Phospholipid/protein co-mediated assembly of Cu2O nanoparticles for specific inhibition of growth and biofilm formation of pathogenic fungi

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  • ReceivedJun 9, 2020
  • AcceptedJul 6, 2020
  • PublishedOct 16, 2020


Funded by

the National Natural Science Foundation of China(31870139,81873961)

the Natural Science Foundation of Tianjin(19JCZDJC33800)

the National Training Program of Innovation and Entrepreneurship for Undergraduates(201810055105)

and the Fundamental Research for the Central Universities.


This work was supported by the National Natural Science Foundation of China (31870139 and 81873961), the Natural Science Foundation of Tianjin (19JCZDJC33800), the National Training Program of Innovation and Entrepreneurship for Undergraduates (201810055105), and the Fundamental Research for the Central Universities.

Interest statement

The authors declare that they have no conflict of interest.

Contributions statement

Peng L, Wei H, and Yu Q designed and performed the experiments, and wrote the paper; Tian L and Xu J performed the data analysis; Yu Q and Li M supervised this study. All authors contributed to the general discussion and revision of the manuscript.

Author information

Liping Peng is currently a PhD student at the Department of Microbiology, College of Life Sciences, Nankai University. Her research focuses on the development of antimicrobial nanosystems for treatment of fungal/bacterial infections, together with investigation of the virulence factors of clinical pathogens.

Henan Wei is currently an undergraduate at the Department of Microbiology, College of Life Sciences, Nankai University. Her research focuses on the design of antifungal nanoplatforms for efficient anti-Candida therapy.

Qilin Yu received his PhD degree in 2013 from the College of Life Sciences, Nankai University. In 2017, he was appointed associated professor in Nankai University. In 2019, he started his visiting scholar research in Prof. Jeffrey Zink’s Lab, Department of Chemistry and Biochemistry, University of California, Los Angeles. His research interests focus on the design and application of nanotechnology-based self-assembly of biological systems.


Supplementary information

Supporting data are available in the online version of the paper.


[1] Crunkhorn S. Fungal infection: Protecting from Candida albicans. Nat Rev Drug Discov, 2016, 15: 604 CrossRef PubMed Google Scholar

[2] Schwartz IS, Govender NP, Sigler L, et al. Emergomyces: The global rise of new dimorphic fungal pathogens. PLoS Pathog, 2019, 15: e1007977 CrossRef PubMed Google Scholar

[3] Lalla RV, Latortue MC, Hong CH, et al. A systematic review of oral fungal infections in patients receiving cancer therapy. Support Care Cancer, 2010, 18: 985-992 CrossRef PubMed Google Scholar

[4] Alangaden GJ. Nosocomial fungal infections: Epidemiology, infection control, and prevention. Infect Dis Clinics North Am, 2011, 25: 201-225 CrossRef PubMed Google Scholar

[5] Moyes DL, Wilson D, Richardson JP, et al. Candidalysin is a fungal peptide toxin critical for mucosal infection. Nature, 2016, 532: 64-68 CrossRef PubMed ADS Google Scholar

[6] Lee KT, So YS, Yang DH, et al. Systematic functional analysis of kinases in the fungal pathogen Cryptococcus neoformans. Nat Commun, 2016, 7: 12766 CrossRef PubMed ADS Google Scholar

[7] Fuller KK, Ringelberg CS, Loros JJ, et al. The fungal pathogen Aspergillus fumigatus regulates growth, metabolism, and stress resistance in response to light. mBio, 2013, 4: e00142-13. Google Scholar

[8] Mayer FL, Wilson D, Hube B. Candida albicans pathogenicity mechanisms. Virulence, 2013, 4: 119-128 CrossRef PubMed Google Scholar

[9] Lewis K. Platforms for antibiotic discovery. Nat Rev Drug Discov, 2013, 12: 371-387 CrossRef PubMed Google Scholar

[10] Peng XM, Cai GX, Zhou CH. Recent developments in azole compounds as antibacterial and antifungal agents. Curr Top Med Chem, 2013, 13: 1963-2010 CrossRef PubMed Google Scholar

[11] Pfaller MA, Carvalhaes C, Messer SA, et al. Activity of a long-acting echinocandin, rezafungin, and comparator antifungal agents tested against contemporary invasive fungal isolates (SENTRY Program, 2016 to 2018). Antimicrob Agents Chemother, 2020, 64: e00099-20. Google Scholar

[12] Low ZJ, Xiong J, Xie Y, et al. Discovery, biosynthesis and antifungal mechanism of the polyene-polyol meijiemycin. Chem Commun, 2020, 56: 822-825 CrossRef PubMed Google Scholar

[13] Berman J, Krysan DJ. Drug resistance and tolerance in fungi. Nat Rev Microbiol, 2020, 18: 319-331 CrossRef PubMed Google Scholar

[14] Whaley SG, Berkow EL, Rybak JM, et al. Azole antifungal resistance in Candida albicans and emerging non-albicans Candida species. Front Microbiol, 2017, 7: 2173 CrossRef PubMed Google Scholar

[15] Chang Z, Yadav V, Lee SC, et al. Epigenetic mechanisms of drug resistance in fungi. Fungal Genets Biol, 2019, 132: 103253 CrossRef PubMed Google Scholar

[16] Feng Y, Zhu J. Copper nanomaterials and assemblies for soft electronics. Sci China Mater, 2019, 62: 1679-1708 CrossRef Google Scholar

[17] Huang WC, Lyu LM, Yang YC, et al. Synthesis of Cu2O nanocrystals from cubic to rhombic dodecahedral structures and their comparative photocatalytic activity. J Am Chem Soc, 2012, 134: 1261-1267 CrossRef PubMed Google Scholar

[18] Chakraborty D, Nandi S, Mullangi D, et al. Cu/Cu2O nanoparticles supported on a phenol-pyridyl COF as a heterogeneous catalyst for the synthesis of unsymmetrical diynes via glaser-hay coupling. ACS Appl Mater Interfaces, 2019, 11: 15670-15679 CrossRef PubMed Google Scholar

[19] Woźniak-Budych MJ, Przysiecka Ł, Maciejewska BM, et al. Facile synthesis of sulfobetaine-stabilized Cu2O nanoparticles and their biomedical potential. ACS Biomater Sci Eng, 2017, 3: 3183-3194 CrossRef Google Scholar

[20] Giannousi K, Sarafidis G, Mourdikoudis S, et al. Selective synthesis of Cu2O and Cu/Cu2O NPs: Antifungal activity to yeast Saccharomyces cerevisiae and DNA interaction. Inorg Chem, 2014, 53: 9657-9666 CrossRef PubMed Google Scholar

[21] Zhou J, Xiang H, Zabihi F, et al. Intriguing anti-superbug Cu2O@ZrP hybrid nanosheet with enhanced antibacterial performance and weak cytotoxicity. Nano Res, 2019, 12: 1453-1460 CrossRef Google Scholar

[22] Du BD, Phu DV, Quoc LA, et al. Synthesis and investigation of antimicrobial activity of Cu2O nanoparticles/zeolite. J Nanoparticles, 2017, 2017: 1-6 CrossRef Google Scholar

[23] Yang J, Dong H. Fungistatic effects of nano-Cu2O suspension with different concentrations against five plant pathogenic fungi (in Chinese). J Changjiang Vegetables, 2014, 8: 67–69. Google Scholar

[24] Hongtao MA, Yang L, Dong H. Inhibitory effects of original nano-Cu2O drug and nano-Cu2O suspension on snake melon Botrytis cinereal (in Chinese). Agr Sci Technol, 2015, 16: 526–561. Google Scholar

[25] Yang J, Dong H, Li Y, et al. Studies on inhibitory effects of nano-Cu2O on Phytophthora capcisi and Fusarium oxysporum of pepper (in Chinese). China Vegetables, 2012, 6: 79–81. Google Scholar

[26] Fan W, Shi Z, Yang X, et al. Bioaccumulation and biomarker responses of cubic and octahedral Cu2O micro/nanocrystals in Daphnia magna. Water Res, 2012, 46: 5981-5988 CrossRef PubMed Google Scholar

[27] Chen D, Zhang D, Yu JC, et al. Effects of Cu2O nanoparticle and CuCl2 on zebrafish larvae and a liver cell-line. Aquat Toxicol, 2011, 105: 344-354 CrossRef PubMed Google Scholar

[28] Grzelczak M, Liz-Marzán LM, Klajn R. Stimuli-responsive self-assembly of nanoparticles. Chem Soc Rev, 2019, 48: 1342-1361 CrossRef PubMed Google Scholar

[29] Zhuang J, Gordon MR, Ventura J, et al. Multi-stimuli responsive macromolecules and their assemblies. Chem Soc Rev, 2013, 42: 7421-7435 CrossRef PubMed Google Scholar

[30] Zhao Y, Thorkelsson K, Mastroianni AJ, et al. Small-molecule-directed nanoparticle assembly towards stimuli-responsive nanocomposites. Nat Mater, 2009, 8: 979-985 CrossRef PubMed ADS Google Scholar

[31] Voltà-Durán E, Cano-Garrido O, Serna N, et al. Controlling self-assembling and tumor cell-targeting of protein-only nanoparticles through modular protein engineering. Sci China Mater, 2020, 63: 147-156 CrossRef Google Scholar

[32] Yu Q, Deng T, Lin FC, et al. Supramolecular assemblies of heterogeneous mesoporous silica nanoparticles to co-deliver antimicrobial peptides and antibiotics for synergistic eradication of pathogenic biofilms. ACS Nano, 2020, 14: 5926-5937 CrossRef PubMed Google Scholar

[33] Nam J, La WG, Hwang S, et al. pH-responsive assembly of gold nanoparticles and “spatiotemporally concerted” drug release for synergistic cancer therapy. ACS Nano, 2013, 7: 3388-3402 CrossRef PubMed Google Scholar

[34] Liu Z, Guo K, Zhao N, et al. Polysaccharides-based nanohybrids: Promising candidates for biomedical materials. Sci China Mater, 2019, 62: 1831-1836 CrossRef Google Scholar

[35] Yu Q, Zhang YM, Liu YH, et al. Magnetism and photo dual-controlled supramolecular assembly for suppression of tumor invasion and metastasis. Sci Adv, 2018, 4: eaat2297 CrossRef PubMed ADS Google Scholar

[36] Tu Y, Peng F, Sui X, et al. Self-propelled supramolecular nanomotors with temperature-responsive speed regulation. Nat Chem, 2017, 9: 480-486 CrossRef PubMed ADS Google Scholar

[37] Lin J, Xin P, An L, et al. Fe3O4-ZIF-8 assemblies as pH and glutathione responsive T2-T1 switching magnetic resonance imaging contrast agent for sensitive tumor imaging in vivo. Chem Commun, 2019, 55: 478-481 CrossRef PubMed Google Scholar

[38] Zhu N, Zhang B, Yu Q. Genetic engineering-facilitated coassembly of synthetic bacterial cells and magnetic nanoparticles for efficient heavy metal removal. ACS Appl Mater Interfaces, 2020, 12: 22948-22957 CrossRef PubMed Google Scholar

[39] Li F, Liang Z, Liu J, et al. Dynamically reversible iron oxide nanoparticle assemblies for targeted amplification of T1-weighted magnetic resonance imaging of tumors. Nano Lett, 2019, 19: 4213-4220 CrossRef PubMed ADS Google Scholar

[40] Lin W, Sun T, Xie Z, et al. A dual-responsive nanocapsule via disulfide-induced self-assembly for therapeutic agent delivery. Chem Sci, 2016, 7: 1846-1852 CrossRef PubMed Google Scholar

[41] Kuo CH, Huang MH. Morphologically controlled synthesis of Cu2O nanocrystals and their properties. Nano Today, 2010, 5: 106-116 CrossRef Google Scholar

[42] Xu Y, Liu P, Cao Y, et al. Room-temperature synthesis of Cu2O nanostructures and their morphology-dependent adsorption properties. Bull Korean Chem Soc, 2016, 37: 1114-1123 CrossRef Google Scholar

[43] Shareck J, Belhumeur P. Modulation of morphogenesis in Candida albicans by various small molecules. Eukaryot Cell, 2011, 10: 1004-1012 CrossRef PubMed Google Scholar

[44] Heilmann CJ, Sorgo AG, Siliakus AR, et al. Hyphal induction in the human fungal pathogen Candida albicans reveals a characteristic wall protein profile. Microbiology, 2011, 157: 2297-2307 CrossRef PubMed Google Scholar

[45] Yu Q, Zhang B, Ma F, et al. Novel mechanisms of surfactants against Candida albicans growth and morphogenesis. Chem-Biol Interact, 2015, 227: 1-6 CrossRef PubMed Google Scholar

[46] Yu Q, Ding X, Xu N, et al. In vitro activity of verapamil alone and in combination with fluconazole or tunicamycin against Candida albicans biofilms. Int J Antimicrob Agents, 2013, 41: 179-182 CrossRef PubMed Google Scholar

[47] Lee YJ, Kim S, Park SH, et al. Morphology-dependent antibacterial activities of Cu2O. Mater Lett, 2011, 65: 818-820 CrossRef Google Scholar

[48] Wilson D, Naglik JR, Hube B. The missing link between Candida albicans hyphal morphogenesis and host cell damage. PLoS Pathog, 2016, 12: e1005867 CrossRef PubMed Google Scholar

[49] Meng L, Zhao H, Zhao S, et al. Inhibition of yeast-to-hypha transition and virulence of Candida albicans by 2-alkylamino-quinoline derivatives. Antimicrob Agents Chemother, 2019, 63: e01891 CrossRef PubMed Google Scholar

[50] Finkel JS, Mitchell AP. Genetic control of Candida albicans biofilm development. Nat Rev Microbiol, 2011, 9: 109-118 CrossRef PubMed Google Scholar

[51] Wall G, Montelongo-Jauregui D, Vidal Bonifacio B, et al. Candida albicans biofilm growth and dispersal: Contributions to pathogenesis. Curr Opin Microbiol, 2019, 52: 1-6 CrossRef PubMed Google Scholar

[52] Nobile CJ, Nett JE, Andes DR, et al. Function of Candida albicans adhesin Hwp1 in biofilm formation. Eukaryot Cell, 2006, 5: 1604-1610 CrossRef PubMed Google Scholar

  • Figure 1

    Characterizations of the synthesized Cu2O NPs and Cu2O-PE-BSA. (a) TEM images. (b) Zeta potential. (c) Size distribution. (d) FT-IR spectra.

  • Scheme 1

    (a) Illustration of construction of Cu2O-PE-BSA assembly and (b) the assembly-mediated inhibition of growth and biofilm formation of the pathogenic fungal cells.

  • Figure 2

    Pathogenic fungus-induced disassembly of the Cu2O-PE-BSA microaggregates. Note that the Cu2O-PE-BSA alone had aggregation with microparticle morphology (top), while the presence of the pathogenic C. albicans cells strongly adsorbed the particles and led to their disassembly (middle). The group of C. albicans alone (bottom) exhibited no autofluorescence on the surface.

  • Figure 3

    Fungal death and inhibition of hyphal growth by Cu2O NPs and Cu2O-PE-BSA. (a) Fungal death induced by Cu2O NPs and Cu2O-PE-BSA at different concentrations. (b) Percent of hyphal cells under treatment of the agents. (c) Microscopy images of the fungal cells under the hypha-inducing condition with 40 mg L−1 of the agents. * indicates significant difference between the control group (0) and the treated group (P < 0.05).

  • Figure 4

    Inhibition of fungal biofilm formation by Cu2O NPs and Cu2O-PE-BSA. (a) Confocal images of biofilms at different layers from the bottom to the top (0–40 μm). (b) Biomass of the fungal biofilms evaluated by XTT assays. (c) Expression of HWP1 in the treated biofilms revealed by Hwp1-GFP fluorescence quantification. * indicates significant difference between the control group (0) and the nanoparticle-treated group (P < 0.05).

  • Figure 5

    Impairment of mammalian cell viability by Cu2O NPs and Cu2O-PE-BSA. (a) Viability of 293T cells. (b) Viability of RAW264.7 cells. * indicates significant difference between the two groups (P < 0.05).

  • Figure 6

    Wound healing and reduction of fungal cells in the mouse wounds by Cu2O NPs and Cu2O-PE-BSA. (a) Illustration of the mouse skin wound model. (b) Wound healing rate of the treated mouse wounds. (c) Fungal burden in the wounds. * indicates significant difference between the groups (P < 0.05).

  • Table 1   IC50 of Cu2O NPs and Cu2O-PE-BSA against C. albicans growth and biofilm formation


    IC50 against growth (mg L−1)

    IC50 against biofilm (mg L−1)

    Cu2O NPs


    Cu2O NPs



    < 20





    < 20


    < 20



    < 20


    < 20



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