Citation: WANG Ping, TIAN Zhuo, QI Xin, ZHENG Qiu-Yue, HU Bing, MA Li-Dan, YANG Li-Li, CHEN Ying, CAO Ji-Juan. Highly Sensitive Detection of Low Concentration Hepatitis A Virus in Small Berries Using Polyethylene Glycol Precipitation Enrichment and Pre-Amplification Reverse Transcription- Quantitative Polymerase Chain Reaction[J]. Chinese Journal of Analytical Chemistry, ;2022, 50(8): 1168-1178. doi: 10.19756/j.issn.0253-3820.210775 shu

Highly Sensitive Detection of Low Concentration Hepatitis A Virus in Small Berries Using Polyethylene Glycol Precipitation Enrichment and Pre-Amplification Reverse Transcription- Quantitative Polymerase Chain Reaction

WANG Ping ¹ ,
TIAN Zhuo ^2,3,5 ,
QI Xin ³ ,
ZHENG Qiu-Yue ² ,
HU Bing ² ,
MA Li-Dan ⁴ ,
YANG Li-Li ² ,
CHEN Ying ^1,, ,
CAO Ji-Juan ^2,,

1.
Chinese Academy of Inspection and Quarantine, Beijing 322001, China;
2.
Key Laboratory of Biotechnology and Bioresources Utilization of Ministry of Education, College of Life Science, Dalian Minzu University, Dalian 116600, China;
3.
Dalian Customs Technology Center, Dalian 116001, China;
4.
Dandong Customs Technology Center, Dandong 118017, China;
5.
School of Life Science, Liaoning Normal University, Dalian 116023, China

Corresponding author: CHEN Ying, CAO Ji-Juan,
Received Date: 2 October 2021
Revised Date: 25 March 2022

Fund Project: Supported by the National Key R&D Program of China (No.2018YFC1603504) and the "Rejuvenating Liaoning Talents Plan" of Liaoning Province, China (No.XLYC2002106).

On the basis of polyethylene glycol (PEG) precipitation enrichment and improved pre-amplified reverse transcription-quantitative polymerase chain reaction (RT-qPCR), a method was established for highly sensitive detection of low-dose contamination of hepatitis A virus (HAV) in small berries. The treatment time of HAV in strawberry enriched by PEG precipitation was optimized, and the pre-amplification steps were set up to accelerate the lysis of sample nucleic acid and enrichment of templates. By using this method, the recovery was 15.51%, the lowest detection sensitivity could be up to 4.49 CCID₅₀/mL of HAV attenuated vaccine titer, and the sensitivity of HAV plasmid was as low as 3.5 copies/μL. Compared with ISO/TS15216-2:2019 standard method, this method had higher detection efficiency and was suitable for detection of trace contaminated HAV in small berries. The positive rate of HAV in 78 batches of commercially available small berry samples was 5.13%, which was higher than that of ISO/TS 15216-2:2019 standard (1.28%). This method showed good applicability and application prospect for detection of low-dose HAV virus contamination in small berries.
- Small berries,
- Low-dose hepatitis A virus,
- Polyethylene glycol,
- Pre-amplification reverse transcription-quantitative polymerase chain reaction,
- Detection
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