Citrus Extracts Inhibit Quorum Sensing and Expression of flaA-B and cadF in Campylobacter jejuni

Introduction:  Campylobacter jejuni is recognized as one of the most common cause of foodborne bacterial gastroenteritis in humans, which can range from asymptomatic to dysentery type illnesses, with severe complications such as Guillain Barre Syndrome. flaA-B and cad F genes are involved in motility and invasion which are crucial steps in C. jejuni pathogenesis. Quorum sensing (QS) regulates several virulence factors including changes in gene expression. Thus, compounds capable to inhibit QS present a novel target in the development of new antimicrobials.

Purpose:  To investigate the effect of citrus extracts in the activity of AI-2 molecules which mediate QS in C. jejuni and their effect on the expression of genes involved in motility and invasion.

Methods: Peel, seeds and bagasse of Citrus limon, C. medica, and C. aurantium were extracted with ethanol. Minimum bactericidal concentrations (MBCs) of citrus extracts against C. jejuni were determined by the microplate-dilution method. The effect of extracts (at subinhibitory concentrations, 25%, 50%, and 75% MBC) on AI-2 activity of C. jejuni was determined by a bioluminescent method using V. harveyi BB170. To measure the relative expression of flaA-B and cadF, RNA from cultures treated with sub-inhibitory concentration of extracts was obtained, and gene expression was determined by real-time quantitative RT-PCR.

Results: The MBC of the citrus extracts ranged from 1.3 to 3.25 mg/ml. Concentrations lower than the MBC did not affect growth of bacteria, however, AI-2 activity decreased in most cases at levels of 90% (Relative light units). The relative expression of flaA-B and cadF was significantly (P ≤ 0.5) reduced or totally inhibited by citrus extracts. 

 Significance : Extracts from citrus waste are an alternative to control Campylobacter by inhibiting growth and virulence factors mediated by QS.