Measuring rhizosphere effects of two tree species in a temperate forest: A comprehensive method comparison

Rhizosphere effects have been increasingly recognized to play an important role in soil biogeochemical cycling. However, various methods were adopted to sample rhizosphere vs. bulk soils and to measure rhizosphere effects. Here we provide a comprehensive comparison of three commonly used methods to measure rhizosphere effects of two tree species (Larix principis-rupprechtii and Pinus sylvestris var. mongolica) in a temperate forest. Specifically, we used two root chamber methods with non-native C-4 soil and native C-3 soil (C-4 and C-3 soil chamber) and the adhering soil method (C-3 soil natural). The rhizosphere effects ranged from negative to neutral and positive, and varied greatly among methods and variables, but not between species. Generally, the two root chamber methods showed negative or neutral rhizosphere effects, while the adhering soil method often indicated positive or neutral rhizosphere effects. For example, rhizosphere effects of Larbc and Pinus on soil carbon mineralization (C-min, based on 30-day lab incubation of sieved soils) ranged from - 25% to 17% for the two chamber methods, while from 13% to 87% for the C-3 soil natural method. However, the non-destructive isotopic tracing method using the C-4 soil chambers (by trapping CO2 from the intact root-soil system) showed the highest rhizosphere effects on C-min (162% for Larbc and 42% for Pinus). Such differences in rhizosphere effects among methods could be due to a few factors, including different degrees of soil drying by root water uptake, different definitions of rhizosphere, and different levels of perturbation of the root-soil system during sampling and measurement. Further, we found positive correlations in rhizosphere effects between soil properties and microbial biomass and enzyme activities across three methods and two species, but rhizosphere effects on C-min were only positively correlated with rhizosphere effects on soil total nitrogen. Taken together, we suggest researchers to use both root chamber and adhering soil method together, better control soil moisture and incubation time for root chamber method, and quantify root traits in relation to rhizosphere effects for both methods to deepen our understanding of rhizosphere effects of woody plants in future work.