Supplementary Figure S2

1. Supplementary Figure S2 Box plots showing the mean fluorescence intensities of Nitrospira-like microcolonies in activated sludge (waste water treatment plant of the veterinary medicine, Vienna, Austria) stained by either Cy3-single- or double-labeled probe Ntsp175. The left panel displays the results from 133 microcolonies stained by the single-labeled probe Ntsp175. The right panel shows the data for 94 aggregates stained by the double-labeled probe Ntsp175. All Nitrospira aggregates were simultaneously identified by the genus Nitrospira specific probe Ntspa662 labeled in Cy5. Confocal microscope settings for recording the images were adjusted for the double-labeled probe Ntsp175 and kept constant for recording the intensities of cells stained with the single-labeled probe Ntsp175. Intensities of the microcolonies were measured using the software package daime. Outliers are indicated by black dots. We used the non parametric Mann-Withney-U-Test to test for significant differences in the signal intensities between the hybridizations with single- and double-Cy3-labeled probe Ntsp175, respectively. Signal intensity of the detected microcolonies was significantly higher in the double-labeled hybridization (Mann-Whitney-U test, Z= -7.3, exact significance 2-tailed P>0,0001) as indicated by the asterisk.