Hybrid RCA-DLS assay combined with aPCR for sensitive Salmonella enteritidis detection

Guoyang Xie; Zhongxu Zhan; Yu Ye; Baoqing Zhou; Ping Tong; Zoraida P Aguilar; Hengyi Xu

doi:10.1016/j.ab.2022.114647

Hybrid RCA-DLS assay combined with aPCR for sensitive Salmonella enteritidis detection

Anal Biochem. 2022 Jun 1:646:114647. doi: 10.1016/j.ab.2022.114647. Epub 2022 Mar 12.

Authors

Guoyang Xie¹, Zhongxu Zhan², Yu Ye¹, Baoqing Zhou³, Ping Tong¹, Zoraida P Aguilar⁴, Hengyi Xu⁵

Affiliations

¹ State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang, 330047, PR China.
² State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang, 330047, PR China; Jiangxi Institute for Food Control, Nanchang, 330001, PR China.
³ State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang, 330047, PR China; Guangdong Institute of Microbiology, Guangdong, 510070, PR China.
⁴ Zystein, LLC, Fayetteville, AR, 72703, USA.
⁵ State Key Laboratory of Food Science and Technology, Nanchang University, Nanchang, 330047, PR China. Electronic address: kidyxu@163.com.

PMID: 35289287
DOI: 10.1016/j.ab.2022.114647

Abstract

Salmonella infection could come from eating contaminated meat or raw eggs, and drinking milk or water contaminated by Salmonella enteritidis. Therefore, it is necessary to explore a fast and easy method for the detection of S. enteritidis in these diverse samples. For this purpose, a novel particle size sensing tool was designed for ultrasensitive and accurate S. enteritidis detection. This assay consisted of rolling circle amplification (RCA) with dynamic light scattering (DLS) using gold nanoparticles (AuNPs) modified with DNA probe as DNA-AuNPs as the capture surface into a hybrid RCA-DLS assay combined with asymmetric polymerase chain reaction (aPCR) and subsequent detection. Under optimal experimental conditions, the novel hybrid RCA-DLS assay combined with aPCR for S. enteritidis reached a limit of detection (LOD) as low as 3 × 10⁰ CFU/mL in pure culture. In spiked milk samples, the LOD was 2.0 × 10⁰ CFU/mL without pre-enriched bacteria. The total time of RCA-DLS assay was about 6 h which including genomic DNA extraction, aPCR, RCA and DLS determination. The hybrid RCA-DLS assay combined with aPCR holds promise in the specific and sensitive S. enteritidis detection.

Keywords: Asymmetric polymerase chain reaction; DNA-AuNPs; Dynamic light scattering; Rolling circle amplification; Salmonella enteritidis.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

DNA, Bacterial / analysis
DNA, Bacterial / genetics
Dynamic Light Scattering
Gold
Limit of Detection
Metal Nanoparticles*
Polymerase Chain Reaction / methods
Salmonella enteritidis* / genetics

Substances

DNA, Bacterial
Gold