School of Life Sciences, Beijing Institute of Technology, Beijing 100081, China.
State Key Laboratory for Biology of Plant Diseases and Insect Pests, Institute of Plant Protection, Chinese Academy of Agricultural Sciences, Beijing 100193, China.
Toxins (Basel). 2020 Apr 13;12(4):252. doi: 10.3390/toxins12040252.
(Bt) is used for insect pest control, and its larvicidal activity is primarily attributed to Cry toxins. Other factors participate in infection, and limited information is available regarding factors acting on the peritrophic matrix (PM). This study aimed to investigate the role of a Bt chitin-binding protein (CBPA) that had been previously shown to be expressed at pH 9 and could therefore be expressed in the alkaline gut of lepidopteron larvae. A ∆cbpA mutant was generated that was 10-fold less virulent than wild-type Bt HD73 towards neonate larvae, indicating its important role in infection. Purified recombinant CBPA was shown to have a chitin affinity, thus indicating a possible interaction with the chitin-rich PM. A translational GFP-CBPA fusion elucidated the localization of CBPA on the bacterial surface, and the transcriptional activity of the promoter P was immediately induced and confirmed at pH 9. Next, in order to connect surface expression and possible gut activity last instar (Gm) larvae (not susceptible to Bt HD-73) were used as a model to follow CBPA in gut expression, bacterial transit, and PM adhesion. CBPA-GFP was quickly expressed in the Gm gut lumen, and more Bt HD73 strain bacteria adhered to the PM than those of the ∆cbpA mutant strain. Therefore, CBPA may help to retain the bacteria, via the PM binding, close to the gut surface and thus takes part in the early steps of Bt gut interactions.
（Bt）被用于防治害虫，其杀虫活性主要归因于 Cry 毒素。其他因素也参与了感染，而对于作用于围食膜（PM）的因素，我们知之甚少。本研究旨在研究一种 Bt 几丁质结合蛋白（CBPA），该蛋白先前已被证明在 pH9 下表达，因此可以在鳞翅目幼虫的碱性肠道中表达。生成了一个 ∆cbpA 突变体，其对 neonate 幼虫的毒力比野生型 Bt HD73 低 10 倍，表明其在感染中起重要作用。纯化的重组 CBPA 显示出对几丁质的亲和力，因此表明与富含几丁质的 PM 可能存在相互作用。GFP-CBPA 的翻译融合阐明了 CBPA 在细菌表面的定位，启动子 P 的转录活性在 pH9 时立即被诱导并得到证实。接下来，为了将表面表达和可能的肠道活性联系起来，最后一龄幼虫（对 Bt HD-73 不敏感）被用作模型，以跟踪 CBPA 在肠道表达、细菌转移和 PM 粘附中的作用。CBPA-GFP 在 Gm 肠道腔中迅速表达，并且与 ∆cbpA 突变体菌株相比，更多的 Bt HD73 菌株细菌粘附到 PM 上。因此，CBPA 可能通过与 PM 结合，帮助将细菌保留在靠近肠道表面的位置，从而参与 Bt 肠道相互作用的早期步骤。
