Page 167 - 《水产学报》2025年第8期

何浩斌，等 水产学报, 2025, 49(8): 089614

Qualitative and quantitative detection of Oreochromis niloticus-derived
components in fish meal based on fluorescent quantitative PCR

HE Haobin 1,2,3,4 , YE Jiawen 1,2,3,4 , LIANG Guanyu 1,2,3,4 , HUANG Yanhua 1,2,3,4 ,
ZHOU Meng 1,2,3,4* , LIANG Rishen 1,2,3,4*
1. Innnovative Institute of Animal Healthy Breeding,
Zhongkai University of Agriculture and Engineering, Guangzhou 510225, China;
2. College of Animal Science and Technology,
Zhongkai University of Agriculture and Engineering, Guangzhou 510225, China;
3. Guangdong Provincial Key Laboratory of Waterfowl Healthy Breeding,
Zhongkai University of Agriculture and Engineering , Guangzhou 510225, China;
4. Guangzhou Key Laboratory of Aquatic Animal Diseases and Waterfowl Breeding,
Zhongkai University of Agriculture and Engineering, Guangzhou 510225, China

Abstract: Fish meal is one of the important raw materials in aquatic feed. It has complete amino acid composition, high cal-
cium, phosphorus and vitamin content, which is widely used in feed production. Recently, its large use and high price lead to
the serious adulteration in fish meal market, tilapia (Oreochromis niloticus) is one of the common adulterated raw materials. In
order to quickly and accurately detect O. niloticus-derived components in fish meal, this study, we established a real-time fluor-
escence PCR method (RT-PCR) on detecting O. niloticus-derived components in fish meal by designing the O. niloticus-spe-
cific primers. The specificity of the primers was verified based on the DNA amplification of various fish, shrimps and crabs.
Sensitivity of the method was detected based on the examination of O. niloticus DNA gradient dilution template and different
contents of O. niloticus component mixed with pure fish meal. Finally, the method was used to detect 8 different fish meal
samples to evaluate whether there were O. niloticus-derived components in the samples. The results showed that the method
had high specificity in identifying O. niloticus-derived components. There was significant amplification signal only found in O.
niloticus and no signal detected in other 50 common fish, shrimp and crab samples. In the sensitive experiment, when the
diluted concentration of O. niloticus DNA was 0.100 0 ng/μL, RT-PCR showed a typical specific amplification curve with a Ct
value 32.60±0.36, when the diluted concentration was 0.01 ng/μL, the Ct value was 35.01 ± 0.26, which was larger than the Ct
value of 35.00. In the experiment on detecting different content of O. niloticus component mixed with pure fish meal, when the
mixing content of O. niloticus component was 0.1%, the Ct value was 33.87±0.49, when the content was 0.010 0%, the Ct value
was 37.16±N/A (>35.00), indicating that the method could detect the minimum concentration of O. niloticus DNA was 0.100 0
ng/μL and the minimum content of O. niloticus component was 0.100 0%. In detecting 8 different fish meal samples from the
market basing on this method, 4 fishmeal samples: Pakistan fishmeal, Myanmar fishmeal, Vietnamese fishmeal and domestic
fishmeal 2 were detected to exist significant amplification signals and their Ct values were all lower than 35.00, revealing that
certain O. niloticus components were mixed in those 4 samples. The Ct value of Peruvian fishmeal, Japanese fishmeal, domestic
fishmeal 1 and imported super fishmeal were larger than 35.00 and they were judged as no O. niloticus components were detec-
ted. The result suggested that the method developed in this study can be used for rapid and effective detection of O. niloticus-
derived components in fish meal.
Key words: aquatic feed; Oreochromis niloticus-derived component; fish meal; real-time fluorescence quantitative PCR (RT-
PCR); detection
Corresponding authors: ZHOU Meng. E-mail: zhoumeng@zhku.edu.cn;
LIANG Rishen. E-mail: cheetahliang@126.com
Funding projects: Guangdong Provincial Key R & D Program (2021B0202050002); Project of Guangdong Province Guiding
Local Science and Technology Development (2023B0208010004); Science and Technology Program of Qingyuan (2022KJJH064)

中国水产学会主办 sponsored by China Society of Fisheries https://www.china-fishery.cn
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