Resilience of a soil-established, genetically modified Sinorhizobium meliloti inoculant to soil management practices

Abstract

Microcosm studies were conducted in the greenhouse to evaluate management strategies for eliminating a genetically modified bacterial inoculant from soil. Sinorhizobium meliloti L33, genetically tagged by the chromosomal insertion of a luciferase ( luc) gene, was inoculated into soil seeded with its leguminous host plant, alfalfa ( Medicago sativa). Within 10 weeks the titer decreased from 10 6 cfu g −1 soil to approximately 10 4 cfu g −1 soil, a level which was maintained without significant deviation until the end of the experiment after 83 weeks. At the point of equilibration, six different treatments were initiated: crop rotation to clover and rye as non-host plants, inoculation with two competitor strains—the non-modified parental strain S. meliloti 2011 or with S. meliloti F16, a strain previously isolated from the experimental soil—and the application of two pesticides, intending to eliminate either the host plant by Roundup ® or the rhizobia by exploiting their heavy metal sensitivity towards copper. The efficiency of these treatments was studied and compared by monitoring the titer of S. meliloti L33 in bulk soil, rhizospheres and in alfalfa root nodules. In bulk soil, only the application of Roundup ® resulted in a significant reduction of S. meliloti L33 to 4.5×10 1 cfu g −1 soil while all other treatments had no significant effect. However, in rhizospheres of clover and rye the concentrations of S. meliloti were approximately 300-fold lower than those found in the rhizosphere of alfalfa after 57 weeks of growth. Additionally, analysis of the alfalfa nodule occupancy revealed a partial replacement of S. meliloti L33 by the strain F16, indicating that cell numbers of the inoculants were not decisive but their competitiveness was. Thus, crop rotation and inoculation with a superior competitor affected S. meliloti L33 on different levels, but only the removal of the host plant by application of a herbicide was an efficient means to reduce the strain in soil.