title: The secreted autotransporter toxin (Sat) does not act as a virulence factor in the probiotic Escherichia coli strain Nissle 1917.
abstract: BACKGROUND
  Escherichia coli Nissle 1917 (EcN) is a probiotic used in the treatment of intestinal diseases. Although it is considered safe, EcN is closely related to the uropathogenic E. coli strain CFT073 and contains many of its predicted virulence elements. Thus, it is relevant to assess whether virulence-associated genes are functional in EcN. One of these genes encodes the secreted autotransporter toxin (Sat), a member of the serine protease autotransporters of Enterobacteriaceae (SPATEs) that are secreted following the type V autotransporter pathway. Sat is highly prevalent in certain E. coli pathogenic groups responsible for urinary and intestinal infections. In these pathogens Sat promotes cytotoxic effects in several lines of undifferentiated epithelial cells, but not in differentiated Caco-2 cells.
  
  
  RESULTS
  Here we provide evidence that sat is expressed by EcN during the colonization of mouse intestine. The EcN protein is secreted as an active serine protease, with its 107 kDa-passenger domain released into the medium as a soluble protein. Expression of recombinant EcN Sat protein in strain HB101 increases paracellular permeability to mannitol in polarized Caco-2 monolayers. This effect, also reported for the Sat protein of diffusely adherent E. coli, is not observed when this protein is expressed in the EcN background. In addition, we show that EcN supernatants confer protection against Sat-mediated effects on paracellular permeability, thus indicating that other secreted EcN factors are able to prevent barrier disruption caused by pathogen-related factors. Sat is not required for intestinal colonization, but the EcNsat::cat mutant outcompetes wild-type EcN in the streptomycin-treated mouse model. Analysis of the presence of sat in 29 strains of the ECOR collection isolated from stools of healthy humans shows 34.8 % positives, with high prevalence of strains of the phylogenetic groups D and B2, related with extra-intestinal infections.
  
  
  CONCLUSIONS
  Sat does not act as a virulence factor in EcN. The role of Sat in intestinal pathogenesis relies on other genetic determinants responsible for the bacterial pathotype.
authors.affiliation: Departament de Bioquímica i Biología Molecular, Institut de Biomedicina de la Universitat de Barcelona, Facultat de Farmàcia, Universitat de Barcelona, E-08028, Barcelona, Spain. lorena-viviana@hotmail.com.
authors.forename: Lorena
authors.initials: L
authors.lastname: Toloza
authors.affiliation: Departament de Bioquímica i Biología Molecular, Institut de Biomedicina de la Universitat de Barcelona, Facultat de Farmàcia, Universitat de Barcelona, E-08028, Barcelona, Spain. rgimenez@ub.edu.
authors.forename: Rosa
authors.initials: R
authors.lastname: Giménez
authors.affiliation: Departament de Bioquímica i Biología Molecular, Institut de Biomedicina de la Universitat de Barcelona, Facultat de Farmàcia, Universitat de Barcelona, E-08028, Barcelona, Spain. mjfabrega@ub.edu.
authors.forename: María Jose
authors.initials: MJ
authors.lastname: Fábrega
authors.affiliation: Departament de Bioquímica i Biología Molecular, Institut de Biomedicina de la Universitat de Barcelona, Facultat de Farmàcia, Universitat de Barcelona, E-08028, Barcelona, Spain. gushi_2@hotmail.com.
authors.forename: Carina Shianya
authors.initials: CS
authors.lastname: Alvarez
authors.affiliation: Departament de Bioquímica i Biología Molecular, Institut de Biomedicina de la Universitat de Barcelona, Facultat de Farmàcia, Universitat de Barcelona, E-08028, Barcelona, Spain. laguilera@ub.edu.
authors.forename: Laura
authors.initials: L
authors.lastname: Aguilera
authors.affiliation: Departament de Bioquímica i Biología Molecular, Institut de Biomedicina de la Universitat de Barcelona, Facultat de Farmàcia, Universitat de Barcelona, E-08028, Barcelona, Spain. mari_canas@ub.edu.
authors.forename: María Alexandra
authors.initials: MA
authors.lastname: Cañas
authors.affiliation: Departament de Fisiologia, Facultat de Farmàcia, Universitat de Barcelona, E-08028, Barcelona, Spain. raquelmartin@ub.edu.
authors.forename: Raquel
authors.initials: R
authors.lastname: Martín-Venegas
authors.affiliation: Departament de Bioquímica i Biología Molecular, Institut de Biomedicina de la Universitat de Barcelona, Facultat de Farmàcia, Universitat de Barcelona, E-08028, Barcelona, Spain. josefabadia@ub.edu.
authors.forename: Josefa
authors.initials: J
authors.lastname: Badia
authors.affiliation: Departament de Bioquímica i Biología Molecular, Institut de Biomedicina de la Universitat de Barcelona, Facultat de Farmàcia, Universitat de Barcelona, E-08028, Barcelona, Spain. lbaldoma@ub.edu.
authors.forename: Laura
authors.initials: L
authors.lastname: Baldomà
mesh_terms: D000818:Animals; D001427:Bacterial Toxins / Q000378:metabolism*; D018938:Caco-2 Cells; D002470:Cell Survival; D004926:Escherichia coli / Q000145:classification / Q000378:metabolism*; D004927:Escherichia coli Infections / Q000378:metabolism / Q000382:microbiology / Q000662:veterinary*; D029968:Escherichia coli Proteins / Q000378:metabolism*; D005243:Feces / Q000382:microbiology; D006367:HeLa Cells; D006801:Humans; D007410:Intestinal Diseases / Q000378:metabolism / Q000382:microbiology* / Q000662:veterinary; D051379:Mice; D010802:Phylogeny
chemical_list: D001427:Bacterial Toxins; D029968:Escherichia coli Proteins; C506813:secreted autotransporter toxin, E coli