Simultaneous oligonucleotide probe hybridization and immunostaining for in situ detection of Gordona species in activated sludge

Abstract

Oligonucleotide probes targeting ribosomal RNA can be designed with high specificity to target microbial populations at different phylogenetic levels if the cellular abundance of ribosomal RNA is sufficiently high. In contrast, polyclonal antibody probes cannot be produced with the same specificity for various phylogenetic groups, but they have the potential to detect slow growing microorganisms, populations with low metabolic activities, or even non-viable cells. We combined a polyclonal antibody stain with a ribosomal RNA targeted oligonucleotide probe for the single cell detection of species of the genus Gordona. Gordona species typically require long generation times, often exhibit filamentous growth, and are commonly encountered in activated sludge foams. Our results suggest that the ribosomal RNA content of individual cells of Gordona in activated sludge is highly variable. Therefore, the combined use of an immunostain and an oligonucleotide probe targeting ribosomal RNA can determine the identity of single cells and provide an approximation of their activity. This approach should result in improved detection limits while maintaining high specificity. z 1999 Federation of European Microbiological Societies.