Browse
Submit Data
Databases
API
Help

Dataset Information

0 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Visual detection of canine parvovirus based on loop-mediated isothermal amplification combined with enzyme-linked immunosorbent assay and with lateral flow dipstick.

ABSTRACT: Loop-mediated isothermal amplification (LAMP) combined with enzyme-linked immunosorbent assay (LAMP-ELISA) and with lateral flow dipstick (LAMP-LFD) are rapid, sensitive and specific methods for the visual detection of clinical pathogens. In this study, LAMP-ELISA and LAMP-LFD were developed for the visual detection of canine parvovirus (CPV). For LAMP, a set of four primers (biotin-labeled forward inner primers) was designed to specifically amplify a region of the VP2 gene of CPV. The optimum time and temperature for LAMP were 60 min and 65°C, respectively. The specific capture oligonucleotide probes, biotin-labeled CPV probe for LAMP-ELISA and fluorescein isothiocyanate-labeled CPV probe for LAMP-LFD were also designed for hybridization with LAMP amplicons on streptavidin-coated wells and LFD strips, respectively. For the comparison of detection sensitivity, conventional PCR and LAMP for CPV detection were also performed. The CPV detection limits by PCR, PCR-ELISA, LAMP, LAMP-ELISA and LAMP-LFD were 10(2), 10(2), 10(-1), 10(-1) and 10(-1) TCID50/ml, respectively. In tests using artificially contaminated dog fecal samples, the samples with CPV inoculation levels of ?1 TCID50/ml gave positive results by both LAMP-ELISA and LAMP-LFD. Our data indicated that both LAMP-ELISA and LAMP-LFD are promising as rapid, sensitive and specific methods for an efficient diagnosis of CPV infection.

SUBMITTER: Sun YL

PROVIDER: S-EPMC4064134 | biostudies-literature | 2014 Apr

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Loop-mediated isothermal amplification-lateral-flow dipstick (LAMP-LFD) to detect Mycoplasma ovipneumoniae.

Project description:In order to establish a rapid detection method for Mycoplasma ovipneumoniae, this study used the loop-mediated isothermal amplification (LAMP) technique to carry out nucleic acid amplification and chromatographic visualization via a lateral flow dipstick (LFD) assay. The M. ovipneumoniae elongation factor TU gene (EF-TU) was detected using a set of specific primers designed for the EF-TU gene, and the EF-TU FIP was detected by biotin labeling, which was used in the LAMP amplification reaction. The digoxin-labeled probe specifically hybridized with LAMP products, which were visually detected by LFD. Here, we established the M. ovipneumoniae LAMP-LFD rapid detection method and tested the specificity, sensitivity, and clinical application of this method. Results showed that the optimized LAMP performed at 60 °C for 60 min, and LFD can specifically and visually detect M. ovipneumoniae with a minimum detectable concentration at 1.0 × 102 CFU/mL. The sensitivity of LAMP-LFD was 1000 times that of the conventional PCR detection methods, and the clinical lung tissue detection rate was 86% of 50 suspected sheep infected with M. ovipneumoniae. In conclusion, LAMP-LFD was established in this study to detect M. ovipneumoniae, a method that was highly specific, sensitive, and easy to operate, and provides a new method for the prevention and diagnosis of M. ovipneumoniae infection.

| S-EPMC6353813 | biostudies-literature

Using loop-mediated isothermal amplification for visual detection of porcine parvovirus 7.

Project description:We developed and optimized a loop-mediated isothermal amplification (LAMP)-based method to detect porcine parvovirus 7 (PPV7). After using three pairs of specific primers to amplify PPV7 isothermally at 62 °C for 40 min, the amplified product was mixed with SYBR Green I, after which the sample turned green. The method detected PPV7 at concentrations as low as 40 copies/μL, and the sensitivity was consistent with that of nested polymerase chain reaction (PCR) analysis, which was tenfold higher than that of conventional PCR. No cross-reactivity occurred with porcine parvovirus 1, porcine circovirus type 3, porcine circovirus type 2, porcine pseudorabies virus, porcine epidemic diarrhea virus, or porcine reproductive and respiratory syndrome virus. Simultaneous analysis of 76 clinical samples was performed using LAMP, conventional PCR, and nested PCR. The results showed that our method is simple, rapid, sensitive, and specific for the rapid diagnosis of PPV7 in pig farms.Supplementary informationThe online version of this article (10.1007/s13205-020-02623-5) contains supplementary material, which is available to authorized users.

| S-EPMC7870734 | biostudies-literature

Development of a rapid loop-mediated isothermal amplification (LAMP) assay for visual detection of porcine parvovirus (PPV) and its application.

Project description:Porcine parvovirus (PPV) infection is one of the most important causes of reproductive failure in pigs impacting the piggery industry globally with huge economic losses. A cost-effective, simple, rapid, specific, and sensitive method is critical for monitoring PPV infection on pig farms. The main aim of the present study was to develop and evaluate a loop-mediated isothermal amplification (LAMP) assay for rapid visual detection of porcine parvovirus (PPV) in pigs. A set of six LAMP primers including two outer primers, two inner primers, and two loop primers were designed utilizing the conserved region of capsid protein VP2 gene sequences of PPV and was applied for detection of PPV from porcine samples. Time and temperature conditions for amplification of PPV genes were optimized to be 30 min at 63 °C. The developed assay was ten-fold more sensitive than conventional PCR with analytical sensitivity of 20 pg and 200 pg, respectively. This is the first report of detection of PPV by LAMP assay from India. The assay did not cross-react with porcine circovirus type 2 (PCV2), porcine reproductive and respiratory syndrome virus (PRRSV), or classical swine fever virus (CSFV). The LAMP assay was assembled into a LAMP assay kit of 20 reactions and was validated in different laboratories in India. The newly developed LAMP assay was proved to be a specific, sensitive, rapid, and simple method for visual detection of PPV which does not require even costly equipments for performing the test. It complements and extends previous methods for PPV detection and provides an alternative approach for detection of PPV.

| S-EPMC8578513 | biostudies-literature

Rapid and visual detection of an isolated and identified goose parvovirus (GPV) strain by a loop-mediated isothermal amplification assay.

Project description:Gosling plague caused by goose parvovirus (GPV), a highly infectious septic disease with high mortality, has caused substantial loss in the waterfowl industry. A method for the rapid detection of GPV is needed. In this study, we isolated the virus strain of GPV in May 2020 and applied it to the loop-mediated isothermal amplification (LAMP) assay. We designed five sets of primers for the goose parvovirus VP3 gene by LAMP. The GV-1 primer set was selected to detect GPV sensitively and rapidly. LAMP was more sensitive compared to PCR. In addition, the LAMP method could complete detection within 60 min which was faster than the PCR assay. The LAMP provided a convenient and effective experimental method for detection of GPV for inspection and quarantine departments and health care units in China, and it is expected to become a simple and routine detection method, especially suitable for goose farms.

| S-EPMC9906619 | biostudies-literature

Rapid Visual Detection of Plasmodium Using Recombinase-Aided Amplification With Lateral Flow Dipstick Assay.

Project description:BackgroundMalaria is a global public health problem. China has had no case of indigenous malaria since 2016. However, imported cases of malaria remain an issue among travelers, overseas workers, and foreign traders. Although these cases are always asymptomatic, if they donate blood, there is a great risk of transfusion transmitted-malaria (TTM). Therefore, blood banks need a rapid screening tool to detect Plasmodium species.MethodsWe designed an assay using recombinase-aided amplification (RAA) and a lateral-flow dipstick (LFD) (RAA-LFD) to detect the 18S ribosomal RNA gene of Plasmodium species. Sensitivity was evaluated using a recombinant plasmid and Plasmodium genomic DNA. Specificity was evaluated using DNA extracted from the blood of patients with malaria or other infectious parasites. For clinical assessment, blood samples from patients with malaria and blood donors were evaluated.ResultsThe RAA-LFD assay was performed in an incubator block at 37°C for 15 min, and the amplicons were visible to the naked eye on the flow dipsticks within 3 min. The sensitivity was 1 copy/μL of recombinant plasmid. For genomic DNA from whole blood of malaria patients infected with P. falciparum, P. vivax, P. ovale, and P. malariae, the sensitivity was 0.1 pg/μL, 10 pg/μL, 10-100 pg/μL, and 100pg/μL, respectively. The sensitivity of this assay was 100pg/μL. No cross-reaction with other transfusion-transmissible parasites was detected.ConclusionsThe results demonstrated that this RAA-LFD assay was suitable for reliable field detection of Plasmodium species in low-resource settings with limited laboratory capabilities.

| S-EPMC9263184 | biostudies-literature

Development of a recombinase polymerase amplification assay with lateral flow dipstick for rapid detection of feline parvovirus.

Project description:Feline panleukopenia caused by feline parvovirus (FPV), a single-stranded DNA virus, is typically highly contagious and often presents with lethal syndrome. The broad spectrum of possible hosts suggests its potential for transmission from animal to person through close contact with pets. FPV thus serves as an example of the importance of new rapid point-of-care field diagnostic tools for the control and prevention of transmission, especially among rare wild animals and pet cats. Recombinase polymerase amplification (RPA), as a real-time and isothermal method, could be a more affordable alternative to PCR when combined with a lateral flow dipstick (LFD) indicator. In this study, we report a novel FPV lateral flow dipstick RPA (LFD-RPA) instant detection method capable of detecting a range of different FPV strains. The LFD-RPA assay consists of specific primers, probe, and nucleic acid strip. It is capable of detecting 102 copies of target nucleic acid per reaction, which is one order of magnitude higher than the sensitivity of traditional PCR. The most suitable reaction conditions for this assay are at 38 ? for 15?min. This paper develops an efficient visual detection system that can eliminate the need for professional staff and expensive and sophisticated equipment for field detection.

| S-EPMC7113848 | biostudies-literature

Development and evaluation of loop-mediated isothermal amplification assay for rapid and sensitive detection of canine parvovirus DNA directly in faecal specimens.

Project description:AIMS:?To develop a specific and highly sensitive loop-mediated isothermal amplification (LAMP) technique for the rapid detection of canine parvovirus (CPV) DNA directly in suspected faecal samples of dogs by employing a simple method of template preparation. METHODS AND RESULTS:?LAMP reaction was developed by designing two sets of outer and inner primers, which target a total of six distinct regions on VP2 gene of CPV. The template DNA was prepared by a simple boiling and chilling method. Of the 140 faecal samples screened by the developed LAMP and the conventional PCR assays, 104 samples (74·28%) were found positive by LAMP, whereas 81 samples (57·85%) were found positive by PCR. The specificity of the LAMP assay was tested by cross-examination of common pathogens of dogs and further confirmed by sequencing. The detection limit of the LAMP was 0·0001 TCID?? ml?¹, whereas the detection limit of the PCR was 1000 TCID?? ml?¹. CONCLUSIONS:?The developed LAMP assay detects CPV DNA in faecal specimens directly within an hour by following a simple and rapid boiling and chilling method of template preparation. The result also shows that the developed LAMP assay is specific and highly sensitive in detecting CPV. SIGNIFICANCE AND IMPACT OF THE STUDY:?The result indicates the potential usefulness of LAMP which is a simple, rapid, specific, highly sensitive and cost-effective field-based method for direct detection of CPV from the suspected faecal samples of dogs.

| S-EPMC7197762 | biostudies-literature

Loop-Mediated Isothermal Amplification Combined with Lateral Flow Dipstick for On-Site Diagnosis of African Swine Fever Virus.

Project description: Not available

| S-EPMC8087745 | biostudies-literature

A loop-mediated isothermal amplification assay for the visual detection of duck circovirus.

Project description:BackgroundDuck circovirus (DuCV) infection in farmed ducks is associated with growth problems or retardation syndromes. Rapid identification of DuCV infected ducks is essential to control DuCV effectively. Therefore, this study aims to develop of an assay for DuCV to be highly specific, sensitive, and simple without any specialized equipment.MethodsA set of six specific primers was designed to target the sequences of the Rep gene of DuCV, and A loop-mediated isothermal amplification (LAMP) assay were developed and the reaction conditions were optimized for rapid detection of DuCV.ResultsThe LAMP assay reaction was conducted in a 62°C water bath condition for 50 min. Then the amplification products were visualized directly for color changes. This LAMP assay is highly sensitive and able to detect twenty copies of DuCV DNA. The specificity of this LAMP assay was supported by no cross-reaction with other duck pathogens.ConclusionThis LAMP method for DuCV is highly specific and sensitive and can be used as a rapid and direct diagnostic assay for testing clinical samples.

| S-EPMC4013541 | biostudies-literature

Development of Loop-Mediated Isothermal Amplification Combined with Lateral Flow Dipstick Assay for a Rapid and Sensitive Detection of Cystic Echinococcosis in Livestock in Kenya.

Project description:BackgroundCystic echinococcosis is a zoonotic disease caused by the metacestode stage of Echinococcus granulosus and occurs worldwide, causing considerable economic losses and public health problems. The currently available methods for the diagnosis of animal hydatidosis are time-consuming and require well-equipped laboratories which make them incompatible with testing in resource-poor settings. This study developed and evaluated a rapid, more sensitive, and specific loop-mediated isothermal amplification combined with a lateral flow dipstick assay for the rapid and sensitive detection of cystic echinococcosis.ResultsIn this study, a specific primer set and FITC-labeled probe targeting the conserved region of the NADH-1 gene were designed. The LAMP reaction was performed at 60°C for 40 minutes, and the amplification products were successfully visualized by LFD strips. The analytical sensitivity of LAMP-LFD was determined using 10-fold serial dilutions of E. granulosus DNA. The minimal concentration detected was 10 fg/μl, and no cross-reactivity was observed with DNA extracted from Taenia solium, Taenia saginata, and Fasciola hepatica. The ability of the developed LAMP-LFD assay to detect cystic echinococcosis was further evaluated with 62 cyst samples from slaughtered cattle in Juja Abattoir, Kiambu County, Kenya. The LAMP-LFD was able to detect 59/62 (95.2%, 95% CI 0.87-0.98) as positive samples of E. granulosus compared to 53/62 (85.5%, 95% CI 0.75-0.92) by nested PCR assay.ConclusionOur results indicated that the developed LAMP-LFD technique was more sensitive than the nested PCR assay, rapid, and easy to perform with a simple visual detection of products. Therefore, it could be an important point-of-care diagnostic tool for cystic echinococcosis.

| S-EPMC8898851 | biostudies-literature

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data