Recombinase Polymerase Amplification (RPA) Combined with Lateral Flow Immunoassay for Rapid Detection of Salmonella in Food

Li, Jiali; Ma, Bach; Fang, Jian Jun; Zhi, Antong; Chen, Erjing; Xu, Ying; Yu, Xiaoping; Sun, Chuanxin; Zhang, Mingzhou

doi:10.3390/foods9010027

Cited by 58 publications

(47 citation statements)

References 30 publications

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“…causes food poisoning, typhoid fever, gastrointestinal inflammation, and septicemia for both humans and animals ( McGuinness et al, 2009 ; Rukambile et al, 2019 ). Currently, there are over 80 million cases of foodborne salmonellosis in the world ( Li et al, 2019 ). Additionally, reports revealed that outbreaks caused by Salmonella spp.…”

Section: Introductionmentioning

confidence: 99%

Development and Evaluation of the Rapid and Sensitive RPA Assays for Specific Detection of Salmonella spp. in Food Samples

Zhao

Wang²,

Sun³

et al. 2021

Front. Cell. Infect. Microbiol.

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Salmonella spp. is among the main foodborne pathogens which cause serious foodborne diseases. An isothermal real-time recombinase polymerase amplification (RPA) and lateral flow strip detection (LFS RPA) were used to detect Salmonella spp. targeting the conserved sequence of invasion protein A (invA). The Real-time RPA was performed in a portable florescence scanner at 39°C for 20 min. The LFS RPA was performed in an incubator block at 39°C for 15 min, under the same condition that the amplifications could be inspected by the naked eyes on the LFS within 5 min. The detection limit of Salmonella spp. DNA using real-time RPA was 1.1 × 101 fg, which was the same with real-time PCR but 10 times higher than that of LFS RPA assay. Moreover, the practicality of discovering Salmonella spp. was validated with artificially contaminated lamb, chicken, and broccoli samples. The analyzing time dropped from 60 min to proximately 5–12 min on the basis of the real-time and LFS RPA assays compared with the real-time PCR assay. Real-time and LFS RPA assays’ results were equally reliable. There was no cross-reactivity with other pathogens in both assays. In addition, the assays had good stability. All of these helped to show that the developed RPA assays were simple, rapid, sensitive, credible, and could be a potential point-of-need (PON) test required mere resources.

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Section: Introductionmentioning

confidence: 99%

Development and Evaluation of the Rapid and Sensitive RPA Assays for Specific Detection of Salmonella spp. in Food Samples

Zhao

Wang²,

Sun³

et al. 2021

Front. Cell. Infect. Microbiol.

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“…The exponential amplification can be detected within 30 min in a temperature range of 37 to 42 °C [ 19 ]. Recently, several applications of RPA in DNA and RNA target detection have been proved [ 20 , 21 ], and the integration of RPA in different platforms has been reported [ 22 , 23 , 24 ]. Lateral flow dipsticks (LFD) has great application prospects in the detection field due to easy-to-use, high sensitivity, less time consuming and low-cost [ 25 , 26 , 27 ].…”

Section: Introductionmentioning

confidence: 99%

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

Zhang

Zhou

et al. 2020

Animals

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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.

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“…But, designing of RPA probe and primers are still difficult, as several sets of primers must be checked for the selection of a working set, as there is no suitable available software so far. RPA has several important advantages over PCR methods, the first being that it does not require a pure template for the analysis, which is required for PCR-based assays ( Mekuria et al., 2014 ; Li et al., 2019 ). The second advantage is that results are available in a much shorter time i.e., 15–20 min for RPA and 1.5-3.0 h for PCR assays.…”

Section: Discussionmentioning

confidence: 99%

Development and Evaluation of Real-Time Reverse Transcription Recombinase Polymerase Amplification Assay for Rapid and Sensitive Detection of West Nile Virus in Human Clinical Samples

Tomar

Kumar

Patel

et al. 2021

Front. Cell. Infect. Microbiol.

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West Nile virus (WNV) causes West Nile fever and encephalitis worldwide. Currently, there are no effective drugs or vaccines available in the market to treat WNV infection in humans. Hence, it is of paramount importance to detect WNV early for the success of the disease control programs and timely clinical management in endemic areas. In the present paper, we report the development of real-time reverse transcription recombinase polymerase amplification (RT-RPA) assay for rapid and real-time detection of WNV targeting the envelope (env) gene of the virus. The RPA reaction was performed successfully at 39°C for 15 min in a real-time thermal cycler. The sensitivity of this assay was found similar to that of the quantitative real-time RT PCR (RT-qPCR) assay, which could detect 10 copies of the gene. The efficacy of the assay was evaluated with a panel of 110 WN suspected human samples showing the signs of retinitis, febrile illness and acute posterior uveitis. In comparison with RT-qPCR, RT-RPA showed a specificity of 100% (CI, 95.07–100%) and sensitivity of 96.15% (CI, 80.36–99.90%) with a negative (NPV) and positive predictive value (PPV) of 98.65 and 100%, respectively. The level of agreement between RT-RPA and reference RT-qPCR assay was shown to be very high. The turnaround time of real-time RPA assay is about 10-20 times faster than the RT-qPCR, which confirms its utility in the rapid and sensitive diagnosis of WNV infection. To the best of our knowledge, this is the first report which deals with the development of real-time RT-RPA assay for simple, rapid, sensitive, and specific detection of WNV in human clinical samples. The present RT-RPA assay proves to be a powerful tool that can be used for the rapid diagnosis of a large number of patient samples in endemic settings.

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Recombinase Polymerase Amplification (RPA) Combined with Lateral Flow Immunoassay for Rapid Detection of Salmonella in Food

Cited by 58 publications

References 30 publications

Development and Evaluation of the Rapid and Sensitive RPA Assays for Specific Detection of Salmonella spp. in Food Samples

Development and Evaluation of the Rapid and Sensitive RPA Assays for Specific Detection of Salmonella spp. in Food Samples

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

Development and Evaluation of Real-Time Reverse Transcription Recombinase Polymerase Amplification Assay for Rapid and Sensitive Detection of West Nile Virus in Human Clinical Samples

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