SURVIVAL AND ACTIVITY OF PSEUDOMONAS SP STRAIN B13(FR1) IN A MARINE MICROCOSM DETERMINED BY QUANTITATIVE PCR AND AN RIBOSOMAL-RNA-TARGETINGPROBE AND ITS EFFECT ON THE INDIGENOUS BACTERIOPLANKTON

Genetically engineered Pseudomonas sp. strain B13(FR1) was released in to laboratory-scale marine ecosystem models (microcosms). Survival of the introduced population in the water column and the sediment was det ermined by plating on a selective medium and by quantitative competiti ve PCR. The activity of the released bacteria was determined by in sit u hybridization of single cells with a specific rRNA-targeting oligonu cleotide probe. Two microcosms were inoculated with 10(6) cells ml(-1) , while an uninoculated microcosm served as a control. The number of P seudomonas sp. strain B13(FR1) cells decreased rapidly to ca. 10(2) ce lls ml(-1) within 2 days after the release, which is indicative of gra zing by protozoa. Three days after the introduction into seawater, cel ls were unculturable, but PCR continued to detect cells in low numbers . Immediately after the release, the ribosomal content of Pseudomonas sp. strain B13(FR1) corresponded to a generation time of 2 h. The grow th rate decreased to less than 0.04 h(-1) in 5 days and remained low, probably because of carbon limitation of the cells. Specific amendment of the microcosms with 10 mM 4-chlorobenzoate resulted in a rapid inc rease of the growth rate and an exponentially increasing number of cel ls detected by PCR, but not in resuscitation of the cells to a cultura ble state. The release of Pseudomonas sp. strain B13(FR1) into the mic rocosms seemed to affect only the indigenous bacterioplankton communit y transiently. Effects on the community were also apparent from the ha ndling of water during filling of the microcosms and the amendment wit h 4-chlorobenzoate.