Colorimetric Integrated PCR Protocol for Rapid Detection of Vibrio parahaemolyticus

Abstract

Rapid detection of pathogens is of great significance for food safety and disease diagnosis. A new colorimetric method for rapid and easy detection of Vibrio parahaemolyticus (V. parahaemolyticus or Vp) has been developed in this research. A specific sequence was designed and integrated with the forward primer for molecular detection of Vp. This specific sequence was tested and treated as the horseradish peroxidase (HRP)-mimicking DNAzyme and could be amplified during the polymerase chain reaction (PCR) process. The products of PCR including the sequence of HRP-mimicking DNAzyme could produce the distinguished color in the presence of catalysis substrates. The optical signal of the catalysis reaction, which is in a linear relationship with the initial template of Vp, could be determined with the naked eye or measured with Ultraviolet-visible (UV-vis) for qualitative and quantitative detections, respectively. Based on the optical signal intensity, rapid and easy detection of Vp was successfully achieved with satisfied sensitivity and specificity. Furthermore, the detection of tdh, trh, tlh and toxR virulence genes of two Vp species (Vp 33847 and Vp 17802) were all performed successfully with this developed colorimetric integrated PCR protocol, which demonstrated potential applicability for the rapid detection of other bacteria.

Self‐signal‐on fluorescent colorimetric protocol for rapid authentication of horsemeat adulterated beef samples with functional designed probes

“…For the reported PCR‐ELISA or PCR‐RFLP‐based protocol (Asensio et al ., ; Kumar et al ., ; Santaclara et al ., ), additional 2 h or overnight is required to achieve the final results of the amplifications respectively. Even compared to our previous results of mimicking DNAzyme‐included primer‐based PCR protocol (Cheng et al ., ), the hairpin‐based fluorescent signal‐on protocol in this research is about 25–30 min faster. Comparatively, the proposed hairpin primer‐based fluorescent signal‐on method in this research is competitive to other methods in detection efficiency and the “rapid” and “easy‐operation” properties are completely highlighted.…”

A simple, portable, electrochemical biosensor to screen shellfish for Vibrio parahaemolyticus

“…The Vibrio parahaemolyticus pathogen is an important seafood-borne enteropathogen that continues to cause acute human gastroenteritis throughout the world (Zhang et al 2016 ) and its rapid detection is of great significance for food safety and disease diagnosis (Cheng et al 2016 ).…”

Rapid and selective recognition of Vibrio parahaemolyticus assisted by perfluorinated alkoxysilane modified molecularly imprinted polymer film

“…The entire detection process only took an average of 5 h, including the capture of bacteria (2 h), lysis of bacteria and DNA extraction (10 min), PCR (2 h), agarose gel electrophoresis (50 min). The developed method was compared with traditional PCR-based methods aiming at detection of bacteria, [36][37][38][39] as shown in Table 4. The method has considerable advantages over most of traditional PCR-based methods in terms of analytical time with a similar detection limit.…”