A standardized and miniaturized method to investigate rhizosphere microorganisms, with a focus on methanogenic archaea and methanotrophic bacteria

Abstract

Plant-microorganism interactions and associations play a fundamental role in terrestrial ecosystems and mainly occur in the rhizosphere, an interaction zone around plant roots. However, the definitions and handling strategies described in the literature differ considerably, and the aim of the present work was to develop a standardized and miniaturized experimental setup that enables a simple and comparable investigation of the rhizosphere. After methodological optimization steps, we applied and evaluated our method to examine the effects of plants and increased methane (CH4) concentrations on the abundance and activity of rhizosphere microorganisms engaged in the CH4-cycle.Seedlings of black radish (Raphanus sativus) and yellow mustard (Sinapis alba) were grown in soil-filled reaction tubes, and the growth conditions were adapted for optimal plant root development. Since plant roots pervaded the entire volume of the soil, we could assume the entire soil to be influenced by the plants and regarded it as rhizosphere soil. Additionally, the application of plant growth chambers allowed to investigate plant and microbial growth at defined headspace compositions. The results demonstrate that bulk and rhizosphere soil differed significantly regarding the dissolved carbon and nitrogen contents. The abundances of cultivable microorganisms (CFU) and of methanotrophic bacteria (type Ia) were significantly increased in rhizosphere soil. Growth under increased CH4 concentration in the headspace revealed an increased abundance of methanotrophic and methanogenic microorganisms in the rhizosphere. A higher capacity to oxidize CH4 in the rhizosphere compared with bulk soil was demonstrated, extending the effect of the influence of plants on microbial activity and confirming the suitability of this approach to study plant-soil-microbe interactions. Since the headspace composition in the plant growth chambers can easily be adjusted, the current setup is also suitable for investigating further gas cycles and the microorganisms involved.