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      Intra-operative biopsy in chronic sinusitis detects pathogenic Escherichia coli that carry fimG/H, fyuA and agn43 genes coding biofilm formation

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          Abstract

          The aim of this study was to investigate whether or not surgical biopsy of sinus tissue in chronic sinusitis, not responsive to treatment, would detect E. coli. We intended to evaluate E. coli virulence genes, therefore dispute the causal role of such an unusual microorganism in chronic sinusitis, as well as consider effective pathogen-targeted therapy. Patients with E. coli isolated by intra-operative puncture biopsy were included in the study. Genetic analysis of E. coli isolates, including phylogenetic grouping and virulence factor characteristics, were done by multiplex PCR. We identified 26 patients with chronic sinusitis, in which 26 E. coli isolates were cultured. The E. coli isolates belonged mainly to pathogenic phylogenetic group B2, and carried multiple virulence genes. Three genes in particular were present in all (100%) of examined isolates, they were (1) marker agn43 gene for forming biofilm, (2) type 1 fimbriae ( fimG/H gene) and (3) yersiniabactin receptor ( fyuA). Furthermore, a pseudo-phylogenetic tree of virulence genes distribution revealed possible cooperation between agn43, fimG/H, and fyuA in the coding of biofilm formation. Intra-operative-biopsy and culture-based therapy, targeting the isolated E. coli, coincided with long-term resolution of symptoms. This is the first report demonstrating an association between a highly pathogenic E. coli, chronic sinus infection, and resolution of symptoms upon E. coli targeted therapy, a significant finding due to the fact that E. coli has not been considered to be a commensal organism of the oropharynx or sinuses. We postulate that the simultaneous presence of three genes, each coding biofilm formation, may in part account for the chronicity of E. coli sinusitis.

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          High prevalence of Escherichia coli belonging to the B2+D phylogenetic group in inflammatory bowel disease.

          It is not clear which species of bacteria may be involved in inflammatory bowel disease (IBD). One way of determining which bacteria might be likely candidates is to use culture-independent methods to identify microorganisms that are present in diseased tissues but not in controls. (1) To assess the diversity of microbial communities of biopsy tissue using culture-independent methods; (2) to culture the bacteria found in the tissues of patients with IBD but not in the controls; (3) to identify potential virulence factors associated with cultured bacteria. 84 biopsy specimens were collected from 15 controls, 13 patients with Crohn's disease (CD) and 19 patients with ulcerative colitis (UC) from a population-based case-control study. Ribosomal intergenic spacer analysis (RISA) was conducted to identify unique DNA bands in tissues from patients with CD and UC that did not appear in controls. RISA followed by DNA sequencing identified unique bands in biopsy specimens from patients with IBD that were classified as Escherichia coli. Targeted culture showed a significantly (p<0.05) higher number of Enterobacteriaceae in specimens from patients with IBD. The B2+D phylogenetic group, serine protease autotransporters (SPATE) and adherence factors were more likely to be associated with tissues from patients with UC and CD than with controls. The abundance of Enterobacteriaceae is 3-4 logs higher in tissues of patients with IBD and the B2+D phylogenetic groups are more prevalent in patients with UC and CD. The B2+D phylogenetic groups are associated with SPATE and adherence factors and may have a significant role in disease aetiology.
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            Prevalence of the "high-pathogenicity island" of Yersinia species among Escherichia coli strains that are pathogenic to humans.

            The fyuA-irp gene cluster contributes to the virulence of highly pathogenic Yersinia (Yersinia pestis, Yersinia pseudotuberculosis, and Yersinia enterocolitica 1B). The cluster encodes an iron uptake system mediated by the siderophore yersiniabactin and reveals features of a pathogenicity island. Two evolutionary lineages of this "high pathogenicity island" (HPI) can be distinguished on the basis of DNA sequence comparison: a Y. pestis group and a Y. enterocolitica group. In this study we demonstrate that the HPI of the Y. pestis evolutionary group is disseminated among species of the family Enterobacteriaceae which are pathogenic to humans. It prevails in enteroaggregative Escherichia coli and in E. coli blood culture isolates (93 and 80%, respectively), but is rarely found in enteropathogenic E. coli, enteroinvasive E. coli, and enterotoxigenic E. coli isolates. In contrast, the HPI was absent from enterohemorrhagic E. coli, Shigella, and Salmonella enterica strains investigated. Polypeptides encoded by the fyuA, irp1, and irp2 genes located on the HPI could be detected in E. coli strains pathogenic to humans. However, these E. coli strains showed a reduced sensitivity to the bacteriocin pesticin, whose uptake is mediated by the FyuA receptor. Escherichia strains do not possess the hms gene locus thought to be a part of the HPI of Y. pestis. Deletions of the juA-irp gene cluster affecting solely the fyuA part of the HPI were identified in 3% of the E. coli strains tested. These results suggest horizontal transfer of the HPI between Y. pestis and some pathogenic E. coli strains.
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              The antigen 43 structure reveals a molecular Velcro-like mechanism of autotransporter-mediated bacterial clumping.

              Aggregation and biofilm formation are critical mechanisms for bacterial resistance to host immune factors and antibiotics. Autotransporter (AT) proteins, which represent the largest group of outer-membrane and secreted proteins in Gram-negative bacteria, contribute significantly to these phenotypes. Despite their abundance and role in bacterial pathogenesis, most AT proteins have not been structurally characterized, and there is a paucity of detailed information with regard to their mode of action. Here we report the structure-function relationships of Antigen 43 (Ag43a), a prototypic self-associating AT protein from uropathogenic Escherichia coli. The functional domain of Ag43a displays a twisted L-shaped β-helical structure firmly stabilized by a 3D hydrogen-bonded scaffold. Notably, the distinctive Ag43a L shape facilitates self-association and cell aggregation. Combining all our data, we define a molecular "Velcro-like" mechanism of AT-mediated bacterial clumping, which can be tailored to fit different bacterial lifestyles such as the formation of biofilms.
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                Author and article information

                Contributors
                Role: ConceptualizationRole: Data curationRole: InvestigationRole: Resources
                Role: ConceptualizationRole: MethodologyRole: Project administrationRole: Writing – review & editing
                Role: Data curationRole: Formal analysisRole: InvestigationRole: MethodologyRole: Validation
                Role: Data curationRole: Formal analysisRole: InvestigationRole: MethodologyRole: SoftwareRole: ValidationRole: Writing – review & editing
                Role: Data curationRole: Formal analysisRole: MethodologyRole: SoftwareRole: ValidationRole: VisualizationRole: Writing – review & editing
                Role: Data curationRole: Formal analysisRole: InvestigationRole: Writing – original draftRole: Writing – review & editing
                Role: Data curationRole: Formal analysisRole: Funding acquisitionRole: MethodologyRole: ResourcesRole: Software
                Role: ConceptualizationRole: Formal analysisRole: ResourcesRole: SupervisionRole: ValidationRole: Writing – review & editing
                Role: Data curationRole: Formal analysisRole: Funding acquisitionRole: InvestigationRole: MethodologyRole: Writing – original draft
                Role: ConceptualizationRole: Formal analysisRole: SupervisionRole: Writing – original draftRole: Writing – review & editing
                Role: Editor
                Journal
                PLoS One
                PLoS ONE
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, CA USA )
                1932-6203
                23 March 2018
                2018
                : 13
                : 3
                : e0192899
                Affiliations
                [1 ] Medical Center MML, Warsaw, Poland
                [2 ] Medical Laboratories, SYNEVO, Warszawa, Poland
                [3 ] Department of Molecular Biotechnology and Microbiology, Faculty of Chemistry, Gdańsk University of Technology, Gdańsk, Poland
                [4 ] Nowicki Institute for Women’s Health Research (Now I for HeR), 114 Governors Way, Brentwood, TN, United States of America
                University of Alabama at Birmingham, UNITED STATES
                Author notes

                Competing Interests: SYNEVO Medical Laboratories provided support in the form of salaries for authors AM and TA, but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript. There is no conflict of interest between the MML Medical Center, SYNEVO Medical Laboratories and Technical School of Gdansk relating to employment, consultancy, patents, products in development, and marketed products. This does not alter our adherence to PLOS ONE policies on sharing data and materials. All authors declare no conflict of interest.

                Author information
                http://orcid.org/0000-0002-3814-3985
                Article
                PONE-D-17-34214
                10.1371/journal.pone.0192899
                5865710
                29570706
                fe28fce0-7c48-436b-b696-cecc04ac4f1d
                © 2018 Michalik et al

                This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.

                History
                : 20 September 2017
                : 18 January 2018
                Page count
                Figures: 1, Tables: 1, Pages: 11
                Funding
                Funded by: Medical Center MML
                Award ID: 2015-01
                Award Recipient :
                This study was supported by research grant from Medical Center MML, Award Number 2015-01 to J. Kur. SYNEVO Medical Laboratories provided support in the form of salaries for authors AM and TA, but did not have any additional role in the study design, data collection and analysis, decision to publish, or preparation of the manuscript.
                Categories
                Research Article
                Biology and Life Sciences
                Microbiology
                Medical Microbiology
                Microbial Pathogens
                Bacterial Pathogens
                Medicine and Health Sciences
                Pathology and Laboratory Medicine
                Pathogens
                Microbial Pathogens
                Bacterial Pathogens
                Medicine and Health Sciences
                Pathology and Laboratory Medicine
                Pathogens
                Virulence Factors
                Medicine and Health Sciences
                Surgical and Invasive Medical Procedures
                Biopsy
                Biology and Life Sciences
                Microbiology
                Bacteriology
                Bacterial Biofilms
                Biology and Life Sciences
                Microbiology
                Biofilms
                Bacterial Biofilms
                Biology and Life Sciences
                Evolutionary Biology
                Evolutionary Systematics
                Phylogenetics
                Biology and Life Sciences
                Taxonomy
                Evolutionary Systematics
                Phylogenetics
                Computer and Information Sciences
                Data Management
                Taxonomy
                Evolutionary Systematics
                Phylogenetics
                Biology and Life Sciences
                Cell Biology
                Cellular Structures and Organelles
                Pili and Fimbriae
                Medicine and Health Sciences
                Pathology and Laboratory Medicine
                Pathogens
                Virulence Factors
                Pathogen Motility
                Pili and Fimbriae
                Medicine and Health Sciences
                Surgical and Invasive Medical Procedures
                Biology and Life Sciences
                Immunology
                Immune Response
                Inflammation
                Medicine and Health Sciences
                Immunology
                Immune Response
                Inflammation
                Medicine and Health Sciences
                Diagnostic Medicine
                Signs and Symptoms
                Inflammation
                Medicine and Health Sciences
                Pathology and Laboratory Medicine
                Signs and Symptoms
                Inflammation
                Custom metadata
                All relevant data are within the paper. There is no supplemental files attached. For Genetic typing files please contact Prof. B. Krawczyk. For patient clinical files contact Dr. Michal Michalik. Queries related to data access may also be submitted directly to the Medical Center MML in Warsaw at: kontakt@ 123456mml.com.pl .

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