1.
Castelain, Mickaël, et al.
(author)
Fast uncoiling kinetics of F1C pili expressed by uropathogenic Escherichia coli are revealed on a single pilus level using force-measuring optical tweezers
2011
In: European Biophysics Journal. - : Springer Science and Business Media LLC. - 0175-7571 .- 1432-1017. ; 40:3, s. 305-316
Journal article (peer-reviewed) abstract
Uropathogenic Escherichia coli (UPEC) expressvarious kinds of organelles, so-called pili or fimbriae, thatmediate adhesion to host tissue in the urinary tract throughspecific receptor-adhesin interactions. The biomechanicalproperties of these pili have been considered important forthe ability of bacteria to withstand shear forces from rinsingurine flows. Force-measuring optical tweezers have beenused to characterize individual organelles of F1C typeexpressed by UPEC bacteria with respect to such properties.Qualitatively, the force-versus-elongation response wasfound to be similar to that of other types of helix-like piliexpressed by UPEC, i.e., type 1, P, and S, with force-inducedelongation in three regions, one of which represents theimportant uncoiling mechanism of the helix-like quaternarystructure. Quantitatively, the steady-state uncoiling forcewas assessed as 26.4 ±1.4 pN, which is similar to those ofother pili (which range from 21 pN for SI to 30 pN for type 1).The corner velocity for dynamic response (1,400 nm/s) wasfound to be larger than those of the other pili (400–700 nm/sfor S and P pili, and 6 nm/s for type 1). The kinetics werefound to be faster, with a thermal opening rate of 17 Hz, afew times higher than S and P pili, and three orders ofmagnitude higher than type 1. These data suggest that F1Cpili are, like P and S pili, evolutionarily selected to primarilywithstand the conditions expressed in the upper urinary tract.
2.
Karah, Nabil, et al.
(author)
Novel Aminoglycoside Resistance Transposons and Transposon-Derived Circular Forms Detected in Carbapenem-Resistant Acinetobacter baumannii Clinical Isolates
2016
In: Antimicrobial Agents and Chemotherapy. - 0066-4804 .- 1098-6596. ; 60:3, s. 1801-1818
Journal article (peer-reviewed) abstract
Acinetobacter baumannii has emerged as an important opportunistic pathogen equipped with a growing number of antibiotic resistance genes. Our study investigated the molecular epidemiology and antibiotic resistance features of 28 consecutive carbapenem-resistant clinical isolates of A. baumannii collected throughout Sweden in 2012 and 2013. The isolates mainly belonged to clonal complexes (CCs) with an extensive international distribution, such as CC2 (n = 16) and CC25 (n = 7). Resistance to carbapenems was related to bla(OXA-23) (20 isolates), bla(OXA-24/40-like) (6 isolates), bla(OXA-467) (1 isolate), and ISAba1-bla(OXA-69) (1 isolate). Ceftazidime resistance was associated with bla(PER-7) in the CC25 isolates. Two classical point mutations were responsible for resistance to quinolones in all the isolates. Isolates with high levels of resistance to aminoglycosides carried the 16S rRNA methylase armA gene. The isolates also carried a variety of genes encoding aminoglycoside-modifying enzymes. Several novel structures involved in aminoglycoside resistance were identified, including Tn6279, Delta Tn6279, Ab-ST3- aadB, and different assemblies of Tn6020 and TnaphA6. Importantly, a number of circular forms related to the IS26 or ISAba125 composite trans-posons were detected. The frequent occurrence of these circular forms in the populations of several isolates indicates a potential role of these circular forms in the dissemination of antibiotic resistance genes.
3.
Song, Tianyan, et al.
(author)
Vibrio cholerae Utilizes Direct sRNA Regulation in Expression of a Biofilm Matrix Protein
2014
In: PLOS ONE. - : PLOS. - 1932-6203. ; 9:7
Journal article (peer-reviewed) abstract
Vibrio cholerae biofilms contain exopolysaccharide and three matrix proteins RbmA, RbmC and Bap1. While much is known about exopolysaccharide regulation, little is known about the mechanisms by which the matrix protein components of biofilms are regulated. VrrA is a conserved, 140-nt sRNA of V. cholerae, whose expression is controlled by sigma factor sigma(E). In this study, we demonstrate that VrrA negatively regulates rbmC translation by pairing to the 5' untranslated region of the rbmC transcript and that this regulation is not stringently dependent on the RNA chaperone protein Hfq. These results point to VrrA as a molecular link between the sigma(E)-regulon and biofilm formation in V. cholerae. In addition, VrrA represents the first example of direct regulation of sRNA on biofilm matrix component, by-passing global master regulators.
4.
Erttmann, Saskia F., et al.
(author)
The gut microbiota prime systemic antiviral immunity via the cGAS-STING-IFN-I axis
2022
In: Immunity. - : Elsevier BV. - 1074-7613 .- 1097-4180. ; 55:5, s. 847-861
Journal article (peer-reviewed) abstract
The microbiota are vital for immune homeostasis and provide a competitive barrier to bacterial and fungal pathogens. Here, we investigated how gut commensals modulate systemic immunity and response to viral infection. Antibiotic suppression of the gut microbiota reduced systemic tonic type I interferon (IFN-I) and antiviral priming. The microbiota-driven tonic IFN-I-response was dependent on cGAS-STING but not on TLR signaling or direct host-bacteria interactions. Instead, membrane vesicles (MVs) from extracellular bacteria activated the cGAS-STING-IFN-I axis by delivering bacterial DNA into distal host cells. DNA-containing MVs from the gut microbiota were found in circulation and promoted the clearance of both DNA (herpes simplex virus type 1) and RNA (vesicular stomatitis virus) viruses in a cGAS-dependent manner. In summary, this study establishes an important role for the microbiota in peripheral cGAS-STING activation, which promotes host resistance to systemic viral infections. Moreover, it uncovers an underappreciated risk of antibiotic use during viral infections.
5.
Aung, Kyaw Min, et al.
(author)
Naturally Occurring IgG Antibodies Provide Innate Protection against Vibrio cholerae Bacteremia by Recognition of the Outer Membrane Protein U
2016
In: Journal of Innate Immunity. - : S. Karger AG. - 1662-811X .- 1662-8128. ; 8:3, s. 269-283
Journal article (peer-reviewed) abstract
Cholera epidemics are caused by Vibrio cholerae serogroups O1 and O139, whereas strains collectively known as non-O1/non-O139 V. cholerae are found in cases of extraintestinal infections and bacteremia. The mechanisms and factors influencing the occurrence of bacteremia and survival of V. cholerae in normal human serum have remained unclear. We found that naturally occurring IgG recognizing V. cholerae outer membrane protein U (OmpU) mediates a serum-killing effect in a complement C1q-dependent manner. Moreover, outer membrane vesicles (OMVs) containing OmpU caused enhanced survival of highly serum-sensitive classical V. cholerae in a dose-dependent manner. OMVs from wild-type and ompU mutant V. cholerae thereby provided a novel means to verify by extracellular transcomplementation the involvement of OmpU. Our data conclusively indicate that loss, or reduced expression, of OmpU imparts resistance to V. cholerae towards serum killing. We propose that the difference in OmpU protein levels is a plausible reason for differences in serum resistance and the ability to cause bacteremia observed among V. cholerae biotypes. Our findings provide a new perspective on how naturally occurring antibodies, perhaps induced by members of the microbiome, may play a role in the recognition of pathogens and the provocation of innate immune defense against bacteremia.
6.
Corkery, Dale, et al.
(author)
Vibrio cholerae cytotoxin MakA induces noncanonical autophagy resulting in the spatial inhibition of canonical autophagy
2021
In: Journal of Cell Science. - : The Company of Biologists. - 0021-9533 .- 1477-9137. ; 134:5
Journal article (peer-reviewed) abstract
Autophagy plays an essential role in the defense against manymicrobial pathogens as a regulator of both innate and adaptive immunity. Some pathogens have evolved sophisticated mechanisms that promote their ability to evade or subvert host autophagy. Here, we describe a novel mechanism of autophagy modulation mediated by the recently discovered Vibrio cholerae cytotoxin, motility-associatedkilling factor A (MakA). pH-dependent endocytosis of MakA by host cells resulted in the formation of a cholesterol-rich endolysosomal membrane aggregate in the perinuclear region. Aggregate formation induced the noncanonical autophagy pathway driving unconventional LC3 (herein referring to MAP1LC3B) lipidation on endolysosomal membranes. Subsequent sequestration of the ATG12-ATG5-ATG16L1 E3-like enzyme complex, required for LC3 lipidation at the membranous aggregate, resulted in an inhibition of both canonical autophagy and autophagy-related processes, including the unconventional secretion of interleukin-1β (IL-1β). These findings identify a novel mechanismof host autophagy modulation and immune modulation employed by V. cholerae during bacterial infection.
7.
Graffeuil, Antoine, et al.
(author)
Polar mutagenesis of polycistronic bacterial transcriptional units using Cas12a
2022
In: Microbial Cell Factories. - : BioMed Central. - 1475-2859 .- 1475-2859. ; 21:1
Journal article (peer-reviewed) abstract
Background: Functionally related genes in bacteria are often organized and transcribed as polycistronic transcrip‑ tional units. Examples are the fim operon, which codes for biogenesis of type 1 fimbriae in Escherichia coli, and the atp operon, which codes for the FoF1 ATP synthase. We tested the hypothesis that markerless polar mutations could be efficiently engineered using CRISPR/Cas12a in these loci.Results: Cas12a‑mediated engineering of a terminator sequence inside the fimA gene occurred with efficiencies between 10 and 80% and depended on the terminator’s sequence, whilst other types of mutations, such as a 97 bp deletion, occurred with 100% efficiency. Polar mutations using a terminator sequence were also engineered in the atp locus, which induced its transcriptional shutdown and produced identical phenotypes as a deletion of the whole atp locus (ΔatpIBEFHAGDC). Measuring the expression levels in the fim and atp loci showed that many supposedly non‑ polar mutants induced a significant polar effect on downstream genes. Finally, we also showed that transcriptional shutdown or deletion of the atp locus induces elevated levels of intracellular ATP during the exponential growth phase.Conclusions: We conclude that Cas12a‑mediated mutagenesis is an efficient simple system to generate polar mutants in E. coli. Different mutations were induced with varying degrees of efficiency, and we confirmed that all these mutations abolished the functions encoded in the fim and atp loci. We also conclude that it is difficult to predict which mutagenesis strategy will induce a polar effect in genes downstream of the mutation site. Furthermore the strategies described here can be used to manipulate the metabolism of E. coli as showcased by the increase in intra‑ cellular ATP in the markerless ΔatpIBEFHAGDC mutant.
8.
Pakharukova, Natalia, et al.
(author)
Archaic chaperone-usher pili self-secrete into superelastic zigzag springs
2022
In: Nature. - : Nature Publishing Group. - 0028-0836 .- 1476-4687. ; 609:7926, s. 335-340
Journal article (peer-reviewed) abstract
Adhesive pili assembled via the chaperone-usher pathway (CUP) are hair-like appendages that mediate host tissue colonization and biofilm formation of Gram-negative bacteria 1-3. Archaic CUP pili, the most diverse and widespread CUP adhesins, are promising vaccine and drug targets due to their prevalence in the most troublesome multidrug-resistant (MDR) pathogens 1,4,5. However, their architecture and assembly-secretion process remain unknown. Here, we present the 3.4 Å resolution cryo-electron microscopy structure of the prototypical archaic Csu pilus that mediates biofilm formation of Acinetobacter baumannii, a notorious MDR nosocomial pathogen. In contrast to the thick helical tubes of the classical type 1 and P pili, archaic pili assemble into a conceptually novel ultrathin zigzag architecture secured by an elegant clinch mechanism. The molecular clinch provides the pilus with high mechanical stability as well as superelasticity, a property observed now for the first time in biomolecules, while enabling a more economical and faster pilus production. Furthermore, we demonstrate that clinch formation at the cell surface drives pilus secretion through the outer membrane. These findings suggest that clinch-formation inhibitors might represent a new strategy to fight MDR bacterial infections.
9.
Zlatkov, Nikola, 1985-, et al.
(author)
Unconventional Cyclic di-GMP Signaling in Escherichia coli
2020. - 1
In: Microbial Cyclic Di-Nucleotide Signaling. - Chambridge : Springer International Publishing. - 9783030333072 - 9783030333089 ; , s. 487-517
Book chapter (other academic/artistic) abstract
The species Escherichia coli represents an unfathomable variety of commensal, pathogenic, and environmental strains. The conventional cyclic di-GMP signaling in E. coli controls sessility-motility changes linked to commensalism and/or pathogenicity. Extraintestinal Pathogenic E. coli (ExPEC) are “commensals” that can cause an array of infections outside the gastrointestinal tract. To accommodate their pathogenic lifestyle with the commensal one, ExPEC biology is shaped not only by the presence of specific virulence genes and pathoadaptive mutations but also by regulatory adaptations. Bioinformatic and genetic studies indicate that the cyclic di-GMP signaling network is included in the adaptation process. For example, some neuroinvasive ExPEC were found to maintain reduced cyclic di-GMP levels due to RpoS deactivation, resulting in loss of appearance of the rugose morphotype. Moreover, ExPEC has a diversified repertoire of cyclic di-GMP degrading enzymes obtained by acquisition of novel genes often associated with fimbrial biogenesis gene clusters (e.g., sfaY/papY/focY) and by modification or deletion of specific core genome genes. For example, the majority of ExPEC contains a shortened allelic variant of the ycgG gene and some ExPEC strains do not even carry the genetic locus. New combinations of regulators offer a new cyclic di-GMP platform for S-fimbrial biogenesis and for new metabolic capabilities leading to citrate utilization and ferric citrate uptake. In this review, we outline the prerequisites for the unconventional signaling network, the rationale behind its existence in ExPEC, and future perspectives in studies of ExPEC.
10.
Balsalobre, Carlos, et al.
(author)
Release of the type I secreted α-haemolysin via outer membrane vesicles from Escherichia coli
2006
In: Molecular Microbiology. - Hoboken, NJ, United States : Wiley-Blackwell. - 0950-382X .- 1365-2958. ; 59:1, s. 99-112
Journal article (peer-reviewed) abstract
The α-haemolysin is an important virulence factor commonly expressed by extraintestinal pathogenic Escherichia coli. The secretion of the α-haemolysin is mediated by the type I secretion system and the toxin reaches the extracellular space without the formation of periplasmic intermediates presumably in a soluble form. Surprisingly, we found that a fraction of this type I secreted protein is located within outer membrane vesicles (OMVs) that are released by the bacteria. The α-haemolysin appeared very tightly associated with the OMVs as judged by dissociation assays and proteinase susceptibility tests. The α-haemolysin in OMVs was cytotoxically active and caused lysis of red blood cells. The OMVs containing the α-haemolysin were distinct from the OMVs not containing α-haemolysin, showing a lower density. Furthermore, they differed in protein composition and one component of the type I secretion system, the TolC protein, was found in the lower density vesicles. Studies of natural isolates of E. coli demonstrated that the localization of α-haemolysin in OMVs is a common feature among haemolytic strains. We propose an alternative pathway for the transport of the type I secreted α-haemolysin from the bacteria to the host cells during bacterial infections.
