Dataset Information

Differences in gene expression between colony and aggregate biofilms of Clostridioides difficile revealed by proteomic profiling

ABSTRACT: The protein inventory of the anaerobic pathogen Clostridioides difficile grown in colony or aggregate biofilms was investigated in a comprehensive LC-MS/MS approach. C. difficile has a versatile metabolism and is perfectly equipped to survive and persist inside the mammalian intestine. However, knowledge on how C. difficile exactly resides inside the intestine is still scare. Aggregate or biofilms attached to the epithelium are discussed as possible growth forms and have become a focus in research. However, recent data suggest that the choice of the model strain, timepoint of analysis and the biofilm model significantly impact the outcome of biofilm experiments leading to contradictory results. To address some of these issues, a comprehensive proteomics approach was applied to investigate the metabolism of C. difficile strain 630erm grown in colony or aggregate biofilms. Besides verification of recent findings, a comprehensive overview on the differential expression profile of C. difficile’s cell surface proteins as well as its energy and stress metabolism during the two different forms of biofilm growth could be provided. Additionally, regulatory circuits that were activated in the two different biofilm models were analysed whereby RpoN was determined to be an important regulator of aggregate biofilm formation in C. difficile.

INSTRUMENT(S): LTQ Orbitrap Elite

ORGANISM(S): Peptoclostridium Difficile (strain 630) (clostridium Difficile)

SUBMITTER: Daniela Zuehlke

LAB HEAD: Katharina Riedel

PROVIDER: PXD022830 | Pride | 2021-07-12

REPOSITORIES: Pride

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Publications

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

Brauer Madita M Lassek Christian C Hinze Christian C Hoyer Juliane J Becher Dörte D Jahn Dieter D Sievers Susanne S Riedel Katharina K

Frontiers in microbiology 20210610

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 fro ...[more]

PMID: 34177868

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Project description:Polymicrobial biofilms are of large medical importance, but little is known about their physiology and the underlying interspecies interactions. Here we studied two human pathogens, the opportunistic fungus Candida albicans and the caries promoting bacterium Streptococcus mutans. Both species formed biofilms in monoculture, with C. albicans growing mainly in the virulence-associated hyphae form, and S. mutans forming a thick layer of extracellular polymeric substances (EPS). Biofilm growth was enhanced in dual-species biofilms, which reached twice the biomass of monospecies biofilms and higher cell numbers of both S. mutans and C. albicans. EPS production by S. mutans was strongly suppressed in dual-species biofilms. Virulence traits of S. mutans, e.g. genetic competence, biofilm formation and bacteriocin synthesis are controlled by quorum sensing through activation of the alternative sigma factor SigX. SigX is induced by the pheromones CSP (competence stimulating factor) or XIP (sigX inducing peptide). Strong induction of sigX was observed in dual species biofilms indicated by fluorescence of a reporter strain for the sigX promoter, S. mutans PcomX-gfp, as well as by qRT-PCR of comX. The peak of sigX expression occurred after 10 h of biofilm growth. Conditioned media from mixed biofilms but not from C. albicans or S. mutans cultivated alone activated sigX in the reporter strain. Deletion mutants for the comC and comS genes encoding the precursors of CSP and XIP, respectively, were constructed. Conditioned media from mixed biofilms with S. mutans DcomS were unable to induce sigX in the reporter strain, while deletion of comC had no effect. These data show that synthesis of XIP was induced in S. mutans by coculture with C. albicans. Transcriptome analysis of S. mutans in single and mixed biofilms confirmed strong induction of comS, sigX, and the downstream late competence genes in dual-species biofilms. Among the late competence genes, fratricins were discovered for the first time. The comCDE operon and bacteriocin related genes were also induced, but much weaker. Genes related to oxidative stress, chaperones and glycosyltransferase genes required for EPS synthesis from sucrose were down-regulated, while glycogen synthesis genes were up-regulated, indicating that S. mutans was protected from oxidative stress and provided with excess sugar for storage polymer synthesis in mixed biofilms. The data show that in dual-species biofilms, C. albicans improves growth of S. mutans, suppresses its EPS formation and induces the complete quorum sensing signalling system, thus fundamentally changing the virulence properties of the caries pathogen, including its potential interactions with other members of the polymicrobial dental plaque community. Dual-species biofilms of S. mutans and C. albicans and single-species biofilms of S. mutans were cultivated in 24-well microtitre plates in YNBB medium. Transcriptional profiles of S. mutans in single- and dual-species biofilms were analysed at early (6 h) and late (10 h) logarithmic phase of the biofilm growth, as well as after 24 h when biofilms entered stationary phase. Transcriptional profiles of S. mutans grown in the dual-species biofilms were compared to profiles obtained for single-species biofilm from the same time point. Three biological and one to two technical replicas were used in the microarray study. RNA samples were labeled with Cy3 or Cy5 using the ULS fluorescent labeling kit (Kreatech, Germany). Seven hundred nanograms of Cy3 or Cy5 labeled RNA after fragmentation were hybridized to the microarray at 65M-BM-0C for 17 h using the Agilent hybridization chamber according to the manufacturer's instructions. The arrays were scanned using the Agilent DNA microarray scanner and the raw data were extracted using Agilent Feature Extraction software (v. 10.7).

Dataset Information

Differences in gene expression between colony and aggregate biofilms of Clostridioides difficile revealed by proteomic profiling

Publications

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

Similar Datasets

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