Development of the Saltatory Rolling Circle Amplification Assay for Rapid and Visual Detection of <i>Alicyclobacillus acidoterrestris</i> in Apple Juice
Ning Yuan, Yunzhe Zhang, Hui Xu, Zhijun Zhou, Xin Lü, Tingting Chen, Qian Yang, Jianxin Tan, Wei Zhang
Abstract
A novel nucleic acid isothermal amplification method based on saltatory rolling circle amplification (SRCA) for rapid and visual detection of Alicyclobacillus acidoterrestris in apple juice was established. Fourteen A. acidoterrestris strains and 44 non-A. acidoterrestris strains were used to confirm the specificity. The sensitivity of SRCA was 4.5 × 101 CFU/mL by observing the white precipitate with the naked eye, while it was 4.5 × 100 CFU/mL by fluorescence visualization. The detection limit of SRCA in artificially inoculated apple juice was 7.1 × 101 and 7.1 × 100 CFU/mL via visualization of the white precipitate and fluorescence, respectively. Compared with the traditional PCR method, SRCA exhibited at least a 100-fold higher sensitivity and 100-fold lower detection limit. Seventy samples were investigated for A. acidoterrestris contamination, and the results showed 100% sensitivity, 97.01% specificity, and 97.14% accuracy compared with those by the conventional microbiological cultivation method. Overall, this method is a potentially useful tool for visual and rapid detection of A. acidoterrestris.