BACKGROUND: Pseudomonas aeruginosa can proliferate in immunocompromised individuals, forming biofilms that increase antibiotic resistance. This bacterium poses a significant global health risk due to its resistance to human defenses, antibiotics, and various environmental stresses. The objective of this study was to evaluate the antibacterial, anti-biofilm, and anti-quorum sensing activities of galloylquinic acid compounds (GQAs) extracted from Copaifera lucens leaves against clinical isolates of multidrug-resistant (MDR) P. aeruginosa. We have investigated the optimal concentration of GQAs needed to eradicate preexisting biofilms and manage wound infections caused by P. aeruginosa, in vitro and in vivo. RESULTS: Our results revealed that GQAs exhibited 25-40 mm inhibition zone diameters, with 1-4 µg/mL MIC and 2-16 µg/mL MBC values. GQAs interfered with the planktonic mode of P. aeruginosa isolates, and significantly inhibited their growth in the pre-formed biofilm architecture, with MBIC80 and MBEC80 values of 64 µg/mL and 128 µg/mL, respectively. The anti-biofilm effect was confirmed by fluorescence staining and confocal microscopy which showed a dramatic reduction in the cell viability and the biofilm thickness (62.5%), after exposure to 128 µg/mL of GQAs in particular. The scanning electron micrographs showed that GQAs impaired biofilm and bacterial structures by interfering with the biomass and the exopolysaccharides forming the matrix. GQAs also interfered with virulence factors and bacterial motility, where 128 µg/mL of GQAs significantly (p < 0.05) reduced rhamnolipid, pyocyanin, and the swarming motility of the organism which play a vital role in the biofilm formation. GQAs downregulated 89% of the quorum-sensing genes (lasI and lasR, pqsA and pqsR) involved in the biofilm formation. CONCLUSION: GQAs demonstrate significant promise as novel and potent antibiofilm and antivirulence agents against clinical isolates of MDR P. aeruginosa, with substantial potential to enhance wound healing in biofilm-associated infections. This promising antibacterial action positions GQAs as a superior alternative for the treatment of biofilm-associated wound infections, with substantial potential to improve wound healing and mitigate the impact of persistent bacterial infections. CLINICAL TRIAL NUMBER: not applicable.

Department of Experimental Biology Faculty of Science Masaryk University Brno 62500 Czech Republic

Department of Microbiology and Immunology Faculty of Pharmacy Tanta University Tanta Egypt

Department of Pathology Faculty of Medicine Tanta University Tanta Egypt

Department of Pharmaceutical Sciences School of Pharmaceutical Sciences of Ribeirão Preto University of São Paulo SP 14040 903 Brazil

Faculty of Pharmacy Department of Microbiology and Immunology Delta University for Science and Technology International Coastal Road Gamasa 11152 Egypt

Faculty of Pharmacy Department of Pharmacognosy Delta University for Science and Technology International Coastal Road Gamasa 11152 Egypt

School of Pharmacy and Biomolecular Sciences Royal College of Surgeons in Ireland Dublin D02 VN51 Ireland

Zobrazit více v PubMed

van Duin D, Paterson DL. Multidrug-resistant Bacteria in the community: an update. Infect Dis Clin North Am. 2020;34(4):709–22. PubMed PMC

Donlan RM, Costerton JW. Biofilms: survival mechanisms of clinically relevant microorganisms. Clin Microbiol Rev. 2002;15(2):167–93. PubMed PMC

Oluwole OM. BIOFILM: FORMATION AND NATURAL PRODUCTS’ APPROACH TO CONTROL - A REVIEW. Afr J Infect Dis. 2022;16(2 Suppl):59–71. PubMed PMC

Sharma D, Misba L, Khan AU. Antibiotics versus biofilm: an emerging battleground in microbial communities. Antimicrob Resist Infect Control. 2019;8:76. PubMed PMC

Vestby LK et al. Bacterial biofilm and its role in the pathogenesis of Disease. Antibiot (Basel), 2020. 9(2). PubMed PMC

Ciofu O, Tolker-Nielsen T. Tolerance and resistance of Pseudomonas aeruginosa Biofilms to Antimicrobial agents-How P. Aeruginosa Can escape antibiotics. Front Microbiol. 2019;10:913. PubMed PMC

Jeong G-J, et al. Natural and synthetic molecules with potential to enhance biofilm formation and virulence properties in Pseudomonas aeruginosa. Crit Rev Microbiol. 2024;50(5):830–58. PubMed

Ahmed S, et al. Natural quorum sensing inhibitors effectively downregulate gene expression of Pseudomonas aeruginosa virulence factors. Appl Microbiol Biotechnol. 2019;103(8):3521–35. PubMed PMC

Hall CW, Mah TF. Molecular mechanisms of biofilm-based antibiotic resistance and tolerance in pathogenic bacteria. FEMS Microbiol Rev. 2017;41(3):276–301. PubMed

Hengzhuang W, et al. In vivo pharmacokinetics/pharmacodynamics of colistin and imipenem in Pseudomonas aeruginosa biofilm infection. Antimicrob Agents Chemother. 2012;56(5):2683–90. PubMed PMC

Mishra R et al. Natural anti-biofilm agents: strategies to Control Biofilm-forming pathogens. Front Microbiol. 2020;11. PubMed PMC

Fernández-Barat L, et al. Phenotypic shift in Pseudomonas aeruginosa populations from cystic fibrosis lungs after 2-week antipseudomonal treatment. J Cyst Fibros. 2017;16(2):222–9. PubMed

Guedes BN, et al. Natural antibiotics against antimicrobial resistance: sources and bioinspired delivery systems. Brazilian J Microbiol. 2024;55(3):2753–66. PubMed PMC

Guimarães AL, et al. Antimicrobial activity of Copaiba (Copaifera officinalis) and Pracaxi (Pentaclethra macroloba) oils against Staphylococcus Aureus: importance in compounding for Wound Care. Int J Pharm Compd. 2016;20(1):58–62. PubMed

Alves JA, et al. Investigation of Copaifera Genus as a new source of antimycobaterial agents. Future Sci OA. 2020;6(7):Fso587. PubMed PMC

Abd El-Salam MA, et al. Novel antitumor activity of the combined treatment of galloylquinic acid compounds with doxorubicin in solid Ehrlich carcinoma model via the notch signaling pathway modulation. Life Sci. 2022;299:120497. PubMed

Al-Madboly LA, et al. Novel preclinical study of Galloylquinic Acid compounds from Copaifera lucens with potent antifungal activity against Vaginal Candidiasis Induced in a murine model via Multitarget modes of Action. Microbiol Spectr. 2022;10(5):e0272421. PubMed PMC

Al-Madboly LA, et al. Characterization of GQA as a novel β-lactamase inhibitor of CTX-M-15 and KPC-2 enzymes. Microb Cell Fact. 2024;23(1):221. PubMed PMC

El-Salam MA, Furtado N, Haskic Z, Lieske J, Bastos J. Antiurolithic activity and biotransformation of galloylquinic acids by Aspergillus alliaceus ATCC10060, Aspergillus brasiliensis ATCC 16404, and Cunninghamella elegans ATCC 10028b. Biocatal Agric Biotechnol. 2019;18:101012. 10.1016/j.bcab.2019.01.050 PubMed PMC

El-Sisi AE, et al. The potential beneficial effects of sildenafil and diosmin in experimentally-induced gastric ulcer in rats. Heliyon. 2020;6(8):e04761. PubMed PMC

Abo Kamer AM, et al. Characterization of newly isolated bacteriophage to control multi-drug resistant Pseudomonas aeruginosa colonizing incision wounds in a rat model: in vitro and in vivo approach. Life Sci. 2022;310:121085. PubMed

El-Mahdy NA, et al. Targeting IL-10, ZO-1 gene expression and IL-6/STAT-3 trans-signaling by a combination of atorvastatin and mesalazine to enhance anti-inflammatory effects and attenuates progression of oxazolone-induced colitis. Fundam Clin Pharmacol. 2021;35(1):143–55. PubMed

Weinstein MP, Lewis JS 2. nd, The Clinical and Laboratory Standards Institute Subcommittee on Antimicrobial susceptibility testing: background, Organization, functions, and processes. J Clin Microbiol, 2020. 58(3). PubMed PMC

Lu L, et al. Quinic acid: a potential antibiofilm agent against clinical resistant Pseudomonas aeruginosa. Chin Med. 2021;16(1):72. PubMed PMC

Abdelaziz AA, et al. A purified and lyophilized Pseudomonas aeruginosa derived pyocyanin induces promising apoptotic and necrotic activities against MCF-7 human breast adenocarcinoma. Microb Cell Fact. 2022;21(1):262. PubMed PMC

Stepanovic S, et al. A modified microtiter-plate test for quantification of staphylococcal biofilm formation. J Microbiol Methods. 2000;40(2):175–9. PubMed

Kamali E, et al. Evaluation of antimicrobial resistance, biofilm forming potential, and the presence of biofilm-related genes among clinical isolates of Pseudomonas aeruginosa. BMC Res Notes. 2020;13(1):27. PubMed PMC

Duarte AC, et al. Synergistic action of phage phiIPLA-RODI and lytic protein CHAPSH3b: a combination strategy to target Staphylococcus aureus biofilms. NPJ Biofilms Microbiomes. 2021;7(1):39. PubMed PMC

Sankar Ganesh P, Ravishankar Rai V. Attenuation of quorum-sensing-dependent virulence factors and biofilm formation by medicinal plants against antibiotic resistant Pseudomonas aeruginosa. J Tradit Complement Med. 2018;8(1):170–7. PubMed PMC

Tkhilaishvili T, et al. Bacteriophage Sb-1 enhances antibiotic activity against biofilm, degrades exopolysaccharide matrix and targets persisters of Staphylococcus aureus. Int J Antimicrob Agents. 2018;52(6):842–53. PubMed

Rachmawati D, et al. In Vitro Assessment on Designing Novel antibiofilms of Pseudomonas aeruginosa using a Computational Approach. Molecules. 2022;27(24):8935. PubMed PMC

El-Shaer S, et al. Control of quorum sensing and virulence factors of Pseudomonas aeruginosa using phenylalanine arginyl β-naphthylamide. J Med Microbiol. 2016;65(10):1194–204. PubMed

Flemming HC, Wingender J. The biofilm matrix. Nat Rev Microbiol. 2010;8(9):623–33. PubMed

Adnan M, et al. Isolation and characterization of bacteriophage to control multidrug-resistant Pseudomonas aeruginosa planktonic cells and biofilm. Biologicals. 2020;63:89–96. PubMed

Deveaux W, Selvarajoo K. Searching for simple rules in Pseudomonas aeruginosa biofilm formation. BMC Res Notes. 2019;12(1):763. PubMed PMC

Kalia VC. Quorum sensing inhibitors: an overview. Biotechnol Adv. 2013;31(2):224–45. PubMed

Pancu DF et al. Antibiotics: conventional therapy and natural compounds with antibacterial Activity-A Pharmaco-Toxicological screening. Antibiot (Basel), 2021. 10(4). PubMed PMC

Grooters KE, et al. Strategies for combating antibiotic resistance in bacterial biofilms. Front Cell Infect Microbiol. 2024;14:1352273. PubMed PMC

Sweet R et al. Comparison of Antibacterial Activity of Phytochemicals against Common Foodborne pathogens and potential for selection of resistance. Microorganisms, 2023. 11(10). PubMed PMC

Suganya T, et al. Tackling multiple-drug-resistant Bacteria with conventional and complex phytochemicals. Front Cell Infect Microbiol. 2022;12:883839. PubMed PMC

da Trindade R, da Silva JK, Setzer WN. Copaifera of the neotropics: a review of the Phytochemistry and Pharmacology. Int J Mol Sci, 2018. 19(5). PubMed PMC

Tuon FF et al. Pathogenesis of the Pseudomonas aeruginosa Biofilm: a review. Pathogens, 2022. 11(3). PubMed PMC

Ryu JH, et al. Attachment and biofilm formation on stainless steel by Escherichia coli O157:H7 as affected by curli production. Lett Appl Microbiol. 2004;39(4):359–62. PubMed

Lima EMF, Winans SC, Pinto UM. Quorum sensing interference by phenolic compounds - A matter of bacterial misunderstanding. Heliyon. 2023;9(7):e17657. PubMed PMC

Hossain MA, et al. Impact of phenolic compounds in the acyl homoserine lactone-mediated quorum sensing regulatory pathways. Sci Rep. 2017;7(1):10618. PubMed PMC

Reen FJ, et al. Coumarin: a novel player in microbial quorum sensing and biofilm formation inhibition. Appl Microbiol Biotechnol. 2018;102(5):2063–73. PubMed PMC

Inchagova KS, Duskaev GK, Deryabin DG. Quorum sensing inhibition in Chromobacterium violaceum by Amikacin Combination with activated charcoal or small plant-derived molecules (pyrogallol and coumarin). Microbiology. 2019;88(1):63–71.