Medical Micology Laboratory, Microbiology Department National School of Biological Sciences, National Polytechnique Institute (IPN), C.P. 11340, Ciudad de México, México.
Research and Development Department, Probiomed S.A. de C.V. Cruce de Carreteras Acatzingo-Zumpahuacan S/N, C.P. 52400, Tenancingo, Estado de México, México.
J Microbiol. 2019 Jun;57(6):485-497. doi: 10.1007/s12275-019-8637-2. Epub 2019 May 27.
Fusarium solani has drawn phytopathogenic, biotechnological, and medical interest. In humans, it is associated with localized infections, such as onychomycosis and keratomycosis, as well as invasive infections in immunocompromised patients. One pathogenicity factor of filamentous fungi is biofilm formation. There is still only scarce information about the in vitro mechanism of the formation and composition of F. solani biofilm. In this work, we describe the biofilm formed by a clinical keratomycosis isolate in terms of its development, composition and susceptibility to different antifungals and ultraviolet light (UV) at different biofilm formation stages. We found five biofilm formation stages using scanning electron microscopy: adherence, germination, hyphal development, maturation, and cell detachment. Using epifluorescence microscopy with specific fluorochromes, it was elucidated that the extracellular matrix consists of carbohydrates, proteins, and extracellular DNA. Specific inhibitors for these molecules showed significant biofilm reductions. The antifungal susceptibility against natamycin, voriconazole, caspofungin, and amphotericin B was evaluated by metabolic activity and crystal violet assay, with the F. solani biofilm preformation to 24 h increased in resistance to natamycin, voriconazole, and caspofungin, while the biofilm preformation to 48 h increased in resistance to amphotericin B. The preformed biofilm at 24 h protected and reduced UV light mortality. F. solani isolate could produce a highly structured extra biofilm; its cellular matrix consists of carbohydrate polymers, proteins, and eDNA. Biofilm confers antifungal resistance and decreases its susceptibility to UV light. The fungal biofilm functions as a survival strategy against antifungals and environmental factors.
茄病镰刀菌具有植物病原性、生物技术和医学方面的研究意义。在人类中，它与局部感染有关，如甲真菌病和角膜真菌病，以及免疫功能低下患者的侵袭性感染。丝状真菌的一个致病性因素是生物膜的形成。目前对于茄病镰刀菌生物膜的形成和组成的体外机制仍知之甚少。在这项工作中，我们根据体外研究描述了由临床角膜真菌病分离株形成的生物膜，包括其在不同生物膜形成阶段的发展、组成和对不同抗真菌药物和紫外线（UV）的敏感性。我们使用扫描电子显微镜发现了五个生物膜形成阶段：附着、发芽、菌丝发育、成熟和细胞脱落。使用带有特定荧光染料的荧光显微镜，阐明了细胞外基质由碳水化合物、蛋白质和细胞外 DNA 组成。这些分子的特定抑制剂显示出显著的生物膜减少。通过代谢活性和结晶紫测定评估了纳他霉素、伏立康唑、卡泊芬净和两性霉素 B 的抗真菌敏感性，与生物膜形成前 24 小时相比，纳他霉素、伏立康唑和卡泊芬净的抗真菌敏感性增加，而生物膜形成前 48 小时对两性霉素 B 的抗真菌敏感性增加。形成前 24 小时的生物膜具有保护和减少紫外线致死作用。茄病镰刀菌分离株可以产生高度结构化的额外生物膜；其细胞基质由碳水化合物聚合物、蛋白质和 eDNA 组成。生物膜赋予抗真菌药物抗性并降低其对紫外线的敏感性。真菌生物膜作为一种对抗真菌药物和环境因素的生存策略。
