RESEARCH PAPER
Rapid Visualized Detection of Escherichia Coli O157:H7 by DNA Hydrogel Based on Rolling Circle Amplification

https://doi.org/10.1016/S1872-2040(21)60085-3Get rights and content

Abstract

Food-borne illnesses caused by pathogenic bacteria are prominent issues in food safety. Rapid and sensitive detection of pathogenic bacteria is significantly essential for food safety. In this work, an aptamer biosensor (aptasensor) based on aldehyde magnetic beads (Mbs), rolling circle amplification (RCA) and DNA hydrogel was developed for visualized, simple and rapid detection of Escherichia coli O157:H7 (E. coli O157:H7). Firstly, the Mbs@double-stranded DNA (dsDNA) complexes were prepared. In the presence of target, the aptamer combined with E. coli O157:H7, releasing E. coli aptamer-initiator (EA-I). The supernatant collected by magnetic separation and those released primers were hybridized with circular sequence to form padlock probe. Then, RCA reaction was initiated by adding T4 DNA ligase, phi29 DNA polymerase, and deoxynucleotides (dNTPs), while RCA cannot be triggered in the absence of target. Two circular sequences were designed to contain partially complementary bases. After RCA, two long single-stranded DNA (ssDNA) products were generated using those two circular sequences. They hybridized with each other and formed the naked-eye visible DNA hydrogel. The method showed high sensitivity and specificity with a detection limit of 4 × 103 CFU/mL for E. coli O157:H7 within 1 h, and the detection time could be finished within 30 min. The aptasensor offers many advantages such as high specificity, easy operation, efficient amplification, rapid and visualized readout, which has potential applications in food safety detection

Graphical abstract

Herein, an aptasensor for the rapid visualization detection of E. coli O157: H7 was developed based on magnetic separation, specific identification of aptamers and characteristics of DNA hydrogel prepared by complementary hybridization of two RCA products. The proposed method was simple, rapid and had high sensitivity and specificity for the detection of E. coli O157: H7. In addition, it exhibited great potential to be a universal strategy for monitoring food safety.

  1. Download : Download high-res image (71KB)
  2. Download : Download full-size image

References (44)

  • M Xu et al.

    Talanta

    (2017)
  • Y Q Jiang et al.

    Sens. Actuators, B

    (2017)
  • S K Miryala et al.

    Genomics

    (2019)
  • T Saxena et al.

    Diagn. Microbiol. Infect. Dis.

    (2015)
  • O Lazcka et al.

    Biosens. Bioelectron.

    (2007)
  • C Wang et al.

    Food Chem.

    (2019)
  • Q Guo et al.

    Biosens. Bioelectron.

    (2016)
  • H L Zhang et al.

    J. Dairy Sci.

    (2018)
  • S Dolati et al.

    Anal. Biochem.

    (2018)
  • A Trapaidze et al.

    Biosens. Bioelectron.

    (2016)
  • L M Wang et al.

    Food Chem.

    (2019)
  • S Yan et al.

    Food Chem.

    (2019)
  • T Li et al.

    Anal. Chim. Acta

    (2020)
  • Y Wang

    Biomaterials

    (2018)
  • Q R Li et al.

    Anal. Biochem.

    (2019)
  • Y Zhang et al.

    Food Chem.

    (2019)
  • J Khang et al.

    Talanta

    (2016)
  • J Y Lim et al.

    J. Microbiol. Biotechnol.

    (2010)
  • D A Susana et al.

    Anal. Chim. Acta

    (2019)
  • B O Sergi et al.

    Sens. Actuators, B

    (2018)
  • E D Hilborn et al.

    Arch. Intern. Med.

    (1999)
  • D D Zhang et al.

    Sci. Rep.

    (2016)
  • Cited by (11)

    • An electrochemical aptasensor based on dual-enzymes-driven target recycling strategy for patulin detection in apple juice

      2022, Food Control
      Citation Excerpt :

      Typically, one or two target molecules only triggers a single biorecognizing event, which limits the sensitivity of the aptasensor. Therefore, a variety of signal amplification strategies have been introduced to aptasensor to improve the utilization rate of the target, including rolling circle amplification (RCA), polymerase chain reaction (PCR), strand displacement amplification (SDA)(Fan et al., 2018; Xia Li et al., 2018; Ma et al., 2019; Zhang, Tao, Bian, Chen, & Yan, 2021). Besides, enzymes-assisted target recycling (EATR) is considered a promising signal amplification mechanism.

    • Recent advances in microchip-based methods for the detection of pathogenic bacteria

      2022, Chinese Chemical Letters
      Citation Excerpt :

      Molecular diagnostics play an important role in medical diagnosis, biological studies, food safety, and environmental monitoring. However, the effective detection of low amounts of target nucleic acids in complex biological samples required signal amplification techniques; loop-mediated isothermal amplification (LAMP) [61,62], CHA [39,63], RCA [64,65], and strand displacement amplification (SDA) [66,67], have been developed in this regard. SDA is an isothermal amplification method that generally depends on an ssDNA template containing complementary bases to the endonuclease recognition sequence; however, the SDA method falls short in terms of detecting short nucleic acids.

    View all citing articles on Scopus

    This work was supported by the National Natural Science Foundation of China (No. 21775102) and the Natural Science Foundation of Shanghai Municipal, China (No. 20ZR1424100).

    View full text