Binder Ulrike, Navarro-Mendoza Maria Isabel, Naschberger Verena, Bauer Ingo, Nicolas Francisco E, Pallua Johannes D, Lass-Flörl Cornelia, Garre Victoriano
Division of Hygiene and Medical Microbiology, Medical University Innsbruck, Schöpfstrasse 41, 6020 Innsbruck, Austria.
Departamento de Genética y Microbiología, Facultad de Biología, Universidad de Murcia, 30100 Murcia, Spain.
Genes (Basel). 2018 Dec 7;9(12):613. doi: 10.3390/genes9120613.
Invasive fungal infections caused by Mucorales (mucormycosis) have increased worldwide. These life-threatening infections affect mainly, but not exclusively, immunocompromised patients, and are characterized by rapid progression, severe tissue damage and an unacceptably high rate of mortality. Still, little is known about this disease and its successful therapy. New tools to understand mucormycosis and a screening method for novel antimycotics are required. Bioluminescent imaging is a powerful tool for in vitro and in vivo approaches. Hence, the objective of this work was to generate and functionally analyze bioluminescent reporter strains of , one mucormycosis-causing pathogen. Reporter strains were constructed by targeted integration of the firefly luciferase gene under control of the promoter P. The luciferase gene was sufficiently expressed, and light emission was detected under several conditions. Phenotypic characteristics, virulence potential and antifungal susceptibility were indifferent to the wild-type strains. Light intensity was dependent on growth conditions and biomass, being suitable to determine antifungal efficacy in vitro. This work describes for the first time the generation of reporter strains in a basal fungus that will allow real-time, non-invasive infection monitoring in insect and murine models, and the testing of antifungal efficacy by means other than survival.
由毛霉目真菌引起的侵袭性真菌感染（毛霉病）在全球范围内呈上升趋势。这些危及生命的感染主要（但不限于）影响免疫功能低下的患者，其特点是进展迅速、组织严重受损且死亡率高得令人难以接受。然而，人们对这种疾病及其有效治疗方法仍知之甚少。需要有了解毛霉病的新工具以及新型抗真菌药物的筛选方法。生物发光成像对于体外和体内研究是一种强大的工具。因此，本研究的目的是构建并功能分析一种引起毛霉病的病原体——的生物发光报告菌株。通过在启动子P的控制下靶向整合萤火虫荧光素酶基因构建报告菌株。荧光素酶基因得到充分表达，并且在多种条件下检测到发光。其表型特征、毒力潜力和抗真菌敏感性与野生型菌株无差异。发光强度取决于生长条件和生物量，适用于体外确定抗真菌效果。本研究首次描述了在一种基础真菌中构建报告菌株，这将允许在昆虫和小鼠模型中进行实时、非侵入性感染监测，并通过存活率以外的方法测试抗真菌效果。
