Bile-Salt-Hydrolases from the Probiotic Strain Lactobacillus johnsonii La1 Mediate Anti-giardial Activity in Vitro and in Vivo

Thibault Allain,Philippe Grellier,Isabelle Vallée,André G Buret,Bruno Polack,Luis G Bermúdez-Humarán,Philippe Langella,Myriam Thomas,Isabelle Florent,Soraya Chaouch

doi:10.3389/fmicb.2017.02707

Abstract

Giardia duodenalis (syn. G. lamblia, G. intestinalis) is the protozoan parasite responsible for giardiasis, the most common and widely spread intestinal parasitic disease worldwide, affecting both humans and animals. After cysts ingestion (through either contaminated food or water), Giardia excysts in the upper intestinal tract to release replicating trophozoites that are responsible for the production of symptoms. In the gut, Giardia cohabits with the host's microbiota, and several studies have revealed the importance of this gut ecosystem and/or some probiotic bacteria in providing protection against G. duodenalis infection through mechanisms that remain incompletely understood. Recent findings suggest that Bile-Salt-Hydrolase (BSH)-like activities from the probiotic strain of Lactobacillus johnsonii La1 may contribute to the anti-giardial activity displayed by this strain. Here, we cloned and expressed each of the three bsh genes present in the L. johnsonii La1 genome to study their enzymatic and biological properties. While BSH47 and BSH56 were expressed as recombinant active enzymes, no significant enzymatic activity was detected with BSH12. In vitro assays allowed determining the substrate specificities of both BSH47 and BSH56, which were different. Modeling of these BSHs indicated a strong conservation of their 3-D structures despite low conservation of their primary structures. Both recombinant enzymes were able to mediate anti-giardial biological activity against Giardia trophozoites in vitro. Moreover, BSH47 exerted significant anti-giardial effects when tested in a murine model of giardiasis. These results shed new light on the mechanism, whereby active BSH derived from the probiotic strain Lactobacillus johnsonii La1 may yield anti-giardial effects in vitro and in vivo. These findings pave the way toward novel approaches for the treatment of this widely spread but neglected infectious disease, both in human and in veterinary medicine.

Highlights

The amino acid sequences of L. johnsonii-BSH12, BSH47, and BSH56 enzymes were blasted against reported sequences from several Gram-positive bacteria using Blastp
Nucleotide homology comparisons previously highlighted the similarities between L. johnsonii La1 bsh12 and bsh56 with cbsHα and cbsHβ from L. johnsonii 100-100, respectively (Elkins and Savage, 1998; Elkins et al, 2001; Pridmore et al, 2004)
Neighbor-joining tree of various BSHs protein sequences from several lactic acid bacteria confirmed that L. johnsonii La1-BSH enzymes are phylogenetically related to BSHs from other species. They share a high degree of similarity to various subgroups of BSHs; for instance, BSH12 shares 99% identity with L. johnsonii 100-100 cbsHα and L. johnsonii PF01 BSHA at amino acid level, and BSH56 shares 99% identity with L. johnsonii 100100 cbsHβ and 98% with L. johnsonii PF01 BSHB at amino acid level

Summary

Introduction

Giardia lamblia and Giardia intestinalis) is a flagellated protozoan parasite responsible for giardiasis, an intestinal zoonotic disease infection that may cause acute or chronic diarrhea, weight loss, malabsorption, abdominal pain, and nausea (Ankarklev et al, 2010; Cotton et al, 2011). Metronidazole is the most frequently used drug for treating G. duodenalis infections, whereas albendazole, tinidazole, and nitazoxanide may be used with efficacy (Gardner and Hill, 2001; Petri, 2005) These drugs have different modes of action, there is an increasing incidence of parasite resistance, and treatment failure is relatively common (Ansell et al, 2015). These observations underscore the need for new therapeutic alternatives for the treatment of giardiasis

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