Giardia extracellular vesicles disrupt intestinal epithelial junctions and inhibit the growth of commensal bacteria while increasing their swimming motility
Authors: Siddiq Affan , Allain Thibault , Dong George , Olivier Martin , Buret Andre 
Published: April 2020

IntroductionExtracellular vesicles (EVs) have emerged as important mediators of host‐parasite interactions. The protozoan parasite Giardia duodenalis, the most common cause of parasitic diarrheal diseases worldwide, produces EVs in response to changes in the environment. The exact role of EVs in the pathogenesis of giardiasis remains unclear. In this study, we investigate the effects of EVs on the intestinal barrier such as the intestinal epithelium and the gut commensal bacteria. Disruption of the intestinal barrier has been implicated in the development of post‐giardiasis gut disorders and EVs may represent a novel mechanism of this disruption.AimsThe aim of this research is to characterize Giardia EVs and investigate their effect on the host epithelium and commensal gut bacteria.MethodsG. duodenalis (isolate NF) trophozoites were either incubated with 10X bile for 1 hour or left untreated and EVs were isolated using Qiagen Exo‐Easy Maxi Kit. The concentration and size of EVs were assessed using Nanosight track analysis (NTA). Proteomic analysis of EVs was conducted using liquid chromatography with tandem mass spectrometry. The effects of Giardia EVs on the host epithelium were assessed by exposing epithelial primary cell line (SCBN) to EVs for 24 hours and examining the tight junction proteins ZO‐1 and claudin‐4 by fluorescence microscopy. Antibacterial effect of Giardia EVs was assessed on gut commensals such as Escherichia coli strain HB101 and strain K12, Enterobacter cloacae (human isolate) and Enterococcus faecalis (human isolate) by measuring bacterial kinetic growth for 12 hours. Swimming motility of non‐invasive commensals was assessed on 0.3% agar after incubating with EVs for 24 hours and the halo sizes were measured as a marker of swimming behavior.ResultsOur findings indicate that bile treated Giardia trophozoites produce more EVs as compared to untreated trophozoites. NTA revealed that Giardia EVs range around 200 nm. Proteomic analysis of EVs revealed the presence of well characterized Giardia virulence factors such as cysteine proteases, tenascins, oxidative defense enzymes, variant surface proteins as well as metabolic enzymes such as arginine deaminase and ornithine carbamoyl transferase. Additionally, exposure of SCBN cells to EVs resulted in the disruption and/or rearrangement of tight junction proteins Zo‐1 and Claudin‐4. Moreover, our results also indicate that Giardia EVs exert bacteriostatic effects on E. coli strains HB101, K12, E. cloacae and E. faecalis. Finally, Giardia EVs also significantly increased the swimming motility of non‐invasive commensals E. coli strain HB101 and E. cloacae.ConclusionOur research highlights a novel mechanism of Giardia’s interaction with the intestinal barrier. Giardia EVs contain virulence factors that disrupt the host’s intestinal epithelium and modify the growth and motility of commensal bacteria. These effects may have significant implications in disease pathophysiology.Support or Funding InformationNatural Sciences and Engineering Research Council (NSERC)