Applicability of non-antibiotic resistance marker genes in ecological studies of introduced bacteria in forest soil

Tagging of an inoculum with marker genes provides a means to monitor the inoculum in the environment in the presence of natural bacteria. In this work a Pseudomonas fluorescens strain originating from the rhizosphere of birch (Betula pendula) was chromosomally tagged with a single copy of a mer-luc marker gene cassette. Survival and activity of this strain were monitored in Finnish forest soil microcosms at different temperatures by determining whole cell and cell extract luminescence, light-producing colonies on selective media containing HgCl2 and by PCR amplification of the luc gene. Monitoring by plate counts was successful using the non-antibiotic resistance marker genes. The luminescence of cell extracts correlated well with plate counts at first, but from day 10 onward, the plate counts were lower than cell numbers estimated by cell extract luminescence. Although PCR detection was less efficient in humus, the lite marker gene was detectable by PCR after 6 months at all temperatures except 30degreesC. Some survival was still observed after a 6-month incubation at temperatures below 4degreesC, representing the prevailing conditions in the boreal region. The inoculum did not cause any detectable prolonged effects on the natural eubacterial community at any temperature examined as determined by denaturing gradient gel electrophoresis. (C) 2001 Federation of European Microbiological Societies. Published by Elsevier Science B.V. All rights reserved.