Infections of Exophiala dermatitidis are often chronic and recalcitrant. Combination therapies with novel compounds and azoles could be an effective solution. Previously, we have demonstrated that pyrvinium pamoate exerted antifungal activity alone and favorable synergy with azoles against planktonic E. dermatitidis. Herein, the underlying antifungal mode of action were investigated. Pyrvinium alone showed sessile MIC50 (SMIC50) of 8->16 μg/ml against E. dermatitidis biofilms. However, synergism of PP with itraconazole, voriconazole, and posaconazole were observed against 16 (88.9%), 9 (50%), and 13 (72.2%) strains of E. dermatitidis biofilms. In accordance with in vitro susceptibilities, pyrvinium alone at concentration of 2 μg/ml resulted in significant growth restriction of planktonic E. dermatitidis. Pyrvinium alone resulted in reduction of biofilm formation. Higher concentration of pyrvinium was associate with more progressive reduction of biofilm mass. The in vivo activity of pyrvinium alone and combined with azoles was evaluated using Galleria mellonella model. Pyrvinium alone significantly improved the survival rate of larvae (P < 0.0001). The combination of pyrvinium and voriconazole or posaconazole acted synergistically in vivo (P < 0.05). Fungal burden determination revealed significant reduction of numbers of colony forming unit (CFU) in larvae treated with pyrvinium-itraconazole and pyrvinium-posaconazole compared to itraconazole or posaconazole alone group, respectively. The effect of pyrvinium on apoptosis, expression of TOR and HSP90, and drug efflux reversal were evaluated by PI/Annexin V staining, Real-Time Quantitative PCR and Rhodamine 6G assay, respectively. Pyrvinium alone or combined with azoles significantly (P < 0.05) increased late apoptosis or necrosis of E. dermatitidis cells. Pyrvinium combined with posaconazole significantly decreased the expression of TOR and Hsp90 compared to posaconazole alone group (P < 0.05). Pyrvinium resulted in significant (P < 0.05) decrease of the efflux of Rhodamine 6G. These findings suggested pyrvinium could be a promising synergist with azoles. The underlying mechanisms could be explained by inducing apoptosis/necrosis, inhibition of drug efflux pumps, and signaling pathways related with stress response and growth control.

感染皮炎外口藻往往是慢性且顽固的。新型化合物和唑类的联合疗法可能是一种有效的解决方案。此前，我们已经证明双羟萘酸吡维铵单独发挥抗真菌活性，并与唑类药物发挥良好的协同作用，对抗浮游生物皮炎艾美耳球菌。在此，研究了潜在的抗真菌作用模式。单独的吡维铵显示出 8->16 μg/ml 的固着 MIC50 (SMIC50)皮炎艾美耳球菌生物膜。然而，观察到 PP 与伊曲康唑、伏立康​​唑和泊沙康唑对 16 种（88.9%）、9 种（50%）和 13 种（72.2%）菌株有协同作用。皮炎艾美耳球菌生物膜。依据体外敏感性，单独使用浓度为 2 μg/ml 的吡维铵会导致浮游生物的显着生长限制皮炎艾美耳球菌。单独的吡维铵导致生物膜形成减少。较高浓度的吡维铵与生物膜质量的进一步减少有关。这体内使用吡维铵单独和与唑类组合的活性进行评估大蜡螟模型。单独使用吡维铵可显着提高幼虫的存活率（磷< 0.0001)。吡维铵与伏立康唑或泊沙康唑的组合具有协同作用体内(磷< 0.05）。真菌负荷测定显示，与单独使用伊曲康唑或泊沙康唑组相比，用吡维铵-伊曲康唑和吡维铵-泊沙康唑处理的幼虫的菌落形成单位（CFU）数量分别显着减少。 吡维铵对细胞凋亡、表达的影响托罗和热休克蛋白90和药物外流逆转分别通过 PI/Annexin V 染色、实时定量 PCR 和罗丹明 6G 测定进行评估。吡维铵单独使用或与唑类合用显着（磷< 0.05) 晚期细胞凋亡或坏死增加皮炎艾美耳球菌细胞。吡维铵联合泊沙康唑显着降低托罗和热休克蛋白90与单独泊沙康唑组相比（磷< 0.05）。吡维铵导致显着（磷< 0.05) 罗丹明 6G 的流出减少。这些发现表明吡维铵可能是一种有前途的唑类增效剂。其潜在机制可以通过诱导细胞凋亡/坏死、抑制药物外排泵以及与应激反应和生长控制相关的信号通路来解释。