Transcriptomic analyses reveal comprehensive responses of insect hemocytes to mycopathogen Beauveria bassiana, and fungal virulence-related cell wall protein assists pathogen to evade host cellular defense

Entomopathogenic fungi naturally infect insect hosts in environment. Fungal invasion and host immune defense are still in the progress of co-evolution. In this study, entomopathogenic fungus <i>Beauveria bassiana</i> and lepidopteran insect <i>Galleria mellonella</i> were used to investigate host cellular immunity and fungal strategy to evade host defense. First of all, genome-wide expression revealed the transcriptomic responses of hemocytes to insect mycopathogen, which dynamically varied during infection process. Enrichment analysis indicated that differentially expressed genes were primarily involved in metabolism, cellular process and immune system. Notably, cellular response involved a series of hydrolytic enzyme and antimicrobial peptide genes which were sorted together in clustering analysis. In <i>B. bassiana</i>, a cell-wall protein gene (<i>BbCwp</i>) contributes to fungal development in host hemocoel and virulence. RT-qPCR analyses indicated that infection by Δ<i>BbCwp</i> mutant strain caused the up-regulated expression of a series of immunity-related genes, including β-1, 3-glucan recognition protein, hydrolytic enzyme and antimicrobial peptide genes. Disruption of <i>BbCwp</i> resulted in a significant change in conidial lectin-binding feature and the enhanced encapsulation by the host hemocytes. After being treated with hydrolytic enzymes, Δ<i>BbCwp</i> mutant displayed a significantly enhanced sensitivity to osmotic and oxidative stresses. In conclusion, fungal invasion initiates comprehensive physiological responses in the host hemocytes. For mycopathogen, cell-wall protein plays an important role in fungal evasion of immunity defense and colonization in host. Our studies provide an initial framework for exploring more mechanistic details about the fungus–host interaction.

虫生真菌（Entomopathogenic fungi）可在自然环境中侵染昆虫宿主。真菌侵染与宿主免疫防御始终处于协同进化的动态进程中。本研究以球孢白僵菌（Beauveria bassiana）与鳞翅目昆虫大蜡螟（Galleria mellonella）为模型，探究宿主细胞免疫机制与真菌逃逸宿主免疫防御的策略。首先，全基因组表达分析揭示了血细胞（hemocytes）对病原真菌的转录组响应，该响应在侵染过程中呈动态变化。富集分析结果显示，差异表达基因主要参与代谢过程、细胞进程与免疫系统调控。值得注意的是，细胞响应相关的一系列水解酶与抗菌肽基因在聚类分析中被聚为一类。在球孢白僵菌中，细胞壁蛋白基因BbCwp可促进真菌在宿主血腔（hemocoel）中的定殖与毒力。实时定量聚合酶链反应（RT-qPCR）分析显示，ΔBbCwp突变菌株侵染宿主后，会诱导一系列免疫相关基因的上调表达，其中包括β-1,3-葡聚糖识别蛋白、水解酶与抗菌肽基因。敲除BbCwp会导致分生孢子（conidia）的凝集素结合特性发生显著改变，并增强宿主血细胞对其的包囊作用。经水解酶处理后，ΔBbCwp突变菌株对渗透压与氧化胁迫的敏感性显著升高。综上，真菌侵染可触发宿主血细胞的全面生理响应。对于病原真菌而言，细胞壁蛋白在其逃逸宿主免疫防御与定殖宿主的过程中发挥关键作用。本研究为深入探究真菌-宿主互作的更多机制细节提供了初步的研究框架。