Untersuchungen zur Induktion von Adhäsion bei Escherichia coli und Salmonella Typhimurium durch murines SPLUNC1 und LPLUNC1

The PLUNC (palate, lung and nasal epithelium clone)-proteins are primarily expressed in the airways and the nasopharynx. They belong to the LT (lipid transfer)/LBP (Lipopolysaccharide binding protein)-protein family and are subdivided into long (L)- and short (S)-PLUNCs (Bingle and Craven 2002, Bingle et al. 2004). SPLUNC1 and LPLUNC1 are the only members of the PLUNC-protein family which are expressed in the upper airways (trachea and bronchia, Bingle und Bingle 2000, Bingle et al. 2010). In this study the influence of these two proteins on the adhesion of different gram-negative bacteria was investigated. For this reason, recombinant murine SPLUNC1 and LPLUNC1 from insect cells was used in a crystal violet-adhesion assay. It was shown that the tested PLUNC-proteins inhibit adhesion of most Klebsiella pneumoniae-isolates and of Serratia marcescens. In Salmonella Typhimurium and Escherichia coli, however, adhesion by type 1 pili was induced in the presence of mSPLUNC1 or mLPLUNC1. A possible receptor for PLUNC-proteins on the bacterial surface is LPS, as addition of exogenous LPS inhibits induction of adhesion in the adhesion assays. By the use of truncated variants of SPLUNC1 it was shown that the adhesion inducing domain is located at the protein's C-terminus. To identify genes involved in the PLUNC-induced adhesion a collection of random S. Typhimurium mutants was analyzed in the context of a crystal violet-adhesion assay. The results of this screening indicate an important role of the sensorkinase ArcB in the formation of type 1 pili. To detect and quantify mSPLUNC1 in following studies a monoclonal α-mSPLUNC1-antibody and a polyclonal α-mSPLUNC1-antiserum were generated. Based on these two antibodies a Sandwich-ELISA was established that allows quantification of mSPLUNC1 with a detection limit of 500 pg/ml.