Mikrobiol. Z. 2021; 83(1):58-67.
Antibiofilm Effect of Adamantane Derivative against Staphylococcus aureus
N.I. Hrynchuk1, N.O. Vrynchanu1, T.A. Buchtyarova1, D.M. Dudikova1,
Yu.V. Korotkyi2, L.B. Bondarenko1
1Institute of Pharmacology and Toxicology, NAMS of Ukraine
14 Anton Tsedik Str., Kуiv, 03057, Ukraine
2Institute of Organic Chemistry, NAS of Ukraine
5 Murmanska Str., Kуiv, 02094, Ukraine
Currently, one of the most urgent problems in clinical practice is the antibiotic therapy ineffectiveness at chronic diseases treatment caused by biofilms-forming microorganisms. One of the ways to its solution is the search for new compounds with antibiofilm activity which can prevent the adhesion of microorganisms, disrupt the structure of the biofilm matrix and affect the Quorum sensing system. The aim of the study was to investigate adamantane derivative 1-[4-(1-adamantyl) phenoxy]-3-(N-benzyl,N-dimethylamino)-2-propanol chloride (KVM-97) antimicrobial activity mechanism against Staphylococcus aureus biofilms. Methods. The ability of the adamantane derivative KVM-97 to prevent S. aureus biofilm formation and to destroy previously formed biofilms has been tested on polystyrene plates by gentian violet sorption on these structures, followed by desorption with organic solvent and use of resazurin (redox indicator). The S. aureus cells viability in mature biofilms was evaluated with specific dyes for living (acridine orange) and dead (propidium iodide) cells. Lowry method was used to assess the effect of KVM-97 on the matrix components for the total protein contents determination, the polysaccharides were detected spectrophotometrically (using phenol and sulfuric acid), Bap-protein – by test with Congo red. Persisters’ subpopulation was detected by activation of the SOS response in bacteria when exposed to high concentrations of antimicrobial substances. Results. It was found that KVM-97 (the compound with the adamantyl radical) showed an antibiofilm effect against S. aureus, decreasing biofilm biomass: at the biofilm formation stage – by 22.5% and 75.0%, while in case of 2-day biofilms treatment – by 34.5% and 32.4% at 0.5 MIC and 5.0 MIC respectively. Compound KVM-97 was able to reduce the number of metabolically active S. aureus cells only at the stage of biofilm formation (reduction by 92.7 and 95.8% at 2.0 and 5.0 MIC). Obtained results indicated that this adamantane-containing compound did not affect the protein and polysaccharides contents of S. aureus biofilms matrix. The changes of Bap-protein level caused by KVM-97 were not statistically significant (p>0.05). It was shown that KVM-97 did not prevent the formation of metabolically inactive persister cells; their share was 0.71% of the control. Conclusions. Thus, adamantane-containing compound KVM-97 is able to prevent S. aureus biofilm formation, causing significant biofilms’ mass reduction, as well as lowering the viable cells number in them and destroying already formed biofilms. Its antibiofilm effects are not associated with matrix protein and polysaccharides synthesis impairments. Further thorough investigations are needed to establish the effect of this compound on eDNA synthesis, the Quorum sensing system, and the ica and arg genes expression of S. aureus responsible for biofilm formation.
Keywords: biofilms, Staphylococcus aureus, adamantane derivative, matrix, persisters.
Full text (PDF, in English)
- Kırmusaoğlu S. Staphylococcal biofilms: pathogenicity, mechanism and regulation of biofilm formation by quorum sensing system and antibiotic resistance mechanisms of biofilm embedded microorganisms. Microbial Biofilms-Importance and Applications. Dhanasekaran D, Thajuddin N, editors. Croatia: Intech; 2016. p. 189–209. https://doi.org/10.5772/62943
- Wolska KI, Grudniak AM, Rudnicka Z, Markowska K. Genetic control of bacterial biofilms. J Appl Genet. 2016; 57(2):225–38. https://doi.org/10.1007/s13353-015-0309-2
- Otto M. Staphylococcal Biofilms. In: Fischetti V, Novick R, Ferretti J, Portnoy D, Braunstein M, Rood J, editors. Gram-Positive Pathogens. Third ed. Washington: ASM Press; 2019.
- Jamal M, Ahmad W, Andleeb S, Jalil F, Imran M, Nawaz MA, et al. Bacterial biofilm and associated infections. J Chin Med Assoc. 2018; 81(1):7–11. https://doi.org/10.1016/j.jcma.2017.07.012
- Hlushanova NA, Blynov AY, Alekseeva NB. [Bakterialnye bioplenki v infekcionnoj patologii cheloveka]. Medicina v Kuzbasse. 2015; 2:30–5. Russian.
- Chernyavskij VI. [Bakterialnye bioplenki i infekcii (lekciya)]. Annals of Mechnikov Institute. 2013; 1:86–90. Russian.
- Chebotar IV, Mayansky AN, Konchakova ED, Lazareva AV, Chistyakova VP. [Antibiotikorezistentnost bioplyonochnyh bakterij]. Klin Mikrobiol Antimikrob Himioter. 2012; 14(1):51–8. Russian.
- Arciola CR, Campoccia D, Ravaioli S, Montanaro L. Polysaccharide intercellular adhesin in biofilm: structural and regulatory aspects. Front Cell Infect Microbiol. 2015; 5(7):1–10. https://doi.org/10.3389/fcimb.2015.00007
- Speziale P, Pietrocola G, Foster TJ, Geoghegan JA. Protein-based biofilm matrices in Staphylococci. Front Cell Infect Microbiol. 2014; 4:171. https://doi.org/10.3389/fcimb.2014.00171
- Taglialegna A, Navarro S, Ventura S, Garnett JA, Matthews S, Penades JR. Staphylococcal Bap proteins build amyloid scaffold biofilm matrices in response to environmental signals. PLoS Pathog. 2016; 12(6):e1005711. https://doi.org/10.1371/journal.ppat.1005711
- Vrynchanu NO. [Antibacterial action of 4-(adamantyl-1)-1-(1-aminobutyl)benzene]. Mikrobiol Z. 2014; 5:42–8. Ukrainian.
- Dudikova DM, Vrynchanu NO, Korotkyi YuV, Dronova ML, Suvorova ZS, Sharova AO, et al. [Antybakterialna aktyvnist aminopropanoliv z adamantylnym i N-alkilarylnym radykalom vidnosno bioplivok E. coli]. Farmakolohiia ta likarska toksykolohiia. 2017; 6:37–42. Ukrainian.
- Sharova AO, Hrynchuk NI, Dudikova DM, Nedashkivska VV, Suvorova ZS, Vrynchanu NO. [Antybakterialna aktyvnist 4-(1-adamantyl)-(1-aminobutyl)benzolu vidnosno bioplivok S. aureus]. Farmakolohiia ta likarska toksykolohiia. 2018; 2 (58):79–85. Ukrainian.
- O’Toole GA. Microtiter dish biofilm formation assay. J Vis Exp. 2011; 47:e2437. https://doi.org/10.3791/2437
- Tote K, Berghe DV, Maes L, Cos P. A new colorimetric microtitre model for the detection of Staphylococcus aureus biofilms. Lett Appl Microbiol. 2008; 46(2):249–54. https://doi.org/10.1111/j.1472-765X.2007.02298.x
- Sandberg ME, Schellmann D, Brunhofer G, Erker T, Busygin I, Leino R, et al. Pros and cons of using resazurin staining for quantification of viable Staphylococcus aureus biofilms in a screening assay. J Microbiol Methods. 2009; 78(1):104–6. https://doi.org/10.1016/j.mimet.2009.04.014
- Chiba A, Sugimoto S, Sato F, Hori S, Mizunoe Y. A refined technique for extraction of extracellular matrices from bacterial biofilms and its applicability. Microb Biotechnol. 2015; 8(3):392–403. https://doi.org/10.1111/1751-7915.12155
- Lowry OH, Rosebrough, NJ, Farr AL, Randall RJ. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951; 193(1):265–75. https://doi.org/10.1016/S0021-9258(19)52451-6
- Dubois M, Gilles KA, Hamilton JK, Rebers PT, Smith F. Colorimetric method for determination of sugars and related substances. Anal Chem. 1956; 28(3):350–6. https://doi.org/10.1021/ac60111a017
- Qadri F, Hossain SA, Ciznár I, Haider K, Ljungh, A., Wadstrom, T, et al. Congo red binding and salt aggregation as indicators of virulence in Shigella species. J Clin Microbiol. 1988; 26(7):1343–8. https://doi.org/10.1128/JCM.26.7.1343-1348.1988
- Taglialegna A, Lasa I, Valle J. Amyloid structures as biofilm matrix scaffolds. J Bacteriol. 2016; 198(19):2579–88. https://doi.org/10.1128/JB.00122-16
- Marques CNH. Isolation of persister cells from biofilm and planktonic populations of Pseudomonas aeruginosa. Bio-protocol. 2015; 5(18):e1590. https://doi.org/10.21769/BioProtoc.1590
- Chen CY, Nace GW, Irwin PL. A 6×6 drop plate method for simultaneous colony counting and MPN enumeration of Campylobacter jejuni, Listeria monocytogenes, and Escherichia coli. J Microbiol Methods. 2003; 55(2):475–9. https://doi.org/10.1016/S0167-7012(03)00194-5
- Mardanova AM, Kabanov DA, Rudakova NL, Sharipova MR. [BIOPLENKI: Osnovnye principy organizacii i metody issledovaniya]. Kazan: K(P)FU. 2016:42. Russian.
- Kayumov AR, Nureeva AA, Trizna EY, Gazizova GR, Bogachev MI, Shtyrlin NV, et al. New derivatives of pyridoxine exhibit high antibacterial activity against biofilm-embedded Staphylococcus cells. Biomed Res Int. 2015:890968. https://doi.org/10.1155/2015/890968
- Bogachev MI, Volkov VYu, Markelov OA, Trizna EYu, Baydamshina DR, Melnikov V, et al. Fast and simple tool for the quantification of biofilm-embedded cells sub-populations from fluorescent microscopic images. PLoS One. 2018; 13(5):e0193267. https://doi.org/10.1371/journal.pone.0193267
- Lapach SN, Chubenko AV, Babich PN. [Statisticheskie metody v mediko-biologicheskih issledovaniyah s ispolzovaniem Excel]. Kyiv: Morion; 2001. Russian.
- Fisenko VP, i dr. [Rukovodstvo po eksperimentalnomu (doklinicheskomu) izucheniyu novyh farmakologicheskih veshestv]. Moscow: Remedium; 2000. Russian.
- Zheng JX, Tu HP, Sun X, Xu GJ, Chen JW, Deng QW, et al. In vitro activities of telithromycin against Staphylococcus aureus biofilms compared with azithromycin, clindamycin, vancomycin and daptomycin. J Med Microbiol. 2020; 69(1):120–31. https://doi.org/10.1099/jmm.0.001122
- Tan F, She P, Zhou L, Liu Y, Chen L, Luo Z, et al. Bactericidal and anti-biofilm activity of the retinoid compound CD437 against Enterococcus faecalis. Front Microbiol. 2019; 10:230–1. https://doi.org/10.3389/fmicb.2019.02301
- Song T, Duperthuy M, Wai SN, Sub-optimal treatment of bacterial biofilms. Antibiotics (Basel). 2016; 5(2):23. https://doi.org/10.3390/antibiotics5020023
- Das MC, Das A, Samaddar S, Daware A, Ghosh C, Acharjee S, et al. Vitexin alters Staphylococcus aureus surface hydrophobicity to interfere with bioﬁlm formation. BioRxiv. 2018. https://doi.org/10.1101/301473
- Grassi L, Di Luca M, Maisetta G, Rinaldi AC, Esin S, Trampuz A, et al. Generation of persister cells of Pseudomonas aeruginosa and Staphylococcus aureus by chemical treatment and evaluation of their susceptibility to membrane-targeting agents. Front Microbiol. 2017; 8:19–17. https://doi.org/10.3389/fmicb.2017.01917