Entomology Research Laboratory, Department of Plant and Soil Science, University of Vermont, 661 Spear Street, Burlington, VT 05405-0105, USA.
Appl Microbiol Biotechnol. 2010 Aug;87(5):1639-48. doi: 10.1007/s00253-010-2543-1. Epub 2010 May 6.
Insect killing fungi have high potential in controlling agriculturally harmful pest, but their slow progress and high variation in killing insect are major impediments to successful industrialization. The present work describes the use of supernatant from the liquid culture of Beauveria bassiana SFB-205 to surmount this problem, particularly efficient production of thermotolerant chitinase, which is one of the major pathogenesis-related enzymes in the supernatant. The chitinase was precipitated using varying mineral precipitants and followed by lyophilization, which was compared with a salting out method using ammonium sulfate in effectiveness. Incorporating of the supernatant fraction of the Beauveria preparation with attagel at 0.5% (w/v) as a precipitant enabled this treatment to show the greatest chitinase-precipitation efficiency (93.4%), followed up with excellent insecticidal activity against cotton aphids when it was mixed with 0.01% (v/v) polyoxyethylene-(3)-isotridecyl ether (TDE-3) as a spreading agent in laboratory conditions. Consequently, lyophilized attagel-mediated precipitation pellet was superior to lyophilized salting out pellet in maintaining chitinase activity against a thermal stress at 50 degrees C. This finding provides that the attagel-mediated precipitation can be exploited to improve the thermotolerance of B. bassiana SFB-205 chitinase as a novel strategy for biopesticide production.
杀虫真菌在防治农业害虫方面具有很大的潜力，但杀虫速度慢、变异性大，是成功实现工业化的主要障碍。本研究描述了利用球孢白僵菌 SFB-205 的液体培养上清液来克服这一问题，特别是高效生产热稳定的几丁质酶，这是上清液中主要的与发病相关的酶之一。采用不同的矿物沉淀剂沉淀几丁质酶，然后进行冻干，与硫酸铵盐析法进行比较，以评估其效果。在 0.5%（w/v）的 Beauveria 制剂上清液中加入 attagel 作为沉淀剂，这种处理方法显示出最高的几丁质酶沉淀效率（93.4%），随后在实验室条件下，当与 0.01%（v/v）聚氧乙烯-（3）-异十三烷基醚（TDE-3）作为展着剂混合时，对棉蚜表现出优异的杀虫活性。因此，在 50°C 的热应激下，冻干 attagel 介导的沉淀颗粒比冻干盐析沉淀颗粒更能保持几丁质酶的活性。这一发现表明，attagel 介导的沉淀可以被用来提高 B. bassiana SFB-205 几丁质酶的耐热性，作为生物农药生产的一种新策略。
