Department of Molecular Microbiology and Immunology, Brown University, Providence, RI United States of America.
PLoS One. 2018 Sep 5;13(9):e0201915. doi: 10.1371/journal.pone.0201915. eCollection 2018.
Pseudogymnoascus destructans is the fungal pathogen responsible for White-nose Syndrome (WNS), a disease that has killed millions of bats in North America over the last decade. A major obstacle to research on P. destructans has been the lack of a tractable infection model for monitoring virulence. Here, we establish a high-throughput model of infection using larvae of Galleria mellonella, an invertebrate used to study host-pathogen interactions for a wide range of microbial species. We demonstrate that P. destructans can kill G. mellonella larvae in an inoculum-dependent manner when infected larvae are housed at 13°C or 18°C. Larval killing is an active process, as heat-killed P. destructans spores caused significantly decreased levels of larval death compared to live spores. We also show that fungal spores that were germinated prior to inoculation were able to kill larvae 3-4 times faster than non-germinated spores. Lastly, we identified chemical inhibitors of P. destructans and used G. mellonella to evaluate these inhibitors for their ability to reduce virulence. We demonstrate that amphotericin B can effectively block larval killing by P. destructans and thereby establish that this infection model can be used to screen biocontrol agents against this fungal pathogen.
拟青霉属(Pseudogymnoascus)破坏菌是导致白鼻综合征(WNS)的真菌病原体,这种疾病在过去十年中导致了北美的数百万只蝙蝠死亡。研究拟青霉属破坏菌的一个主要障碍是缺乏一种可控制的感染模型来监测其毒力。在这里,我们使用黄粉虫幼虫建立了一种高通量的感染模型,黄粉虫是一种用于研究宿主-病原体相互作用的无脊椎动物,适用于多种微生物物种。我们证明,当感染幼虫在 13°C 或 18°C 下饲养时,拟青霉属破坏菌可以以剂量依赖的方式杀死黄粉虫幼虫。幼虫死亡是一个主动的过程,因为与活孢子相比,热杀死的拟青霉属破坏菌孢子导致幼虫死亡率显著降低。我们还表明,在接种前萌发的真菌孢子能够比未萌发的孢子更快地杀死幼虫 3-4 倍。最后,我们鉴定了拟青霉属破坏菌的化学抑制剂,并使用黄粉虫幼虫评估这些抑制剂降低毒力的能力。我们证明两性霉素 B 可以有效地阻止拟青霉属破坏菌对幼虫的杀伤,从而确立了这种感染模型可用于筛选针对这种真菌病原体的生物防治剂。