assay were 102 copies/μL recombinant plasmid pMD19-T-VP1, 6.38 × 10−7 ng/μL PPV DNA, and 10−1 TCID50/mL virus, respectively. This method was highly specific for PPV detection with no cross-reactivity for other swine pathogens. In contrast to polymerase chain reaction (PCR), the PPV RAA-LFD assay is more sensitive and cost-saving. Hence, the established PPV RAA-LFD assay provided an alternative for PPV detection, especially in resource-limited regions.

Rapid and visual detection of Meloidogyne hapla using recombinase polymerase amplification combined with a lateral flow dipstick (RPA-LFD) assay

Plant Disease ◽  
2020 ◽  
Author(s):  
Zhiqiang Song ◽  
Xiai Yang ◽  
Xiaowei Zhang ◽  
Mingbao Luan ◽  
Bing Guo ◽  
...  
Keyword(s):  
Rapid Detection ◽  
Lateral Flow ◽  
Recombinase Polymerase Amplification ◽  
Economic Losses ◽  
Meloidogyne Hapla ◽  
Root Knot Nematode ◽  
Resource Limited ◽  
Lateral Flow Dipstick ◽  
Wide Range ◽  
And Control

The northern root-knot nematode, Meloidogyne hapla, is a biotrophic parasite that infects many crops and causes severe economic losses worldwide. Rapid and accurate detection of M. hapla is crucial for disease forecasting and control. We developed a recombinase polymerase amplification combined with a lateral flow dipstick (RPA-LFD) assay for rapid detection of M. hapla. The primers and a probe were designed based on the effector gene 16D10 sequence and were highly specific to M. hapla. The RPA reaction was performed at a wide range of temperatures from 25 to 45°C within 5 to 25 min, and the amplicon was visualized directly on the LFD within 5 min. The detection limits of the RPA-LFD assay were 10-3 female and 10-2 J2/0.5 g of soil, which was 10 times more sensitive than the conventional PCR assay. In addition, the RPA-LFD assay can detect M. hapla from infested plant roots and soil samples, and the entire detection process can be completed within 1.5 h. These results indicate that the RPA-LFD assay is a simple, rapid, specific, sensitive, and visual method that can be used for rapid detection of M. hapla in the field and in resource-limited conditions.

scholarly journals The Development of Highly Specific and Sensitive Primers for the Detection of Potentially Allergenic Soybean (Glycine max) Using Loop-Mediated Isothermal Amplification Combined with Lateral Flow Dipstick (LAMP-LFD)

Foods ◽  
2020 ◽  
Vol 9 (4) ◽  
pp. 423 ◽  
Author(s):  
Stefanie M. Allgöwer ◽  
Chris A. Hartmann ◽  
Thomas Holzhauser
Keyword(s):  
Glycine Max ◽  
Detection Method ◽  
Lateral Flow ◽  
Isothermal Amplification ◽  
Loop Mediated Isothermal Amplification ◽  
Lateral Flow Dipstick ◽  
Soybean Allergy ◽  
Systematic Development ◽  
Multicopy Genes ◽  
Selection Of

The soybean (Glycine max) has been recognized as a frequent elicitor of food allergy worldwide. A lack of causative immunotherapy of soybean allergy makes soybean avoidance essential. Therefore, sensitive and specific methods for soybean detection are needed to allow for soybean verification in foods. Loop-mediated isothermal amplification (LAMP) represents a rapid and simple DNA-based detection method principally suitable for field-like applications or on-site analytical screening for allergens during the manufacturing of foods. This work describes the systematic development and selection of suitable LAMP primers based on soybean multicopy genes. The chemistry applied allows for a versatile detection of amplified DNA, using either gel electrophoresis, fluorescence recording, or a simple Lateral Flow Dipstick (LFD). LAMP based on the ORF160b gene was highly specific for the soybean and may allow for a detection level equivalent to approximately 10 mg soy per kg food. Various soybean cultivars were detectable at a comparable level of sensitivity. LAMP combined with LFD-like detection facilitates a simple, highly specific and sensitive detection of the soybean without the need for expensive analytical equipment. In contrast to the majority of antibody-based methods for soybean detection, all identified primer sequences and optimized protocols are disclosed and broadly available to the community.

scholarly journals Diagnosis of Histoplasmosis Using the MVista Histoplasma Galactomannan Antigen Qualitative Lateral Flow–Based Immunoassay: A Multicenter Study

2021 ◽  
Vol 8 (9) ◽  
Author(s):  
Wassim Abdallah ◽  
Thein Myint ◽  
Richard LaRue ◽  
Melissa Minderman ◽  
Suphansa Gunn ◽  
...  
Keyword(s):  
Severe Disease ◽  
The United States ◽  
Cross Reactivity ◽  
Lateral Flow ◽  
Medical Centers ◽  
Resource Limited ◽  
Pulmonary Histoplasmosis ◽  
Lateral Flow Assays ◽  
Poor Outcomes ◽  
Endemic Mycoses

Abstract Background Accurate and timely methods for the diagnosis of histoplasmosis in resource-limited countries are lacking. Histoplasma antigen detection by enzyme immunoassay (EIA) is widely used in the United States (US) but not in resource-limited countries, leading to missed or delayed diagnoses and poor outcomes. Lateral flow assays (LFAs) can be used in this setting. Methods Frozen urine specimens were submitted to MiraVista diagnostics for antigen testing from 3 medical centers in endemic areas of the US. They were blinded and tested for the MVista Histoplasma LFA. Patients were classified as controls or cases of histoplasmosis. Cases were divided into proven or probable; pulmonary or disseminated; immunocompetent or immunosuppressed; and mild, moderate, or severe. Results Three hundred fifty-two subjects were enrolled, including 66 cases (44 proven, 22 probable) and 286 controls. Most of the cases were immunocompromised (71%), and 46 had disseminated and 20 had pulmonary histoplasmosis. Four cases were mild, 42 moderate, and 20 severe. LFA and EIA were highly concordant (κ = 0.84). Sensitivity and specificity of the LFA were 78.8% and 99.3%, respectively. LFA sensitivity was higher in proven cases (93.2%), patients with disseminated (91.3%), moderate (78.6%), and severe disease (80%), and those with galactomannan levels >1.8 ng/mL (97.8%). Specificity was 99.3% in proven cases, 99.3% in patients with moderate or severe disease, and 96.8% in those with galactomannan levels >1.8 ng/mL. Cross-reactivity was noted with other endemic mycoses. Conclusions The MVista Histoplasma LFA meets the need for accurate rapid diagnosis of histoplasmosis in resource-limited countries, especially in patients with high disease burden, potentially reducing morbidity and mortality.

scholarly journals A recombinase polymerase amplification-lateral flow dipstick assay for rapid detection of the quarantine citrus pathogen in China, Phytophthora hibernalis

PeerJ ◽  
2019 ◽  
Vol 7 ◽  
pp. e8083 ◽  
Author(s):  
Tingting Dai ◽  
Tao Hu ◽  
Xiao Yang ◽  
Danyu Shen ◽  
Binbin Jiao ◽  
...  
Keyword(s):  
Reaction System ◽  
Citrus Fruit ◽  
Pcr Amplification ◽  
Brown Rot ◽  
Fungal Species ◽  
Lateral Flow ◽  
Recombinase Polymerase Amplification ◽  
Isolation And Identification ◽  
Resource Limited ◽  
Lateral Flow Dipstick

Phytophthora hibernalis, the causal agent of brown rot of citrus fruit, is an important worldwide pathogen and a quarantine pest in China. Current diagnosis of the disease relies on disease symptoms, pathogen isolation and identification by DNA sequencing. However, symptoms caused by P. hibernalis can be confused with those by other Phytophthora and fungal species. Moreover, pathogen isolation, PCR amplification and sequencing are time-consuming. In this study, a rapid assay including 20-min recombinase polymerase amplification targeting the Ypt1 gene and 5-min visualization using lateral flow dipsticks was developed for detecting P. hibernalis. This assay was able to detect 0.2 ng of P. hibernalis genomic DNA in a 50-µL reaction system. It was specific to P. hibernalis without detection of other tested species including P. citrophthora, P. nicotianae, P. palmivora and P. syringae, four other important citrus pathogens. Using this assay, P. hibernalis was also detected from artificially inoculated orange fruits. Results in this study indicated that this assay has the potential application to detect P. hibernalis at diagnostic laboratories and plant quarantine departments of customs, especially under time- and resource-limited conditions.

scholarly journals Evaluation of the diagnostic accuracy of lateral flow devices as a tool to diagnose rabies in post-mortem animals

2020 ◽  
Vol 14 (11) ◽  
pp. e0008844
Author(s):  
Kazunori Kimitsuki ◽  
Nobuo Saito ◽  
Kentaro Yamada ◽  
Chun-Ho Park ◽  
Satoshi Inoue ◽  
...  
Keyword(s):  
Sensitivity And Specificity ◽  
Fluorescent Antibody ◽  
Negative Control ◽  
Lateral Flow ◽  
Resource Limited Settings ◽  
Diagnostic Laboratory ◽  
Diagnostic Efficacy ◽  
Resource Limited ◽  
Efficient Detection ◽  
Flow Devices

Implementation of lateral flow devices (LFDs) for rabies antigen detection is expected to improve surveillance through the efficient detection of rabid animals in resource-limited settings; however, the use of LFDs for diagnosis remains controversial because some commercially available kits show low sensitivity. Therefore, we compared the diagnostic efficacy of three LFDs (ADTEC, Bionote, and Elabscience kits) paralleled with the direct fluorescent antibody test (dFAT) using fresh samples and investigated the diagnostic accuracies. To do so, we evaluated rabies-suspected samples submitted to the Regional Animal Disease Diagnostic Laboratory III, Philippines. Furthermore, we conducted real-time RT-PCR and sequencing to measure the accuracy of field laboratory diagnosis. The total number of animals submitted during this study period was 184 cases, including negative control samples. Of these, 53.9% (84 cases) were positive in the dFAT. Dogs were the most common rabies-suspected animal (n = 135). The sensitivities of the ADTEC and Bionote kits were 0.88 (74 cases) and 0.95 (80 cases), respectively. The specificity of both kits was 1.00 (100 cases). Furthermore, the sensitivity and specificity of the ADTEC kit after directly homogenizing the samples in assay buffer without dilution in phosphate-buffered saline (ADTEC kit DM) were 0.94 (79 cases) and 1.00 (100 cases), respectively. By contrast, there were no positive results using the Elabscience kit among all dFAT-positive samples. The sensitivity and specificity of LFDs make these tests highly feasible if properly used. Therefore, LFD tests can be used to strengthen the surveillance of rabies-infected animals in endemic and resource-limited settings.

scholarly journals Development of an accurate lateral flow immunoassay for PEDV detection in swine fecal samples with a filter pad design

2021 ◽  
Vol 1 (1) ◽  
Author(s):  
Siyi Zou ◽  
Lei Wu ◽  
Gan Li ◽  
Juan Wang ◽  
Dongni Cao ◽  
...  
Keyword(s):  
Limit Of Detection ◽  
Cross Reactivity ◽  
Rapid Diagnosis ◽  
Lateral Flow ◽  
Economic Losses ◽  
Lateral Flow Immunoassay ◽  
High Morbidity ◽  
Swine Industry ◽  
Fecal Samples ◽  
Diarrhea Virus

AbstractPorcine epidemic diarrhea virus (PEDV), as the main causative pathogen of viral diarrhea in pigs, has been reported to result in high morbidity and mortality in neonatal piglets and cause significant economic losses to the swine industry. Rapid diagnosis methods are essential for preventing outbreaks and transmission of this disease. In this study, a paper-based lateral flow immunoassay for the rapid diagnosis of PEDV in swine fecal samples was developed using stable color-rich latex beads as the label. Under optimal conditions, the newly developed latex bead-based lateral flow immunoassay (LBs-LFIA) attained a limit of detection (LOD) as low as 103.60 TCID50/mL and no cross-reactivity with other related swine viruses. To solve swine feces impurity interference, by adding a filtration unit design of LFIA without an additional pretreatment procedure, the LBs-LFIA gave good agreement (92.59%) with RT-PCR results in the analysis of clinical swine fecal samples (n = 108), which was more accurate than previously reported colloidal gold LFIA (74.07%) and fluorescent LFIA (86.67%). Moreover, LBs-LFIA showed sufficient accuracy (coefficient of variance [CV] < 15%) and stable (room temperature storage life > 56 days) performance for PEDV detection, which is promising for on-site analysis and user-driven testing in pig production system.

scholarly journals Recombinase Polymerase Amplification Based Multiplex Lateral Flow Dipstick for Fast Identification of Duck Ingredient in Adulterated Beef

Animals ◽  
2020 ◽  
Vol 10 (10) ◽  
pp. 1765
Author(s):  
Ming Fu ◽  
Quanwang Zhang ◽  
Xiang Zhou ◽  
Bang Liu
Keyword(s):  
Room Temperature ◽  
Detection Method ◽  
Lateral Flow ◽  
Recombinase Polymerase Amplification ◽  
Processed Meat ◽  
Visual Method ◽  
Lateral Flow Dipstick ◽  
Good Potential ◽  
Meat Species ◽  
Meat Samples

Meat adulteration has become a global social problem. In order to protect consumers from meat adulteration, several methods have been developed to identify meat species. However, the conventional methods are labor-intensive, time-consuming and require instruments. In the present study, a rapid and visual method based on recombinase polymerase amplification (RPA) and multiplex lateral flow dipstick (MLFD) was developed to detect duck ingredient in adulterated beef. Using recombinase and strand displacement polymerase enable RPA to amplify different double-labeled DNA amplicons at room temperature, which can be further detected by MLFD. The whole reaction process can be finished within 35 min, and the results can be determined by naked eyes. As low as 5% of duck ingredient in adulterated beef can be easily measured. Moreover, we confirmed that our new method held good potential in the detection of commercially processed meat samples. In conclusion, this study reported a useful animal derived meat adulteration detection method, which have potential application in future.

Development of a reverse-transcription recombinase polymerase amplification assay with a lateral flow assay for rapid detection of avian orthoavulavirus 1

2021 ◽  
Vol 33 (2) ◽  
pp. 308-312
Author(s):  
Jindai Fan ◽  
Wenxian Chen ◽  
Yuanyuan Zhang ◽  
Zhixiang Liu ◽  
Xiaoming Li ◽  
...  
Keyword(s):  
Reverse Transcription ◽  
Newcastle Disease ◽  
Cross Reactivity ◽  
Lateral Flow ◽  
Lateral Flow Assay ◽  
Recombinase Polymerase Amplification ◽  
Control Measures ◽  
Economic Losses ◽  
Resource Limited ◽  
Simple Detection

Newcastle disease is an avian infectious disease caused by avian orthoavulavirus 1, also known as Newcastle disease virus (NDV). This disease has caused significant economic losses to the poultry industry worldwide. The rapid and simple detection of NDV infection is crucial to inform the appropriate control measures. We developed a reverse-transcription recombinase polymerase amplification (RT-RPA) assay combined with a lateral flow assay (LFA) for NDV detection. The RPA assay can be completed at 37°C within 20 min, and the RPA result can be visualized by the LFA within 5 min. The NDV RT-RPA-LFA detected NDV specifically with no cross-reactivity with other pathogens. The detection limit of NDV cDNA with our RT-RPA-LFA was 3.34 × 10−3 ng/μL. Consequently, the RT-RPA-LFA showed good potential for the detection of NDV infection in the field, especially in resource-limited settings.

scholarly journals Ultrasensitive detection of Mycobacterium tuberculosis by a rapid and specific probe-triggered one-step, simultaneous DNA hybridization and isothermal amplification combined with a lateral flow dipstick

2020 ◽  
Vol 10 (1) ◽  
Author(s):  
Wansadaj Jaroenram ◽  
Jantana Kampeera ◽  
Narong Arunrut ◽  
Sarawut Sirithammajak ◽  
Sarinya Jaitrong ◽  
...  
Keyword(s):  
Mycobacterium Tuberculosis ◽  
Dna Hybridization ◽  
Cross Reactivity ◽  
Lateral Flow ◽  
Isothermal Amplification ◽  
Clinical Samples ◽  
High Resource ◽  
Lateral Flow Dipstick ◽  
Probe System ◽  
One Step

Abstract Mycobacterium tuberculosis (Mtb) is an insidious scourge that has afflicted millions of people worldwide. Although there are many rapid methods to detect it based on loop-mediated isothermal amplification (LAMP) and a lateral flow dipstick (LFD), this study made further improvements using a new set of primers to enhance LAMP performance and a novel DNA probe system to simplify detection and increase specificity. The new probe system eliminates the post-LAMP hybridization step typically required for LFD assays by allowing co-hybridization and amplification of target DNA in one reaction while preventing self-polymerization that could lead to false-positive results. The improved assay was named Probe-Triggered, One-Step, Simultaneous DNA Hybridization and LAMP Integrated with LFD (SH-LAMP-LFD). SH-LAMP-LFD was simpler to perform and more sensitive than previously reported LAMP-LFD and PCR methods by 100 and 1000 times, respectively. It could detect a single cell of Mtb. The absence of cross-reactivity with 23 non-TB bacteria, and accurate test results with all 104 blind clinical samples have highlighted its accuracy. Its robustness and portability make SH-LAMP-LFD suitable for users in both low and high resource settings.

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