哈茨木霉 CBS226.95 的分离鉴定及产几丁质酶工艺优化

在植物病害的防治中，生物防治具有安全和环保的优点，丝状真菌哈茨木霉（Trichoderma harzianum）及其近缘种能抑制多种植物病原菌的生长。文章从腐烂的猕猴桃果实中获得一株分离菌株CBS226.95，并对分离株进行体外拮抗试验，通过形态学及ITS、BenA和TEF1多基因序列方法对分离株进行鉴定，以单因素实验和响应面试验设计对菌株几丁质酶生产的发酵条件进行优化。结果显示，分离株能够抑制葡萄座腔菌（Botryosphaeria dothidea）、拟茎点霉菌（Diaporthe）和灰霉菌（Botrytis cinerea）的生长，抑制率分别为45.35%、44.99%、47.31%。综合形态学特征和系统发育分析结果可确定菌株CBS226.95为哈茨木霉。单因素实验和响应面试验分析结果发现，菌株CBS226.95-4发酵的最适碳源为葡萄糖，最适氮源为蛋白胨，最佳发酵条件为培养温度33.23℃、培养时间4.11d、pH值6.108。该菌株的分离鉴定及发酵条件优化为木霉的生防应用和几丁质酶的工业化生产提供了技术支持。

In the plant disease management, biological control offers distinct advantages in terms of safety and environmental friendliness. Filamentous fungi such as Trichoderma harzianum and its related strains exhibit broad-spectrum inhibitory effects against various plant pathogens. A fungal strain CBS226.95 was isolated from decayed kiwifruit and evaluated through in vitro antagonist assays. The strain identification was confirmed through morphological features and sequencing information of multiple genes, including ITS, BenA, and TEF1. Subsequently, fermentation conditions of CBS226.95 were optimized to improve chitinase production by employing a one-factor-at-a-time approach followed by a response surface methodology design. The results demonstrated that CBS226.95 significantly inhibited the growth of plant pathogens Botryosphaeria do-thidea, Diaporthe, and Botrytis cinerea, exhibiting inhibition rates of 45.35%, 44.99%, and 47.31%, respectively. Through a comprehensive integration of morphological traits and phylogenetic analyses, CBS226.95 was identified as Trichoderma harzianum. The one-factor-at-a-time and response surface experiments revealed glucose as the optimal carbon source and peptone as the preferred nitrogen source for increasing fermentation efficiency of CBS226.95-4. Additionally, the most favorable fermentation conditions were determined to be a temperature of 33.23 °C, a duration of 4.11 days, and a pH level of 6.108. The isolation, identification, and optimization of fermentation conditions of Trichoderma harzianum CBS226.95-4 provide technical support for the biocontrol application of Trichoderma and the industri- al production of chitinase.