茶园扁刺蛾生防菌筛选、鉴定及其发酵条件优化

贺望兴; 谢小群; 杨普香; 李文金; 李延升; 江新凤; 毛平生

doi:10.19303/j.issn.1008-0384.2022.011.012

茶园扁刺蛾生防菌筛选、鉴定及其发酵条件优化

Identification and Culture Optimization of Effective Biocontrol Agent on Thosea senensis for Tea Plantations

摘要

摘要:
目的筛选适宜生态有机茶园防治扁刺蛾的生防菌。
方法采用温度筛选法从自然罹病死亡的扁刺蛾肠道体内初筛生防菌，浸液饲喂法测定生防菌对扁刺蛾幼虫的室内毒力；综合菌株的形态特征、分子生物学鉴定以及构建的系统发育树分析进行菌株的鉴定；通过单因素和正交试验对生防菌的发酵参数条件进行优化。
结果筛选到1株对扁刺蛾幼虫具有较高防效的内生生防菌X-11，扁刺蛾3龄幼虫取食经2.56×10⁸个·mL⁻¹ X-11菌液处理的茶枝条7 d后扁刺蛾累计校正死亡率达到90.5%，致死中时（LT₅₀）仅为2.88 d。菌株X-11经16s rDNA分子生物学鉴定以及构建的系统发育树分析确定为偶氮还原拟杆菌（Paenibacillus azoreducens）。发酵条件优化试验表明，菌株X-11的适宜发酵条件为：菌株接种量2%，在发酵温度36℃条件下发酵培养60 h，菌株含量最高达1.34×10⁹个·mL⁻¹。
结论筛选到1株对扁刺蛾幼虫具有较高防效的内生生防菌x-11，可作为防治扁刺蛾幼虫的生物防治菌剂使用。

Abstract:
Objective A biocontrol microbe on Thosea senensis Walker suitable for organic tea agriculture was identified and optimal culture conditions studied.
Method From the intestines of T. senensis died from disease in nature, microbes were cultured and screened by means of varying the temperature. Indoor virulence of the selected microbes on T. senensis was determined by immersion feeding and further identified by morphological, molecular biological, and phylogenetic tree analyses. Culture of the isolated strain was optimized by univariate and orthogonal experiments.
Result The endophytic strain X-11 with high efficacy in controlling the pest was isolated. The 3^rd instar larvae of T. senensis fed with a 2.56×10⁸ conidia·mL⁻¹ X-11 solution displayed a cumulative corrected mortality rate of 90.5% with a median lethal time LT₅₀ of 2.88 d on the 7^th day after treatment. Based on the 16S rDNA molecular identification and phylogenetic analysis, X-11 was determined to be Paenibacillus azoreducens. The optimized culture conditions for the bacterium included the use of 2% inoculum to culture for 60 h at 36℃ with a final count approaching 1.34×10⁹ conidia·mL⁻¹.
Conclusion The endophytic strain with high control efficacy on T. senensis larvae was identified as P. azoreducens. It could be cultured to be used as a bioagent for the pest control at tea plantations.

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