Bioengineered Silver Nanoparticle from Spirulina platensis in Attenuating Biofilm Mediated Virulence in Vibrio parahemolyticus: An in vitro and in vivo Approach

Abstract

Objectives: In this study, bioengineered AgNPs (silver nanoparticles) from Spirulina platensis was used to counter the biofilm mediated virulence by Vibrio parahemolyticus.Methods: AgNPs were synthesized from crude Spirulina platensis extract and characterized using UV spectrophotometry, Transmission electron microscope (TEM). Antibacterial effect against aquatic pathogen Vibrio parahemolyticus was carried out. Further, the nanoparticle at sub MIC (minimum inhibitory concentration) level were screened for anti-virulence property under in vitro studies which include biofilm inhibition using microtiter plate assay, microscopic analysis under light microscope and SEM (Scanning electron microscope) imaging, EPS (Exopolysaccharide) inhibition, CSH(Cell surface hydrophobicity) inhibition using BATH(Bacterial adhesion to hydrocarbons) assay, swarming and swimming motility inhibition, growth pattern analysis followed by in vivo study in aquatic animal model brine shrimp and its survival percentage along with light microscopic imaging was performed.Results: AgNPs, at 320µg/ml showed bactericidal activity, minimal inhibitory concentration was found to be 160 µg/ml. All anti-virulence assay performed at below sub MIC concentration. Biofilm and EPS showed 63% and 53% inhibition respectively, CSH was found to be decreased 3-fold. Growth pattern at sub MIC compared with non-treated showed similar result. In vivo study increased survival percentage (90%) when treated with AgNPs, compared to untreated (20%). A 7-fold increased resistance to pathogen was found and light microscope also confirms the efficacy of Ag nanoparticle.Conclusion: AgNPs become a substitute to the current commercial antibiotics used in aquaculture against Vibriosis, by functioning as an efficient quorum sensing inhibitor.

Keywords: AgNPs; Vibrio parahemolyticus; Anti-virulence; Biofilm; Survival rate