Interaction of a genetically modified Pseudomonas fluorescens with the soil-feeding earthworm Octolasion cyaneum (Lumbricidae)

Abstract

The geophagous earthworm Octolasion cyaneum was maintained in microcosms for up to 14 d in the presence of a genetically-modified microorganism (GEMMO), Pseudomonas fluorescens KTG. The GEMMO contained a marker cassette, which was inserted into the chromosome, consisting of the genes coding for kanamycin and gentamycin resistance and also a cryIVB sequence. Plate counts of P. fluorescens KTG were higher in the burrow wall on day 2, and lower on days 7 and 14 than those in the unworked bulk soil. Numbers of P. fluorescens KTG were consistently significantly lower in cast material than in the unworked soil. Counts for total bacteria revealed no significant differences between bulk soil, burrow wall and casts. When earthworms were fed on soil containing P. fluorescens KTG, the population size of the GEMMO declined progressively on passage from the foregut to the hindgut, then increased slightly in the casts relative to the hindgut. However counts in fresh casts were still significantly lower than the corresponding uningested soil. Populations of P. fluorescens KTG in casts increased by up to approximately 10-fold over the first 2 d of the ageing period. Thereafter, plate counts of the GEMMO were slightly less than the corresponding soil kept under the same conditions, showing a similar rate of decline over the 50-d period. Total bacterial plate counts in the aged casts increased by approximately 25-fold during the first 2 d of incubation, subsequently declining whilst remaining significantly higher than the total bacterial plate counts in the corresponding soil which remained relatively constant throughout the experiment. Following a single exposure of the earthworms to the GEMMO, counts of the modified bacterium were detected in casts for upto 15 d. The interactions between bacteria and earthworms are discussed in relation to the potential for dispersal of GEMMOs by soil invertebrates.