title: Investigating the effects of probiotics on pneumococcal colonization using an in vitro adherence assay.
abstract: Adherence of Streptococcus pneumoniae (the pneumococcus) to the epithelial lining of the nasopharynx can result in colonization and is considered a prerequisite for pneumococcal infections such as pneumonia and otitis media. In vitro adherence assays can be used to study the attachment of pneumococci to epithelial cell monolayers and to investigate potential interventions, such as the use of probiotics, to inhibit pneumococcal colonization. The protocol described here is used to investigate the effects of the probiotic Streptococcus salivarius on the adherence of pneumococci to the human epithelial cell line CCL-23 (sometimes referred to as HEp-2 cells). The assay involves three main steps: 1) preparation of epithelial and bacterial cells, 2) addition of bacteria to epithelial cell monolayers, and 3) detection of adherent pneumococci by viable counts (serial dilution and plating) or quantitative real-time PCR (qPCR). This technique is relatively straightforward and does not require specialized equipment other than a tissue culture setup. The assay can be used to test other probiotic species and/or potential inhibitors of pneumococcal colonization and can be easily modified to address other scientific questions regarding pneumococcal adherence and invasion.
authors.affiliation: Pneumococcal Research, Murdoch Childrens Research Institute; eileen.dunne@mcri.edu.au.
authors.forename: Eileen M
authors.initials: EM
authors.lastname: Dunne
authors.affiliation: Allergy & Immune Disorders, Murdoch Childrens Research Institute.
authors.forename: Zheng Q
authors.initials: ZQ
authors.lastname: Toh
authors.affiliation: Department of Otolaryngology, The University of Melbourne.
authors.forename: Mary
authors.initials: M
authors.lastname: John
authors.affiliation: Pneumococcal Research, Murdoch Childrens Research Institute; Department of Microbiology & Immunology at the Peter Doherty Institute for Infection & Immunity, The University of Melbourne.
authors.forename: Jayne
authors.initials: J
authors.lastname: Manning
authors.affiliation: Pneumococcal Research, Murdoch Childrens Research Institute; Department of Microbiology & Immunology at the Peter Doherty Institute for Infection & Immunity, The University of Melbourne.
authors.forename: Catherine
authors.initials: C
authors.lastname: Satzke
authors.affiliation: Allergy & Immune Disorders, Murdoch Childrens Research Institute.
authors.forename: Paul
authors.initials: P
authors.lastname: Licciardi
mesh_terms: D001422:Bacterial Adhesion / Q000187:drug effects*; D002460:Cell Line; D004847:Epithelial Cells / Q000187:drug effects / Q000382:microbiology; D006801:Humans; D009305:Nasopharynx / Q000166:cytology / Q000382:microbiology; D019936:Probiotics / Q000494:pharmacology*; D013291:Streptococcus / Q000502:physiology*; D013296:Streptococcus pneumoniae / Q000166:cytology / Q000187:drug effects* / Q000502:physiology