title: Quantification of bacterial invasion into adherent cells by flow cytometry.
abstract: Quantification of invasive, intracellular bacteria is critical in many areas of cellular microbiology and immunology. We describe a novel and fast approach to determine invasion of bacterial pathogens in adherent cell types such as epithelial cells or fibroblasts based on flow cytometry. Using the CEACAM-mediated uptake of Opa-expressing Neisseria gonorrhoeae as a well-characterized model of bacterial invasion, we demonstrate that the flow cytometry-based method yields results comparable to a standard antibiotic protection assay. Furthermore, the quantification of intracellular bacteria by the novel approach is not biased by intracellular killing of the microbes and correctly discriminates between cell-associated extracellular and bona fide intracellular bacteria. As flow cytometry-based quantification is also applicable to other pathogen-host interactions such as the integrin-mediated internalization of Staphylococcus aureus, this approach provides a fast and convenient alternative for the quantification of bacterial uptake and should be particularly useful in elucidating the molecular mechanisms of pathogen-triggered host cell invasion.
authors.affiliation: Zentrum für Infektionsforschung, Universität Würzburg, Röntgenring 11, 97070 Würzburg, Germany.
authors.forename: Stefan
authors.initials: S
authors.lastname: Pils
authors.forename: Tim
authors.initials: T
authors.lastname: Schmitter
authors.forename: Florian
authors.initials: F
authors.lastname: Neske
authors.forename: Christof R
authors.initials: CR
authors.lastname: Hauck
mesh_terms: D000818:Animals; D015703:Antigens, CD / Q000378:metabolism; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics / Q000378:metabolism; D002272:Carcinoembryonic Antigen; D015815:Cell Adhesion Molecules / Q000378:metabolism; D002460:Cell Line; D004847:Epithelial Cells / Q000382:microbiology*; D005347:Fibroblasts / Q000382:microbiology*; D005434:Flow Cytometry / Q000379:methods*; D006801:Humans; D016023:Integrins / Q000378:metabolism; D007668:Kidney / Q000166:cytology* / Q000382:microbiology; D051379:Mice; D009344:Neisseria gonorrhoeae / Q000472:pathogenicity*; D013211:Staphylococcus aureus / Q000472:pathogenicity*
chemical_list: D015703:Antigens, CD; D001425:Bacterial Outer Membrane Proteins; C069847:CD66 antigens; C075729:CEACAM3 protein, human; D002272:Carcinoembryonic Antigen; D015815:Cell Adhesion Molecules; D016023:Integrins; C090348:Opa protein, Neisseria