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Zhen Li,1,* Bin Zhu,2,* Weiqin Chen,1 Jun Hu,1 Yingjun Xue,1 Hongmei Yin,1 Xiaobo Hu,3 Weiwei Liu1 1Department of Laboratory Medicine, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, 200032, People’s Republic of China; 2Department of Rehabilitation, Ruijin Hospital, Shanghai Jiaotong University School of Medicine, Shanghai, 200025, People’s Republic of China; 3General Office, Centre for Clinical Laboratory, Shanghai, 200126, People’s Republic of China*These authors contributed equally to this workCorrespondence: Weiwei Liu, Department of Laboratory Medicine, Longhua Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, 200032, People’s Republic of China, Email huashanvivian@126.com Xiaobo Hu, General Office, Centre for Clinical Laboratory, Shanghai, 200126, People’s Republic of China, Email huxiaobo@vip.sina.comPurpose: The non-albicans Candida (NAC) species have recently gained great importance worldwide due to the increasing proportion in candida causing bloodstream infections. This investigation aimed to explore the efficacy of Pseudolaric acid A (PAA, a diterpenoid derived from Pseudolarix kaempferi) and its synergistic effect with fluconazole (FLC) against NAC species, including C. tropicalis, C. parapsilosis complex, and C. glabrata.Methods: The microdilution checkerboard assay and time-killing curves were performed to detect the antifungal efficiency. To examine the integrity of cell walls and membranes, calcofluor white stain and propidium iodide stain were used. The changes of intracellular ultrastructure in Candida cells after treatment were observed using transmission electron microscopy. Changes in cell viability with the autophagy inhibitor 3-MA were assessed by the XTT method.Results: It was revealed that PAA alone is effective on C. tropicalis, C. parapsilosis sensu stricto, C. orthopsilosis, and C. metapsilosis (MIC 8– 128 μg/mL). Strong synergism against FLC-resistant C. tropicalis was observed (FICI 0.07– 0.281), when PAA and FLC were combined. PAA had dose-dependently detrimental effects on C. tropicalis cell membranes. Moreover, increased vacuoles and autophagosome formation were found in C. tropicalis exposed to PAA. And the inhibitory effect of PAA against C. tropicalis can be relieved by autophagy inhibitor 3-MA in a certain concentration range. Ultrastructural alterations of C. tropicalis were more pronounced under the combination of PAA and FLC, including separation of the cell membrane from the cell wall, increased number of vacuoles, and degradation of organelles.Conclusion: These observations indicated that PAA and its combination with FLC could be a promising therapeutic candidate for treating infections caused by NAC species.Keywords: pseudolaric acid A, fluconazole, antifungal, combination, non-albicans Candida, synergy, autophagy |
