Interactions in the rhizosphere: Plant responses to bacterivorous soil protozoa

Plant growth promotion by bacterivorous soil protozoa is generally assigned to an improved nitrogen supply due to the mobilisation of nitrogen fixed in bacterial biomass. However, there is evidence that protozoa may also stimulate plant growth by non-nutrient effects with the phytohormone auxin (indole-3-acetic acid; IAA) being likely involved. This PhD Thesis was performed to investigate morphological, physiological and transcriptional responses of plants to soil protozoa and to assess the involvement of nitrogen and plant hormones in the protozoa-induced plant growth promotion. In the first experiment (Chapter 2) modifications of root architecture and internal auxin metabolism of Lepidium sativum and Arabidopsis thaliana due to the presence of a diverse soil bacterial community and the protozoan species Acanthamoeba castellanii were analysed. Soil bacteria enhanced concentrations of conjugated IAA in L. sativum shoots without affecting root architecture, whereas soil bacteria plus A. castellanii increased free bioactive IAA concentrations and root branching. The results indicate that soil protozoa stimulate root foraging via affecting plant internal modifications of auxin metabolism and thus enable enhanced nutrient capture and plant growth. However, despite increased root branching, A. thaliana reporter plants for auxin and cytokinin did not respond to auxin but to cytokinin. Since soil protozoa increased nitrate concentrations in the rhizosphere the results suggest that nitrate caused an accumulation of cytokinin in the plant and interacted with its hormonal antagonist auxin, which finally induced increased root branching. In the second experiment (Chapter 3) a defined laboratory system using A. thaliana as model plant was designed allowing to investigate effects of interactions between a diverse soil bacterial community and A. castellanii on plant performance in detail. Soil bacteria and protozoa increased growth of A. thaliana already three days past plant inoculation (dpi) with the effects of protozoa exceeding those of bacteria only. The immediate growth response was accompanied by an increased carbon but not nitrogen allocation. However, three days later protozoa enhanced ammonium availability and plant uptake of nitrogen from organic material, which presumably was responsible for prolonged vegetative growth and increased seed production. The results suggest that A. thaliana sensed the upcoming mobilization of nitrogen presumably by changes in rhizosphere signalling and initiated carbon fixation and root carbon allocation which payed off later by increased nutrient capture and strongly increased plant reproduction. In the third experiment (Chapter 4) transcriptional changes of A. thaliana genes involved in plant signalling and stress response as well as nitrogen responsive genes were investigated by performing a DNA array and quantitative real time PCR. Nitrogen responsive genes were not immediately regulated by soil protozoa, but later ammonium responsive genes were up-regulated supporting the results obtained in the experiment reported in Chapter 3. Transcription analysis further demonstrated that soil protozoa down-regulate defence mechanisms in plant roots, but induce plant defence in plant shoots. This suggests that soil protozoa inhibit detrimental soil bacteria by selective grazing leading to a reduced defence in roots and thus reduced investment in secondary metabolite production. Improved nutrient and energy status of A. thaliana may be responsible for increased shoot growth in presence of protozoa despite plant defence concurrently being enhanced. Overall, the results suggest that the effect of protozoa on plant growth in fact initially may not be caused by increased nitrogen availability. Rather, the plants appear to anticipate the subsequent up-coming nitrogen mobilization due to changes in rhizosphere signalling and increase carbon assimilation and allocation to roots resulting in strongly increased plant growth and seed production, i.e. plant fitness. Further, protozoa-mediated reduction in detrimental bacteria may have contributed to increased plant growth by saving costs for secondary metabolite production. Notably, the induction of plant defence in shoots by protozoa was not associated with reduced plant growth but rather the opposite, suggesting that due to increasing nitrogen supply protozoa enable plants to invest in defence in shoots and in parallel increase plant growth and reproduction.