Design and application of oligonucleotide probes for fluorescent in situ identification of the filamentous bacterial morphotype Nostocoida limicola in activated sludge

16S rRNA targeted probes, designed using sequence data from pure cultures o f the three morphotypes of the filamentous bulking bacteria Nostocoida limi cola I, II and III and their successful application to the in situ identifi cation of these bacteria in activated sludge biomass samples are described here. Two probes were required to detect all the sequenced N. limicola II i solates. Results from fluorescent in situ hybridization suggest that the mo rphotypes N. limicola I and II contain at least two phylogenetically unrela ted bacteria. The N. limicola II filaments that did not respond to the prob es designed in this study fluoresced instead with the probes previously des igned for the alpha -Proteobacteria. The data also suggest that both N. lim icola I and III can exist in activated sludge as single, paired or clumped cells and thus in a form not recognizable microscopically as this morphotyp e. Some N. limicola II filaments which responded to the probes designed her e were much thinner than the filaments conventionally 'identified' as this morphotype and better fitted the descriptions often used in the literature for N. limicola I.