Antimicrobial Activity And Mechanism Of Ajoene And Metal Oxide Nanoparticles To Campylobacter Jejuni

Campylobacter jejuni, a main pathogen, may lead to enteritis by infecting intestinal epithelial cells, and in some instances may be associated with severe enteritis, septicaemia, Crohn’s disease, reactive arthritis and a higher incidence of Guillain-Barre syndrome. In order to control the widely spread and infection of Campylobacter jejuni, we present the first study characterizing pathogen stress under synergistic antimicrobial treatment of organic antibacterial material ajoene and metal oxide nanoparticles (TiO2 nanoparticles and Al2O3 nanoparticles). The inhibitory effects of different compounds on Campylobacter jejuni were determined with broth dilution method. Scanning electron microsopy and confocal laser scanning microscopy were applied to observe the morphology of the damaged bacterial cells. And by gathering and analysising Raman spectroscopy of damaged cells, the inhibitory mechanism was studied.Ajoene, a garlic-derived organosulfur compound decomposed from unstable thiosulfinates, was found to be highly effective antimicrobial against Campylobacter jejuni. TiO2 nanoparticles and Al2O3 nanoparticles could inhibit the growth of Campylobacter jejuni when their concentrations were high enough. We also found ajoene and metal oxide nanoparticles had a synergistic antimicrobial effect on this bacteria.Scanning electron microscopy (SEM) confirmed cell injury showing significant morphological variations in bacterial cells following treatments by these different compounds.With the appropriate mixture of the SYTO 9 and propidium iodide stains, bacteria with intact cell membranes fluoresce bright green, whereas bacteria with damaged membranes exhibit significantly less green fluorescence and often also fluoresce red. Confocal laser scanning microscopy (CLSM) confirmed cell injury intuitively in bacterial cells following treatments by these different compounds. Synergistic antimicrobial effect on this bacteria of ajoene and metal oxide nanoparticles could be observed with CLSM.Confocal micro-Raman spectroscopy characterizes macromolecular changes in the bacterial cell resulting from cell injury. The bacterial response to ajoene differed from metal oxide nanoparticles. Confocal micro-Raman spectroscopy revealed that ajoene was able to penetrate through the cell membrane and bind to thiol-containing proteins/enzymes in bacterial cells generating a disulfide stretching band. Then the structures of relevant enzymes and proteins (band at 1606 cm-1) were transformed, resulting in biological dysfunction. Metal oxide nanoparticles appeared to have the same phase I antimicrobial mechanism of disorganizing disulfide stretching bands in bacterial membrane and forming hydrocarbons, but a different phase II showing alterations in the structures of amino acids, nucleotides and proteins in two different ways. Al2O3 nanoparticles primarily altered the a-helix and β-sheet as reflected in changes in the amide I (bands at 759 cm-1 and 775 cm-1) while TiO2 nanoparticles altered the structures containing tyrosine and guanine (band at 654 cm-1).Bayesian probability validated the ability of unsupervised chemometric models [principal component analysis (PCA)] to differentiate treated and control Campylobacter jejuni cells.Results of this study can aid in the development of more effective intervention strategies to reduce the risk of Campylobacter jejuni contamination in the food supply, specifically in dehydrated products consumed by susceptible populations.