SINGLE-CELL RNA-CONTENT OF NATURAL MARINE PLANKTONIC BACTERIA MEASURED BY HYBRIDIZATION WITH MULTIPLE 16S RIBOSOMAL-RNA-TARGETED FLUORESCENT-PROBES

Despite the potential of cellular RNA as a tool for the study of bacterial activities in natural environments, cellular RNA has rarely been measured because of methodological limitations. In a previous study, we quantitatively analyzed single-cell RNA content with multiple 16S rRNA-targeted fluorescent probes. In the present study, we explore the potential of this approach with 12 samples of natural planktonic bacteria collected from coastal water over a 6-month period (late winter to summer). We measured fluorescence from single cells hybridized with multiple probes, cellular DNA and RNA by ethidium bromide (EtBr) fluorometry, and [H-3]thymidine (TdR) incorporation rates. Fluorescence of the probe-hybridized cells was highly correlated to cellular RNA determined by EtBr fluorometry. Winter and summer samples showed different patterns of fluorescence-frequency distribution. Temperature had a large positive effect on TdR incorporation rates, but there was a negative correlation with cellular RNA content; only the samples taken at elevated summer temperatures showed a positive relationship between TdR incorporation and cellular RNA content. We estimated that the RNA content of the cells not visibly labeled with five probes was <0.3 fg.