Warwick Crop Centre, School of Life Sciences, University of Warwick, Wellesbourne, Warwick CV35 9EF, UK.
Warwick Crop Centre, School of Life Sciences, University of Warwick, Wellesbourne, Warwick CV35 9EF, UK.
J Invertebr Pathol. 2020 Nov;177:107480. doi: 10.1016/j.jip.2020.107480. Epub 2020 Oct 3.
Hypocrealean entomopathogenic fungi (EPF) (Sordariomycetes, Ascomycota) are natural regulators of insect populations in terrestrial environments. Their obligately-killing life-cycle means that there is likely to be strong selection pressure for traits that allow them to evade the effects of the host immune system. In this study, we quantified the effects of cordycepin (3'-deoxyadenosine), a secondary metabolite produced by Cordyceps militaris (Hypocreales, Cordycipitaceae), on insect susceptibility to EPF infection and on insect immune gene expression. Application of the immune stimulant curdlan (20 µg ml, linear beta-1,3-glucan, a constituent of fungal cell walls) to Drosophila melanogaster S2r+ cells resulted in a significant increase in the expression of the immune effector gene metchnikowin compared to a DMSO-only control, but there was no significant increase when curdlan was co-applied with 25 µg ml cordycepin dissolved in DMSO. Injection of cordycepin into larvae of Galleria mellonella (Lepidoptera: Pyralidae) resulted in dose-dependent mortality (LC of cordycepin = 2.1 mg per insect 6 days after treatment). Incubating conidia of C. militaris and Beauveria bassiana (Hypocreales, Cordycipitaceae; an EPF that does not synthesize cordycepin) with 3.0 mg ml cordycepin had no effect on the numbers of conidia germinating in vitro. Co-injection of G. mellonella with a low concentration of cordycepin (3.0 mg ml) plus 10 or 100 conidia per insect of C. militaris or B. bassiana caused a significant decrease in insect median survival time compared to injection with the EPF on their own. Analysis of predicted vs. observed mortalities indicated a synergistic interaction between cordycepin and the EPF. The injection of C. militaris and B. bassiana into G. mellonella resulted in increased expression of the insect immune effector genes lysozyme, IMPI and gallerimycin at 72 h post injection, but this did not occur when the EPF were co-injected with 3.0 mg ml cordycepin. In addition, we observed increased expression of IMPI and lysozyme at 48 h after injection with C. militaris, B. bassiana and sham injection (indicating a wounding response), but this was also prevented by application of cordycepin. These results suggest that cordycepin has potential to act as a suppressor of the immune response during fungal infection of insect hosts.
Hypocrealean 昆虫病原真菌（EPF）（Sordariomycetes，Ascomycota）是陆地环境中昆虫种群的天然调节剂。它们的专性致死生命周期意味着，它们很可能会受到强烈的选择压力，以产生允许它们逃避宿主免疫系统影响的特性。在这项研究中，我们量化了蛹虫草（Hypocreales，Cordycipitaceae）产生的次生代谢物虫草素（3'-脱氧腺苷）对昆虫对 EPF 感染的敏感性和对昆虫免疫基因表达的影响。用免疫刺激剂几丁质（20 µg ml，线性β-1,3-葡聚糖，真菌细胞壁的组成部分）处理黑腹果蝇 S2r+细胞，与仅用 DMSO 处理的对照组相比，免疫效应基因 metchnikowin 的表达显著增加，但当几丁质与 25 µg ml 溶于 DMSO 的虫草素一起应用时，没有显著增加。将虫草素注入欧洲金龟子幼虫（鳞翅目：Pyralidae）中，导致剂量依赖性死亡率（处理后 6 天，虫草素的 LC 为每只昆虫 2.1 毫克）。将虫草素（3.0 mg ml）与蛹虫草和球孢白僵菌（Hypocreales，Cordycipitaceae；一种不合成虫草素的 EPF）的分生孢子一起孵育，对体外萌发的分生孢子数量没有影响。与单独注射 EPF 相比，向欧洲金龟子中共同注射低浓度的虫草素（3.0 mg ml）加每只昆虫 10 或 100 个分生孢子的虫草素或白僵菌，导致昆虫中位生存时间显著缩短。对预测死亡率与观察死亡率的分析表明，虫草素与 EPF 之间存在协同相互作用。将虫草素和白僵菌注入欧洲金龟子中，导致昆虫免疫效应基因溶菌酶、IMPI 和 gallerimycin 在注射后 72 小时表达增加，但当 EPF 与 3.0 mg ml 虫草素共同注射时则不会发生这种情况。此外，我们观察到在注射虫草素、白僵菌和假注射（表明创伤反应）后 48 小时，IMPI 和溶菌酶的表达增加，但这也被虫草素的应用所阻止。这些结果表明，虫草素有可能在真菌感染昆虫宿主时充当免疫反应的抑制剂。
