Citrus paradisi and Citrus reticulata essential oils interfere with Pseudomonas aeruginosa quorum sensing in vivo on Caenorhabditis elegans

Abstract

Background: Pseudomonas aeruginosa is a Gram-negative opportunistic pathogen, considered a leading cause of acute and chronic infections in immunocompromised patients. P. aeruginosa uses quorum sensing to control virulence and biofilm formation. To combat this human-resistant pathogen increasing attention has been paid to anti-QS compounds from natural products as potential therapeutic agents. Purpose: To assess the efficacy of C. paradisi (Grapefruit) and C. reticulata (Mandarin) commercial EOs (obtained by cold-pressing and cold-pressing followed by steam distillation, named EOP and EOPD, respectively) and their majority component, limonene, in inhibiting the effect of QS-controlled virulence factors of P. aeruginosa PAO1 and PA14 in a C. elegans infection model. Results: C. paradisi and C. reticulata EOs at 0.125% (v/v) significantly inhibited the in vitro biofilm formation of PAO1 and PA14 strains between 40 and 50%. This EOs concentration, safe for C. elegans according to the survival and brood size assays, rescued the nematodes from P. aeruginosa infections, reducing their death rate by 20–30% in the paralysis assay, and increasing worm lifespan with median survivals of 6 (Grapefruit) and 5 (Mandarin) in the slow killing assay (3 for non-treated worms). Limonene protected nematodes from death in these assays although less effectively than both EOs species. Grapefruit EOs rescued the nematodes in 45–50% from phenazine-death, while mandarin EOs and limonene rescued them in 30%. In the food choice assay, worms preferred the PA14 grown with 0.125% of mandarin and grapefruit EOs or limonene after 3-4 hours (CI=0.11 to 0.18). Conclusion: Our results suggest that C. paradisi and C. reticulata EOPDs, with lower commercial value but similar in vivo effects than EOPs, are sources of anti-QS agents for controlling P. aeruginosa infections, therefore should be considered as potential candidates for the development of novel therapeutics against persistent microorganisms.