Dose‑dependent effect of lysozyme upon Candida albicans biofilm

Sebaa,Sarra; Hizette,Nicolas; Boucherit‑Otmani,Zahia; Courtois,Philippe

doi:10.3892/mmr.2017.6148

Dose‑dependent effect of lysozyme upon Candida albicans biofilm

Authors:
- Sarra Sebaa
- Nicolas Hizette
- Zahia Boucherit‑Otmani
- Philippe Courtois
View Affiliations

Affiliations: Laboratory of Physiology and Pharmacology, Université Libre de Bruxelles, B‑1070 Brussels, Belgium, Laboratory of Antibiotics Antifungals: Physico‑Chemistry, Synthesis and Biological Activity, University of Aboubekr Belkaïd‑Tlemcen, Tlemcen 13000, Algeria
Published online on: January 25, 2017 https://doi.org/10.3892/mmr.2017.6148
Pages: 1135-1142
Copyright: © Sebaa et al. This is an open access article distributed under the terms of Creative Commons Attribution License.

Metrics: Total Views: 0 (Spandidos Publications: | PMC Statistics: )

Metrics: Total PDF Downloads: 0 (Spandidos Publications: | PMC Statistics: )

Cited By (CrossRef): 0 citations Loading Articles...

Abstract

The present study investigated the in vitro effect of lysozyme (0-1,000 µg/ml) on Candida albicans (C. albicans) biofilm development. Investigations were conducted on C. albicans ATCC 10231 and on 10 clinical isolates from dentures. Strains were cultured aerobically at 37˚C in Sabouraud broth. Yeast growth was evaluated by turbidimetry. Biofilm biomass was quantified on a polystyrene support by crystal violet staining and on acrylic surfaces by counts of colony forming units. Lysozyme affected biofilm formation to a greater extent than it affected growth. For the ATCC 10231 reference strain, lysozyme acted as a biofilm promotor on polystyrene at the highest concentration tested (1,000 µg/ml, non‑physiological). When the reference strain was investigated on acrylic resin support, lysozyme acted as a significant biofilm promotor on rough resin, but less on smooth resin. The attached biomass in the presence of physiological concentrations of lysozyme (10‑30 µg/ml) was significantly decreased compared with the hypothetical value of 100% using a one‑sample t‑test, but a comparison between the different lysozyme conditions using analysis of variance and post hoc tests did not reveal significant differences. In 10 wild strains, different patterns of biofilm formation on polystyrene were observed in the presence of lysozyme. Some strains, characterized by large amounts of biofilm formation in the presence of 1,000 µg/ml lysozyme, were poor biofilm producers at low concentrations of lysozyme. In contrast, some strains that were poor biofilm producers with a high lysozyme concentration were more inhibited by low concentrations of lysozyme. The present study emphasizes the need to develop strategies for biofilm control based on in vitro experiments, and to implement these in clinical trials prior to approval of hygiene products enriched with exocrine proteins, such as lysozyme. Further studies will extend these investigations to other Candida species, and to fungi and bacteria present in oral biofilms.

View Figures

View References

1	Radford DR, Challacombe SJ and Walter JD: Denture plaque and adherence of Candida albicans to denture-base materials in vivo and in vitro. Crit Rev Oral Biol Med. 10:99–116. 1999. View Article : Google Scholar : PubMed/NCBI
2	Courtois Ph: Candida biofilms on oral biomaterials. Biomaterials-Physics and chemistry. Pignatello R: Intech; Rjieka: pp. 475–490. 2011
3	Epstein JB, Truelove EL and Izutzu KT: Oral candidiasis: Pathogenesis and host defense. Rev Infect Dis. 6:96–106. 1984. View Article : Google Scholar : PubMed/NCBI
4	Webb BC, Thomas CJ, Willcox MD, Harty DW and Knox KW: Candida-associated denture stomatitis. Aetiology and management: A review. Part 1. Factors influencing distribution of Candida species in the oral cavity. Aust Dent J. 43:45–50. 1998. View Article : Google Scholar : PubMed/NCBI
5	Ramage G, Tomsett K, Wickes BL, López-Ribot JL and Redding SW: Denture stomatitis: A role for Candida biofilms. Oral Surg Oral Med Oral Pathol Oral Radiol Endod. 98:53–59. 2004. View Article : Google Scholar : PubMed/NCBI
6	Fleming A: On a remarkable bacteriolytic element found in tissues and secretions. Proc R Soc Lond B Biol Sci. 93:306–317. 1922. View Article : Google Scholar
7	Gordon S, Todd J and Cohn ZA: In vitro synthesis and secretion of lysozyme by mononuclear phagocytes. J Exp Med. 139:1228–1248. 1974. View Article : Google Scholar : PubMed/NCBI
8	Vukosavljevic D, Custodio W and Siqueira WL: Salivary proteins as predictors and controls for oral health. J Cell Commun Signal. 5:271–275. 2011. View Article : Google Scholar : PubMed/NCBI
9	Fábián TK, Hermann P, Beck A, Fejérdy P and Fábián G: Salivary defense proteins: Their network and role in innate and acquired oral immunity. Int J Mol Sci. 13:4295–4320. 2012. View Article : Google Scholar : PubMed/NCBI
10	Phillips DC: The three-dimensional structure of an enzyme molecule. Sci Am. 215:78–90. 1966. View Article : Google Scholar : PubMed/NCBI
11	Masschalck B and Michiels CW: Antimicrobial properties of lysozyme in relation to foodborne vegetative bacteria. Crit Rev Microbiol. 29:191–214. 2003. View Article : Google Scholar : PubMed/NCBI
12	MacKay BJ, Goodman H, Cox D, Grossbard BL, Iacono VJ and Pollock JJ: Development of an enzyme-linked immunosorbent assay for determination of lysozyme in human parotid and submandibular-sublingual salivas. J Clin Microbiol. 19:844–848. 1984.PubMed/NCBI
13	Noble RE: Salivary alpha-amylase and lysozyme levels: A non-invasive technique for measuring parotid vs submandibular/sublingual gland activity. J Oral Sci. 42:83–86. 2000. View Article : Google Scholar : PubMed/NCBI
14	Brandtzaeg P and Mann WV Jr: A comparative study of the lysozyme activity of human gingival pocket fluid, serum and saliva. Acta Odontol Scand. 22:441–455. 1964. View Article : Google Scholar : PubMed/NCBI
15	Eisenberg RJ, Bowers GM and Bergquist JJ: Lysozyme activity in gingival crevicular fluid. J Baltimore Coll Dent Surg. 32:83–85. 1977.PubMed/NCBI
16	Sakalauskiene J, Surna A, Ivanauskiene E, Zekonis G and Gleiznys A: Secretory function of neutrophilic leucocytes of the patients with periodontal diseases. Stomatologija. 7:90–94. 2005.PubMed/NCBI
17	Stuchell RN and Mandel ID: A comparative study of salivary lysozyme in caries-resistant and caries-susceptible adults. J Dent Res. 62:552–554. 1983. View Article : Google Scholar : PubMed/NCBI
18	Woods CM, Hooper DN, Ooi EH, Tan LW and Carney AS: Human lysozyme has fungicidal activity against nasal fungi. Am J Rhinol Allergy. 25:236–240. 2011. View Article : Google Scholar : PubMed/NCBI
19	Marquis G, Montplaisir S, Garzon S, Strykowski H and Auger P: Fungitoxicity of muramidase. Ultrastructural damage to Candida albicans. Lab Invest. 46:627–636. 1982.PubMed/NCBI
20	Marquis G, Garzon S, Strykowsky H and Auger P: Cell walls of normal and lysozyme-damaged blastoconidia of Candida albicans: Localization of surface factor 4 antigen and vicinal-glycol staining. Infect Immun. 59:1312–1318. 1991.PubMed/NCBI
21	Edgerton M and Koshlukova SE: Salivary histatin 5 and its similarities to the other antimicrobial proteins in human saliva. Adv Dent Res. 14:16–21. 2000. View Article : Google Scholar : PubMed/NCBI
22	Kamaya T: Flocculation phenomenon of Candida albicans by lysozyme. Mycopathol Mycol Appl. 37:320–330. 1969. View Article : Google Scholar : PubMed/NCBI
23	Tenovuo J: Clinical applications of antimicrobial host proteins lactoperoxidase, lysozyme and lactoferrin in xerostomia: Efficacy and safety. Oral Dis. 8:23–29. 2002. View Article : Google Scholar : PubMed/NCBI
24	Gil-Montoya JA, Guardia-López I and González-Moles MA: Evaluation of the clinical efficacy of a mouthwash and oral gel containing the antimicrobial proteins lactoperoxidase, lysozyme and lactoferrin in elderly patients with dry mouth: A pilot study. Gerodontology. 25:3–9. 2008. View Article : Google Scholar : PubMed/NCBI
25	Güneri P, Alpöz E, Epstein JB, Çankaya H and Ates M: In vitro antimicrobial effects of commercially available mouth-wetting agents. Spec Care Dentist. 31:123–128. 2011. View Article : Google Scholar : PubMed/NCBI
26	Lee JY, Kim YY, Chang JY, Park MS and Kho HS: The effects of peroxidase on the enzymatic and candidacidal activities of lysozyme. Arch Oral Biol. 55:607–612. 2010. View Article : Google Scholar : PubMed/NCBI
27	Cho MA, Kim YY, Chang JY and Kho HS: Interactions between hyaluronic acid, lysozyme, and the glucose oxidase-mediated lactoperoxidase system in enzymatic and candidacidal activities. Arch Oral Biol. 58:1349–1356. 2013. View Article : Google Scholar : PubMed/NCBI
28	Samaranayake YH, Samaranayake LP, Wu PC and So M: The antifungal effect of lactoferrine and lysozyme on Candida krusei and Candida albicans. APMIS. 105:875–883. 1997. View Article : Google Scholar : PubMed/NCBI
29	Kang JH, Kim YY, Chang JY and Kho HS: Influences of hyaluronic acid on the anticandidal activities of lysozyme and the peroxidase system. Oral Dis. 17:577–583. 2011. View Article : Google Scholar : PubMed/NCBI
30	Collins MS and Pappagianis D: Lysozyme-enhanced killing of Candida albicans and Coccidioides immitis by amphoteracin B. Sabouraudia. 12:329–340. 1974. View Article : Google Scholar : PubMed/NCBI
31	Nishiyama Y, Nakaoka C, Hiratani T, Abe S, Uchida K and Yamaguchi H: Synergy of lysozyme and lanoconazole on the morphology of Candida albicans. J Electron Microsc (Tokyo). 50:41–49. 2001. View Article : Google Scholar : PubMed/NCBI
32	Anil S and Samaranayake LP: Impact of lysozyme and lactoferrin on oral Candida isolates exposed to polyene antimycotics and fluconazole. Oral Dis. 8:199–206. 2002. View Article : Google Scholar : PubMed/NCBI
33	Samaranayake YH, Cheung BP, Parahitiyawa N, Seneviratne CJ, Yau JY, Yeung KW and Samaranayake LP: Synergistic activity of lysozyme and antifungal agents against Candida albicans biofilms on denture acrylic surfaces. Arch Oral Biol. 54:115–126. 2009. View Article : Google Scholar : PubMed/NCBI
34	Tanida T, Okamoto T, Okamoto A, Wang H, Hamada T, Ueta E and Ozaki T: Decreased excretion of antimicrobial proteins and peptides in saliva of patients with oral candidiasis. J Oral Pathol Med. 32:586–594. 2003. View Article : Google Scholar : PubMed/NCBI
35	Silva MP, Junior Chibebe J, Jorjão AL, Machado AK, Oliveira LD, Junqueira JC and Jorge AO: Influence of artificial saliva in biofilm formation of Candida albicans in vitro. Braz Oral Res. 26:24–28. 2012. View Article : Google Scholar : PubMed/NCBI
36	Stepanovic S, Vukovic D, Dakic I, Savic B and Svabic-Vlahovic M: A modified microtiter-plate test for quantification of staphylococcal biofilm formation. J Microbiol Methods. 40:175–179. 2000. View Article : Google Scholar : PubMed/NCBI
37	Iacono VJ, MacKay BJ, DiRienzo S and Pollock JJ: Selective antibacterial properties of lysozyme for oral microorganisms. Infect Immun. 29:623–632. 1980.PubMed/NCBI
38	Salton MR: Cell structure and the the enzymic lysis of bacteria. J Gen Microbiol. 9:512–523. 1953. View Article : Google Scholar : PubMed/NCBI
39	Edgerton M and Levine MJ: Characterization of acquired denture pellicle from healthy and stomatitis patients. J Prosthet Dent. 68:683–691. 1992. View Article : Google Scholar : PubMed/NCBI
40	Kamaya T: Lytic action of lysozyme on Candida albicans. Mycopathol Mycol Appl. 42:197–207. 1970. View Article : Google Scholar : PubMed/NCBI