东北地区水稻、小麦和玉米镰孢菌鉴定与ZEN毒素检测

Identification of Fusarium spp. Isolated in Northeastern China and Detection of ZEN Toxin

在黑龙江、吉林和辽宁三省的13个市县,水稻、小麦和玉米的主要产区共分离到214株镰孢菌株,其中分离频率最高的是禾谷镰孢菌(58.1%)。镰孢菌株产生玉米赤霉烯酮(ZEN),具有较强生物毒性,对人畜有很强的毒害作用,因此检测作物中玉米赤霉烯酮含量是十分必要的。单克隆抗体进行特异性分析结果显示,抗-ZEN单克隆抗体与ZEN、α-玉米赤霉醇、β-玉米赤霉醇和β-玉米赤霉烯醇结合率分别为100%、188.7%、43.9%和25.6%,而与脱氧雪腐镰刀菌烯醇和黄曲霉毒素的结合率则均小于1.0%,由此证明试验制备的单克隆抗体具有较高特异性。以该单抗为依据建立的ELISA检测法,检测限为0.4 ng/m L,检测区间为0.4~25.00 ng/m L,曲线回归方程y=-0.7673x+3.1318,与相似毒素的交叉反应率均小于1.0%,回收率在79%~112%之间。用ELISA产毒能范围在0.27~1.61μg/L,其中有6株禾谷镰孢菌(Fusarium graminearum)菌株具有产毒能力。研究结果说明建立的间接竞争ELISA方法可以用于检测玉米赤霉烯酮。

214 strains of Fusarium spp. were isolated in 13 cities and counties of the main producing areas of rice, wheat and maize in Heilongjiang, Jilin and Liaoning provinces. Of which, the highest isolation frequency was Fusarium graminearum （58.1%）. Fusarium spp. produces zearalenone （ZEN）, which has a strong biological toxicity and makes a strong poisoning effect to humans and animals. Therefore, the detection of zearalenone content in crops is very necessary. In this paper, ZEN-BSA conjugated to bovine serum albumin （ZEN-BSA） was used as antigen to immunize mice. The results of specific analysis for monoclonal antibodies showed that the binding rates of anti-ZEN monoclonal antibody to ZEN, α-zeranol, β-zeranol and β-zearalenol were 100%, 188.7%, 43.9% and 25.6%, respectively. And the binding rate to deoxynivalenol and aflatoxin was less than 1.0%, which proved that the monoclohal antibody prepared by the experiment had higher specificity. The monoclonal antibody was prepared for ELISA detection method based on the model, the detection limit is 0.4 ng/mL, the detection range of 0.4-25.00 ng/mL, the curve regression equation is y=-0.7673x＋3.1318, and the cross reaction of similar toxins were less than 1%, the recovery was 79%-112%. The range of toxin production with ELISA was 0.27-1.61μg/L. There were 6 strains of Fusarium graminearum toxin producing strains with ability. The results showed that the indirect competitive ELISA method can be used for the detection of zearalenone.