scholarly journals Extracellular DNA, cell surface proteins and c-di-GMP promote biofilm formation in Clostridioides difficile

2021 ◽  
Vol 11 (1) ◽  
Author(s):  
Lisa F. Dawson ◽  
Johann Peltier ◽  
Catherine L. Hall ◽  
Mark A. Harrison ◽  
Maria Derakhshan ◽  
...  
Keyword(s):  
Cell Surface ◽  
Biofilm Formation ◽  
Bacterial Species ◽  
Surface Proteins ◽  
Extracellular Dna ◽  
Cell Surface Proteins ◽  
Secondary Messenger ◽  
Clostridioides Difficile ◽  
Antibiotic Efficacy ◽  
Biofilm Matrix

AbstractClostridioides difficile is the leading cause of nosocomial antibiotic-associated diarrhoea worldwide, yet there is little insight into intestinal tract colonisation and relapse. In many bacterial species, the secondary messenger cyclic-di-GMP mediates switching between planktonic phase, sessile growth and biofilm formation. We demonstrate that c-di-GMP promotes early biofilm formation in C. difficile and that four cell surface proteins contribute to biofilm formation, including two c-di-GMP regulated; CD2831 and CD3246, and two c-di-GMP-independent; CD3392 and CD0183. We demonstrate that C. difficile biofilms are composed of extracellular DNA (eDNA), cell surface and intracellular proteins, which form a protective matrix around C. difficile vegetative cells and spores, as shown by a protective effect against the antibiotic vancomycin. We demonstrate a positive correlation between biofilm biomass, sporulation frequency and eDNA abundance in all five C. difficile lineages. Strains 630 (RT012), CD305 (RT023) and M120 (RT078) contain significantly more eDNA in their biofilm matrix than strains R20291 (RT027) and M68 (RT017). DNase has a profound effect on biofilm integrity, resulting in complete disassembly of the biofilm matrix, inhibition of biofilm formation and reduced spore germination. The addition of exogenous DNase could be exploited in treatment of C. difficile infection and relapse, to improve antibiotic efficacy.

scholarly journals What’s a Biofilm?—How the Choice of the Biofilm Model Impacts the Protein Inventory of Clostridioides difficile

2021 ◽  
Vol 12 ◽  
Author(s):  
Madita Brauer ◽  
Christian Lassek ◽  
Christian Hinze ◽  
Juliane Hoyer ◽  
Dörte Becher ◽  
...  
Keyword(s):  
Cell Surface ◽  
Biofilm Formation ◽  
Surface Proteins ◽  
Type Iv ◽  
Clostridioides Difficile ◽  
Biofilm Model ◽  
Proteomics Approach ◽  
Surface Polysaccharides ◽  
Mammalian Intestine

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.

scholarly journals Identification of Bicarbonate as a Trigger and Genes Involved with Extracellular DNA Export in Mycobacterial Biofilms

mBio ◽  
2016 ◽  
Vol 7 (6) ◽  
Author(s):  
Sasha J. Rose ◽  
Luiz E. Bermudez
Keyword(s):  
Nontuberculous Mycobacteria ◽  
Bacterial Species ◽  
Genomic Region ◽  
Surface Proteins ◽  
Extracellular Dna ◽  
Carbonic Anhydrases ◽  
Independent Manner ◽  
Genes Encoding ◽  
Library Screen ◽  
Biofilm Matrix

ABSTRACTExtracellular DNA (eDNA) is an integral biofilm matrix component of numerous pathogens, including nontuberculous mycobacteria (NTM). Cell lysis is the source of eDNA in certain bacteria, but the source of eDNA remains unidentified for NTM, as well as for other eDNA-containing bacterial species. In this study, conditions affecting eDNA export were examined, and genes involved with the eDNA export mechanism were identified. After a method for monitoring eDNA in real time in undisturbed biofilms was established, different conditions affecting eDNA were investigated. Bicarbonate positively influenced eDNA export in a pH-independent manner inMycobacterium avium,M. abscessus, andM. chelonae. The surface-exposed proteome ofM. aviumin eDNA-containing biofilms revealed abundant carbonic anhydrases. Chemical inhibition of carbonic anhydrases with ethoxzolamide significantly reduced eDNA export. An unbiased transposon mutant library screen for eDNA export inM. aviumidentified many severely eDNA-attenuated mutants, including one not expressing a unique FtsK/SpoIIIE-like DNA-transporting pore, two with inactivation of carbonic anhydrases, and nine with inactivation of genes belonging to a unique genomic region, as well as numerous mutants involved in metabolism and energy production. Complementation of nine mutants that included the FtsK/SpoIIIE and carbonic anhydrase significantly restored eDNA export. Interestingly, several attenuated eDNA mutants have mutations in genes encoding proteins that were found with the surface proteomics, and many more mutations are localized in operons potentially encoding surface proteins. Collectively, our data strengthen the evidence of eDNA export being an active mechanism that is activated by the bacterium responding to bicarbonate.IMPORTANCEMany bacteria contain extracellular DNA (eDNA) in their biofilm matrix, as it has various biological and physical functions. We recently reported that nontuberculous mycobacteria (NTM) can contain significant quantities of eDNA in their biofilms. In some bacteria, eDNA is derived from dead cells, but that does not appear to be the case for all eDNA-containing organisms, including NTM. In this study, we found that eDNA export in NTM is conditionally dependent on the molecules to which the bacteria are exposed and that bicarbonate positively influences eDNA export. We also identified genes and proteins important for eDNA export, which begins to piece together a description of a mechanism for eDNA. Better understanding of eDNA export can give new targets for the development of antivirulence drugs, which are attractive because resistance to classical antibiotics is currently a significant problem.

scholarly journals Role of SrtA in Pathogenicity of Staphylococcus lugdunensis

2020 ◽  
Vol 8 (12) ◽  
pp. 1975
Author(s):  
Muzaffar Hussain ◽  
Christian Kohler ◽  
Karsten Becker
Keyword(s):  
Cell Wall ◽  
Cell Surface ◽  
Biofilm Formation ◽  
Surface Proteins ◽  
Transcription Level ◽  
Coagulase Negative Staphylococci ◽  
Staphylococcus Lugdunensis ◽  
Cell Surface Proteins ◽  
Mutant Strains

Among coagulase-negative staphylococci (CoNS), Staphylococcus lugdunensis has a special position as causative agent of aggressive courses of infectious endocarditis (IE) more reminiscent of IEs caused by Staphylococcus aureus than those by CoNS. To initiate colonization and invasion, bacterial cell surface proteins are required; however, only little is known about adhesion of S. lugdunensis to biotic surfaces. Cell surface proteins containing the LPXTG anchor motif are covalently attached to the cell wall by sortases. Here, we report the functionality of Staphylococcus lugdunensis sortase A (SrtA) to link LPXTG substrates to the cell wall. To determine the role of SrtA dependent surface proteins in biofilm formation and binding eukaryotic cells, we generated SrtA-deficient mutants (ΔsrtA). These mutants formed a smaller amount of biofilm and bound less to immobilized fibronectin, fibrinogen, and vitronectin. Furthermore, SrtA absence affected the gene expression of two different adhesins on transcription level. Surprisingly, we found no decreased adherence and invasion in human cell lines, probably caused by the upregulation of further adhesins in ΔsrtA mutant strains. In conclusion, the functionality of S. lugdunensis SrtA in anchoring LPXTG substrates to the cell wall let us define it as the pathogen’s housekeeping sortase.

scholarly journals The dynamics and pH-dependence of Ag43 adhesins’ self-association probed by atomic force spectroscopy

Nanoscale ◽  
2014 ◽  
Vol 6 (21) ◽  
pp. 12665-12681 ◽  
Author(s):  
Adrien Jacquot ◽  
Chizuko Sakamoto ◽  
Angelina Razafitianamarahavo ◽  
Céline Caillet ◽  
Jenny Merlin ◽  
...  
Keyword(s):  
Cell Surface ◽  
Biofilm Formation ◽  
Ph Dependence ◽  
Force Spectroscopy ◽  
Surface Proteins ◽  
Cell Surface Proteins ◽  
Atomic Force Spectroscopy ◽  
Atomic Force ◽  
Self Association

Self-associating auto-transporter (SAAT) adhesins are two-domain cell surface proteins involved in bacteria auto-aggregation and biofilm formation.

What do nanometer molecular trajectories tell us about heterogeneous cell surface domaines?

1995 ◽  
Vol 53 ◽  
pp. 786-787
Author(s):  
Watt W. Webb
Keyword(s):  
Cell Surface ◽  
Lipid Membranes ◽  
Surface Proteins ◽  
Protein Diffusion ◽  
Coated Pits ◽  
Cell Surface Proteins ◽  
Membrane Heterogeneity ◽  
Fluorescence Photobleaching Recovery ◽  
Photobleaching Recovery ◽  
Plasma Membrane Heterogeneity

Plasma membrane heterogeneity is implicit in the existence of specialized cell surface organelles which are necessary for cellular function; coated pits, post and pre-synaptic terminals, microvillae, caveolae, tight junctions, focal contacts and endothelial polarization are examples. The persistence of these discrete molecular aggregates depends on localized restraint of the constituent molecules within specific domaines in the cell surface by strong intermolecular bonds and/or anchorage to extended cytoskeleton. The observed plasticity of many of organelles and the dynamical modulation of domaines induced by cellular signaling evidence evanescent intermolecular interactions even in conspicuous aggregates. There is also strong evidence that universal restraints on the mobility of cell surface proteins persist virtually everywhere in cell surfaces, not only in the discrete organelles. Diffusion of cell surface proteins is slowed by several orders of magnitude relative to corresponding protein diffusion coefficients in isolated lipid membranes as has been determined by various ensemble average methods of measurement such as fluorescence photobleaching recovery(FPR).

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