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Capability of iturin from Bacillus subtilis to inhibit Candida albicans in vitro and in vivo

  • Shuzhen Lei
  • Haobin Zhao
  • Bing Pang
  • Rui Qu
  • Ziyang Lian
  • Chunmei Jiang
  • Dongyan Shao
  • Qingsheng Huang
  • Mingliang Jin
  • Junling ShiEmail author
Biotechnological products and process engineering
  • 32 Downloads

Abstract

Candida albicans is a fungal pathogen that is difficult to cure clinically. The current clinic C. albicans-inhibiting drugs are very harmful to humans. This study revealed the potential of iturin fractions from Bacillus subtilis to inhibit C. albicans in free status (MIC = 32 μg/mL) and natural biofilm in vitro. The inhibition mechanism was identified as an apoptosis pathway via the decrease of mitochondrial membrane potential, the increase of the reactive oxygen species (ROS) accumulation, and the induction of nuclear condensation. For in vivo experiments, the C. albicans infection model was constructed via intraperitoneal injection of 1 × 108C. albicans cells into mice. One day after the infection, iturin was used to treat infected mice at different concentrations alone and in combination with amphotericin B (AmB) by intraperitoneal injection. The treatment with AmB alone could cause the death of infected mice, whereas treatment with 15 mg/kg iturin per day alone led to the survival of all infected mice throughout the study. After continuously treated for 6 days, all mice were sacrificed and analyzed. As results, the combination of 15 mg/kg iturin and AmB at a ratio of 2:1 had the most efficient effect to remove the fungal burden in the kidney and cure the infected mice by reversing the symptoms caused by C. albicans infection, such as the loss of body weight, change of immunology cells in blood and cytokines in serum, and damage of organ structure and functions. Overall, iturin had potential in the development of efficient and safe drugs to cure C. albicans infection.

Keywords

Clinic fungal infection Lipopeptides Apoptosis Mouse model Amphotericin B 

Notes

Acknowledgments

The authors thank Northwestern Polytechnical University Analytica & Testing Center, Central Laboratory and Key Laboratory for Space Bioscience and Biotechnology, School of Life Sciences, Northwestern Polytechnical University for providing the instruments used in this study.

Funding information

This work was supported by the National Natural Science Foundation of China (grant no. 31701722), the Modern Agricultural Industry Technology System (CARS-30), the National Key R&D Program of China (2017YFE0105300), the Key Research and Development Plan of Shaanxi Province (2017ZDXL-NY-0304), the China Postdoctoral Science Foundation (No. 2017M620471), the Shaanxi Provincial Natural Science Foundation (No. 2018JQ3054), and the Postdoctoral Research Project of Shaanxi Province (No. 2017BSHEDZZ103).

Compliance with ethical standards

Conflict of interest

The authors declare that they have no conflict of interest.

Ethical statement

The experimentation, transportation, and care of the animals used in these experiments were performed in compliance with the relevant laws and institutional guidelines of Shaanxi Province, China. This study was approved by the Experimental Animal Care and Ethics Committees of Northwestern Polytechnical University, Shaanxi Province, China.

Supplementary material

253_2019_9805_MOESM1_ESM.pdf (432 kb)
ESM 1 (PDF 432 kb)

References

  1. Arikan S, Lozano-Chiu M, Paetznick V, Rex JH (2002) In vitro synergy of caspofungin and amphotericin B against Aspergillus and Fusarium spp. Antimicrob Agents Chemother 46:245–247.  https://doi.org/10.1128/AAC.46.1.245-247.2002 CrossRefGoogle Scholar
  2. Biniarz P, Baranowska G, Feder-kubis J, Krasowska A (2015) The lipopeptides pseudofactin II and surfactin effectively decrease Candida albicans adhesion and hydrophobicity. Antonie Van Leeuwenhoek 108:343–353.  https://doi.org/10.1007/s10482-015-0486-3 CrossRefGoogle Scholar
  3. Borghi E, Iatta R, Sciota R, Biassoni C, Cuna T, Montagna MT, Morace G (2010) Comparative evaluation of the vitek 2 yeast susceptibility test and CLSI broth microdilution reference method for testing antifungal susceptibility of invasive fungal isolates in Italy: the GISIA3 study. J Clin Microbiol 48:3153–3157.  https://doi.org/10.1128/JCM.00952-10 CrossRefGoogle Scholar
  4. Ceresa C, Rinaldi M, Chiono V, Carmagnola I, Allegrone G, Fracchia L (2016) Lipopeptides from Bacillus subtilis AC7 inhibit adhesion and biofilm formation of Candida albicans on silicone. Antonie Van Leeuwenhoek 109:1375–1388.  https://doi.org/10.1007/s10482-016-0736-z CrossRefGoogle Scholar
  5. Dey G, Bharti R, Dhanarajan G, Das S, Dey KK, Kumar BNP, Sen R, Mandal M (2015) Marine lipopeptide Iturin A inhibits Akt mediated GSK3 β and FoxO3a signaling and triggers apoptosis in breast cancer. Sci Reports 5:10316.  https://doi.org/10.1038/srep10316 CrossRefGoogle Scholar
  6. Elefanti A, Mouton JW, Verweij PE, Tsakris A, Zerva L, Meletiadis J (2013) Amphotericin B- and voriconazole-echinocandin combinations against Aspergillus spp.: effect of serum on inhibitory and fungicidal interactions. Antimicrob Agents Chemother 57:4656–4663.  https://doi.org/10.1128/AAC.00597-13 CrossRefGoogle Scholar
  7. Gaofu Q, Fayin Z, Peng D, Xiufen Y, Dewen Q, Ziniu Y, Jingyuan C, Xiuyun Z (2010) Lipopeptide induces apoptosis in fungal cells by a mitochondria-dependent pathway. Peptides 31:1978–1986.  https://doi.org/10.1016/j.peptides.2010.08.003 CrossRefGoogle Scholar
  8. Grau A, Ortiz A, De Godos A, Gómez-Fernández JC (2000) A biophysical study of the interaction of the lipopeptide antibiotic iturin A with aqueous phospholipid bilayers. Arch Biochem Biophys 377:315–323.  https://doi.org/10.1006/abbi.2000.1791 CrossRefGoogle Scholar
  9. Harris MR, Coote PJ (2010) Combination of caspofungin or anidulafungin with antimicrobial peptides results in potent synergistic killing of Candida albicans and Candida glabrata in vitro. Int J Antimicrob Agents 35:347–356.  https://doi.org/10.1016/j.ijantimicag.2009.11.021 CrossRefGoogle Scholar
  10. Hong SY, Oh JE, Lee KH (1999) In vitro antifungal activity and cytotoxicity of a novel membrane-active peptide. Antimicrob Agents Chemother 43:1704–1707.  https://doi.org/10.1016/S0584-8547(02)00057-5 CrossRefGoogle Scholar
  11. Kim P, Ryu J, Kim YH, Chi YT (2010) Production of biosurfactant lipopeptides iturin A, fengycin, and surfactin A from Bacillus subtilis CMB32 for control of Colletotrichum gloeosporioides. J Microbiol Biotechnol 20:138–145.  https://doi.org/10.4014/jmb.0905.05007 Google Scholar
  12. Kovács R, Gesztelyi R, Berényi R, Domán M, Kardos G, Juhász B, Majoros L (2014) Killing rates exerted by caspofungin in 50% serum and its correlation with in vivo efficacy in a neutropenic murine model against Candida krusei and Candida inconspicua. J Med Microbiol 63:186–194.  https://doi.org/10.1099/jmm.0.066381-0 CrossRefGoogle Scholar
  13. Kriengkauykiat J, Ito JI, Dadwal SS (2011) Epidemiology and treatment approaches in management of invasive fungal infections. Clin Epidemiol 3:175–191.  https://doi.org/10.2147/CLEP.S12502 Google Scholar
  14. Kullberg BJ, Verweij PE, Akova M, Arendrup MC, Bille J, Calandra T, Cuenca-Estrella M, Herbrecht R, Jacobs F, Kalin M, Kibbler CC, Lortholary O, Martino P, Meis JF, Muñoz P, Odds FC, De Pauw BE, Rex JH, Roilides E, Rogers TR, Ruhnke M, Ullmann AJ, Uzun Ö, Vandewoude K, Vincent JL, Donnelly JP (2011) European expert opinion on the management of invasive candidiasis in adults. Clin Microbiol Infect 17:1–12.  https://doi.org/10.1111/j.1469-0691.2011.03615.x CrossRefGoogle Scholar
  15. Kumar SN, Nambisan B, Mohandas C, Sundaresan A (2013) In vitro synergistic activity of diketopiperazines alone and in combination with amphotericin B or clotrimazole against Candida albicans. Folia Microbiol (Praha) 58:475–482.  https://doi.org/10.1007/s12223-013-0234-x CrossRefGoogle Scholar
  16. Leunk RD, Moon RJ (1979) Physiological and metabolic alterations accompanying systemic candidiasis in mice. Infect Immun 26:1035–1041.  https://doi.org/10.1007/BF01646257 Google Scholar
  17. Lin PY, Tsai CT, Chuang WL, Chao YH, Pan IH, Chen YK, Lin CC, Wang BY (2017) Chlorella sorokiniana induces mitochondrial-mediated apoptosis in human non-small cell lung cancer cells and inhibits xenograft tumor growth in vivo. BMC Complement Altern Med 17:88.  https://doi.org/10.1186/s12906-017-1611-9 CrossRefGoogle Scholar
  18. Linares CEB, Giacomelli SR, Altenhofen D, Alves SH, Morsch VM, Schetinger MRC (2013) Fluconazole and amphotericin-B resistance are associated with increased catalase and superoxide dismutase activity in Candida albicans and Candida dubliniensis. Rev Soc Bras Med Trop 46:752–758.  https://doi.org/10.1590/0037-8682-0190-2013 CrossRefGoogle Scholar
  19. Liu S, Hou Y, Chen X, Gao Y, Li H, Sun S (2014) Combination of fluconazole with non-antifungal agents: a promising approach to cope with resistant Candida albicans infections and insight into new antifungal agent discovery. Int J Antimicrob Agents 43:395–402.  https://doi.org/10.1016/j.ijantimicag.2013.12.009 CrossRefGoogle Scholar
  20. Lu M, Li T, Wan J, Li X, Yuan L, Sun S (2017) Antifungal effects of phytocompounds on Candida species alone and in combination with fluconazole. Int J Antimicrob Agents 49:125–136.  https://doi.org/10.1016/j.ijantimicag.2016.10.021 CrossRefGoogle Scholar
  21. Lu M, Yu C, Cui X, Shi J, Yuan L, Sun S (2018) Gentamicin synergises with azoles against drug-resistant Candida albicans. Int J Antimicrob Agents 51:107–114.  https://doi.org/10.1016/j.ijantimicag.2017.09.012 CrossRefGoogle Scholar
  22. Lv Y, Wang J, Gao H, Wang Z, Dong N, Ma Q, Shan A (2014) Antimicrobial properties and membrane-active mechanism of a potential α-helical antimicrobial derived from cathelicidin PMAP-36. PLoS One 9:e86364.  https://doi.org/10.1371/journal.pone.0086364 CrossRefGoogle Scholar
  23. Maget-Dana R, Peypoux F (1994) Iturins , a special class of pore-forming lipopeptides: biological and physicochemical properties. Toxicology 87:151–174.  https://doi.org/10.1016/0300-483x(94)90159-7 CrossRefGoogle Scholar
  24. Meena KR, Kanwar SS (2015) Lipopeptides as the antifungal and antibacterial agents: applications in food safety and therapeutics. Biomed Res Int 2015:1–9.  https://doi.org/10.1155/2015/473050 CrossRefGoogle Scholar
  25. Nasir MN, Besson F (2011) Specific interactions of mycosubtilin with cholesterol-containing artificial membranes. Langmuir 27:10785–10792.  https://doi.org/10.1021/la200767e CrossRefGoogle Scholar
  26. Nasir MN, Besson F (2012a) Conformational analyses of bacillomycin D, a natural antimicrobial lipopeptide, alone or in interaction with lipid monolayers at the air-water interface. J Colloid Interface Sci 387:187–193.  https://doi.org/10.1016/j.jcis.2012.07.091 CrossRefGoogle Scholar
  27. Nasir MN, Besson F (2012b) Interactions of the antifungal mycosubtilin with ergosterol-containing interfacial monolayers. Biochim Biophys Acta 1818:1302–1308.  https://doi.org/10.1016/j.bbamem.2012.01.020 CrossRefGoogle Scholar
  28. Nolte FS, Parkinson T, Falconer DJ, Dix S, Williams J, Gilmore C, Geller R, Wingard JR (1997) Isolation and characterization of fluconazole- and amphotericin B- resistant Candida albicans from blood of two patients with leukemia. Antimicrob Agents Chemother 41:196–199.  https://doi.org/10.1016/j.msea.2006.12.193 CrossRefGoogle Scholar
  29. Onishi J, Meinz M, Thompson J, Curotto J, Dreikorn S, Rosenbach M, Douglas C, Abruzzo G, Flattery A, Kong L, Cabello A, Vicente F, Pelaez F, Diez MT, Martin I, Bills G, Giacobbe R, Dombrowski A, Schwartz R, Morris S, Harris G, Tsipouras A, Wilson K, Kurtz MB (2000) Discovery of novel antifungal (1,3)-β-D-glucan synthase inhibitors. Antimicrob Agents Chemother 44:368–377.  https://doi.org/10.1128/AAC.44.2.368-377.2000 CrossRefGoogle Scholar
  30. Pathak KV, Keharia H (2014) Identification of surfactins and iturins produced by potent fungal antagonist, Bacillus subtilis K1 isolated from aerial roots of banyan (Ficus benghalensis) tree using mass spectrometry. Biotech 4(3):283–295.  https://doi.org/10.1007/s13205-013-0151-3 Google Scholar
  31. Ramage G, López-ribot JL (2005) Techniques for antifungal susceptibility testing of Candida albicans biofilms. Methods Mol Med 118:71–79.  https://doi.org/10.1385/1-59259-943-5:071 Google Scholar
  32. Rautela R, Singh AK, Shukla A, Cameotra SS (2014) Lipopeptides from Bacillus strain AR2 inhibits biofilm formation by Candida albicans. Antonie Van Leeuwenhoek 105:809–821.  https://doi.org/10.1007/s10482-014-0135-2 CrossRefGoogle Scholar
  33. Ruhnke M, Rickerts V, Cornely OA, Buchheidt D, Glöckner A, Heinz W, Höhl R, Horré R, Karthaus M, Kujath P, Willinger B, Presterl E, Rath P, Ritter J, Glasmacher A, Lass-Flörl C, Groll AH (2011) Diagnosis and therapy of Candida infections: joint recommendations of the German Speaking Mycological Society and the Paul-Ehrlich-Society for Chemotherapy. Mycoses 54:279–310.  https://doi.org/10.1111/j.1439-0507.2011.02040.x CrossRefGoogle Scholar
  34. Ryder NS, Leitner I (2001) Synergistic interaction of terbinafine with triazoles or amphotericin B against Aspergillus species. Med Mycol 39:91–95.  https://doi.org/10.1080/714030977 CrossRefGoogle Scholar
  35. Sanglard D, Odds FC (2002) Resistance of Candida species to antifungal agents : molecular mechanisms and clinical consequences. Lancet Infect Dis 2:73–85.  https://doi.org/10.1016/S1473-3099(02)00181-0 CrossRefGoogle Scholar
  36. Sarkar S, Rajput S, Tripathi AK, Mandal M (2013) Targeted therapy against EGFR and VEGFR using ZD6474 enhances the therapeutic potential of UV-B phototherapy in breast cancer cells. Mol Cancer 12:122.  https://doi.org/10.1186/1476-4598-12-122 CrossRefGoogle Scholar
  37. Shin S, Pyun MS (2004) Anti-Candida effects of estragole in combination with ketoconazole or amphotericin B. Phyther Res 18:827–830.  https://doi.org/10.1002/ptr.1573 CrossRefGoogle Scholar
  38. Tabbene O, Di Grazia A, Azaiez S, Ben Slimene I, Elkahoui S, Alfeddy MN a, Casciaro B, Luca V, Limam F, uis MML (2015) Synergistic fungicidal activity of the lipopeptide Bacillomycin D with Amphotericin B against pathogenic Candida species. FEMS Yeast Res 15:fov022.  https://doi.org/10.1093/femsyr/fov022 CrossRefGoogle Scholar
  39. Tang Q, Bie X, Lu Z, Lv F, Tao Y, Qu X (2014) Effects of fengycin from Bacillus subtilis fmbJ on apoptosis and necrosis in Rhizopus stolonifer. J Microbiol 52:675–680.  https://doi.org/10.1007/s12275-014-3605-3 CrossRefGoogle Scholar
  40. van de Veerdonk FL, Netea MG, Joosten LA, van der Meer JWM, Kullberg BJ (2010) Novel strategies for the preventionand treatment of Candida infections: the potential of immunotherapy. FEMS Microbiol Rev 34:1063–1075.  https://doi.org/10.1111/j.1574-6976.2010.00232.x CrossRefGoogle Scholar
  41. Wisplinghoff H, Bischoff T, Tallent SM, Seifert H, Wenzel RP, Edmond MB (2004) Nosocomial bloodstream infections in US hospitals: analysis of 24,179 cases from a prospective nationwide surveillance study. Clin Infect Dis 39:309–317.  https://doi.org/10.1086/421946 CrossRefGoogle Scholar
  42. Yuan D, Wan JZ, Deng LL, Zhang CC, Dun YY, Dai YW, Zhou ZY, Liu CQ, Wang T (2014) Chikusetsu saponin V attenuates MPP+-induced neurotoxicity in SH-SY5Y cells via regulation of Sirt1/Mn-SOD and GRP78/Caspase-12 pathways. Int J Mol Sci 15:13209–13222.  https://doi.org/10.3390/ijms150813209 CrossRefGoogle Scholar
  43. Zhang B, Dong C, Shang Q, Han Y, Li P (2013) New insights into membrane-active action in plasma membrane of fungal hyphae by the lipopeptide antibiotic bacillomycin L. Biochim Biophys Acta 1828:2230–2237.  https://doi.org/10.1016/j.bbamem.2013.05.033 CrossRefGoogle Scholar
  44. Zhang X, Jacob MR, Rao RR, Wang Y-H, Agarwal AK, Newman DJ, Khan IA, Clark AM, Li X (2012) Antifungal cyclic peptides from the marine sponge Microscleroderma herdmani. Res Rep Med Chem 2:7–14.  https://doi.org/10.2147/RRMC.S30895 Google Scholar
  45. Zhao H, Li J, Zhang Y, Lei S, Zhao X, Shao D, Jiang C, Shi J, Sun H (2018a) Potential of iturins as functional agents: safe, probiotic, and cytotoxic to cancer cells. Food Funct 9:5580–5587.  https://doi.org/10.1039/c8fo01523f CrossRefGoogle Scholar
  46. Zhao H, Shao D, Jiang C, Shi J, Li Q, Huang Q, Rajoka MSR, Yang H, Jin M (2017) Biological activity of lipopeptides from Bacillus. Appl Microbiol Biotechnol 101:5951–5960.  https://doi.org/10.1007/s00253-017-8396-0 CrossRefGoogle Scholar
  47. Zhao H, Xu X, Lei S, Shao D, Jiang C, Shi J, Zhang Y, Liu L, Lei S, Sun H, Huang Q (2019) Iturin A-like lipopeptides from Bacillus subtilis trigger apoptosis, paraptosis, and autophagy in Caco-2 cells. J Cell Physiol 234:6414–6427.  https://doi.org/10.1002/jcp.27377 CrossRefGoogle Scholar
  48. Zhao H, Yan L, Xu X, Jiang C, Shi J, Zhang Y, Liu L, Lei S (2018b) Potential of Bacillus subtilis lipopeptides in anti - cancer I: induction of apoptosis and paraptosis and inhibition of autophagy in K562 cells. AMB Express 8:78.  https://doi.org/10.1186/s13568-018-0606-3 CrossRefGoogle Scholar

Copyright information

© Springer-Verlag GmbH Germany, part of Springer Nature 2019

Authors and Affiliations

  1. 1.Key Laboratory for Space Bioscience and Space Biotechnology, School of Life SciencesNorthwestern Polytechnical UniversityXi’anChina

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