In situ detection of a virulence factor mRNA and 16S rRNA inListeria monocytogenes

Michael Wagner, Michael Schmid, Stefan Juretschko, Karl-Heinz Trebesius, Andreas Bubert, Werner Goebel, Karl-Heinz Schleifer
1998 FEMS Microbiology Letters  
Simultaneous in situ analysis of the structure and function of bacterial cells present within complex communities is a key for improving our understanding of microbial ecology. A protocol for the in situ identification of Listeria spp. using fluorescently tagged, rRNA-targeted oligonucleotide probes was developed. Ethanol fixation and enzymatic pretreatment with lysozyme and proteinase K were used to optimize whole cell hybridization of exponential phase and stationary phase Listeria spp.
more » ... In parallel, transcript probes carrying multiple digoxigenin molecules were combined with anti-digoxigenin Fab antibody fragments labeled with horseradish peroxidase to detect, via the catalytic deposition of fluorescein-tyramide, the iap-mRNA in single Listeria monocytogenes cells. The iap gene encodes the associated virulence factor p60. Application of the new signal amplification technique resulted in strong signals comparable in intensity to those obtained with fluorescently labeled rRNAtargeted oligonucleotide probes. z 1998 Federation of European Microbiological Societies. Published by Elsevier Science B.V.
doi:10.1111/j.1574-6968.1998.tb12906.x pmid:9495027 fatcat:kzbwf266tnhltliheyuoyium7e