基于核酸适配体的PCR法检测溶藻弧菌及其灭活菌

溶藻弧菌(Vibrio alginolyticus)分布广,数量多,发病率高,是水产养殖中常见的条件致病菌,而对溶藻弧菌进行快速准确的识别鉴定是其病害防治的前提和基础。核酸适配体,因为具有较高的亲和特异性,在微生物的识别鉴定方面展现出了巨大的优势。本文利用核酸适配体和适配体筛选产物,通过结合、洗涤、加热分离、PCR扩增以及电泳检测等步骤,对溶藻弧菌进行了检测鉴定。结果表明,适配体和筛选产物都能对溶藻弧菌及其灭活菌进行较好的识别鉴定,适配体筛选产物对溶藻弧菌的检测下限为10~3cfu/mL,而对其灭活菌的检测下限为10~2cfu/mL,适配体对溶藻弧菌及其灭活菌的检测下限都可达到10 cfu/mL。该方法对溶藻弧菌有较好的亲和特异性,并能较好地区分溶藻弧菌与哈维氏弧菌等水产常见病原菌,在水产病害的检测中显示了较好的应用前景。

Detection of Vibrio alginolyticus and its inactivated form based on aptamers and polymerase chain reaction

Vibrio alginolyticus is a common conditional pathogen in aquaculture, which is widely distributed in the world. Rapid detection of the pathogen is necessary for the prevention and control of the diseases caused by the bacterium. Aptamers exhibit great advantages in the detection of microorganisms owing to their high affinities and specificities. In the present study, Vibrio alginolyticus was detected based on its aptamer and its selection products following various steps, such as binding, washing, separation, amplification, and electrophoresis. The results showed that the aptamer and the selection products could superiorly detect V. alginolyticus and its inactivated form. The lowest detection limits were 10 cfu/mL of V. alginolyticus or its inactivated form by the aptamer, and 10² cfu/mL of inactivated V. alginolyticus and 10³ cfu/mL of V. alginolyticus by the selection products. The system of detection had better affinity and specificity for V. alginolyticus and could distinguish V. alginolyticus from other aquatic pathogens such as V. harveyi, and therefore shows good application potential in the detection of pathogens in aquaculture.