What's a Biofilm?-How the Choice of the Biofilm Model Impacts the Protein Inventory of Clostridioides difficile.

Madita Brauer,Dieter Jahn,Christian Lassek,Susanne Sievers,Juliane Hoyer,Katharina Riedel,Dörte Becher,Christian Hinze

doi:10.3389/fmicb.2021.682111

Madita Brauer, Dieter Jahn + Show 6 more

Open Access

https://doi.org/10.3389/fmicb.2021.682111

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Abstract

The anaerobic pathogen Clostridioides difficile is perfectly equipped to survive and persist inside the mammalian intestine. When facing unfavorable conditions C. difficile is able to form highly resistant endospores. Likewise, biofilms are currently discussed as form of persistence. Here a comprehensive proteomics approach was applied to investigate the molecular processes of C. difficile strain 630Δerm underlying biofilm formation. The comparison of the proteome from two different forms of biofilm-like growth, namely aggregate biofilms and colonies on agar plates, revealed major differences in the formation of cell surface proteins, as well as enzymes of its energy and stress metabolism. For instance, while the obtained data suggest that aggregate biofilm cells express both flagella, type IV pili and enzymes required for biosynthesis of cell-surface polysaccharides, the S-layer protein SlpA and most cell wall proteins (CWPs) encoded adjacent to SlpA were detected in significantly lower amounts in aggregate biofilm cells than in colony biofilms. Moreover, the obtained data suggested that aggregate biofilm cells are rather actively growing cells while colony biofilm cells most likely severely suffer from a lack of reductive equivalents what requires induction of the Wood-Ljungdahl pathway and C. difficile’s V-type ATPase to maintain cell homeostasis. In agreement with this, aggregate biofilm cells, in contrast to colony biofilm cells, neither induced toxin nor spore production. Finally, the data revealed that the sigma factor SigL/RpoN and its dependent regulators are noticeably induced in aggregate biofilms suggesting an important role of SigL/RpoN in aggregate biofilm formation.

Highlights

In recent years, the anaerobic gastrointestinal pathogen Clostridioides difficile has established itself as one of the major causative agents of pseudomembranous colitis and toxic megacolon (Wiegand et al, 2012; Nasiri et al, 2018; Martínez-Meléndez et al, 2020; Doll et al, 2021)
Two different types of biofilms were investigated: 1. colony biofilms on agar plates and 2. aggregate biofilms formed in 6well plates
Colonies on agar plates do not meet the traditional criteria of a biofilm, the densely packed cells likewise present a form of multicellular growth attached to a biotic surface (Gingichashvili et al, 2017; Kesel et al, 2017) and it cannot be excluded that the growth conditions that bacteria face during this form of multicellular living are relevant during infection

Summary

Introduction

The anaerobic gastrointestinal pathogen Clostridioides difficile has established itself as one of the major causative agents of pseudomembranous colitis and toxic megacolon (Wiegand et al, 2012; Nasiri et al, 2018; Martínez-Meléndez et al, 2020; Doll et al, 2021). Persistence does obviously not solely rely on sporulation and on additional features of C. difficile (Smits, 2013) In this context, biofilm formation was proposed to be a major additional factor (Dawson et al, 2012; Ðapa et al, 2013). Many nosocomial infections caused by Staphylococcus aureus, Streptococci and numerous other pathogens rely on biofilms (Jamal et al, 2018). In this context, biofilm formation has been frequently linked to pro-longed infection and persistence (Burmølle et al, 2010; Jamal et al, 2018). Anaerobic bacteria, including C. difficile have been found to produce biofilms in vitro, and biofilm-like structures have been observed on the intestinal mucosal surface of C. difficile infected mice (Donelli et al, 2012; Lawley et al, 2012; Soavelomandroso et al, 2017)

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