Modification of spatial distribution of 2,4-dichlorophenoxyacetic acid degrader microhabitats during growth in soil columns

Abstract

Bacterial processes in soil, including biodegradation, require contact between bacteria and substrates. Knowledge of the three-dimensional spatial distribution of bacteria at the microscale is necessary to understand and predict such processes. Using a soil microsampling strategy combined with a mathematical spatial analysis, we studied the spatial distribution of 2,4-dichlorophenoxyacetic acid (2,4-D) degrader microhabitats as a function of 2,4-D degrader abundance. Soil columns that allowed natural flow were percolated with 2,4-D to increase the 2,4-D degrader abundance. Hundreds of soil microsamples (minimum diameter, 125 microm) were collected and transferred to culture medium to check for the presence of 2,4-D degraders. Spatial distributions of bacterial microhabitats were characterized by determining the average size of colonized soil patches and the average number of patches per gram of soil. The spatial distribution of 2,4-D degrader microhabitats was not affected by water flow, but there was an overall increase in colonized patch sizes after 2,4-D amendment; colonized microsamples were dispersed in the soil at low 2,4-D degrader densities and clustered in patches that were more than 0.5 mm in diameter at higher densities. During growth, spreading of 2,4-D degraders within the soil and an increase in 2,4-D degradation were observed. We hypothesized that spreading of the bacteria increased the probability of encounters with 2,4-D and resulted in better interception of the degradable substrate. This work showed that characterization of bacterial microscale spatial distribution is relevant to microbial ecology studies. It improved quantitative bacterial microhabitat description and suggested that sporadic movement of cells occurs. Furthermore, it offered perspectives for linking microbial function to the soil physicochemical environment.

FIG. 1.

Principles of the spatial analysis. (A) Distribution of soil patches colonized by bacteria in a two-dimensional grid with an indication of the four sizes of microsamples used. (B) Same distribution after the test for the presence of bacteria. The black and white elementary units represent positive and negative results, respectively. (C) Corresponding curves obtained after sampling, showing the percentages of positive microsamples as a function of the four microsample sizes. Different types of distribution of bacteria are shown in rows a, b, and c.

FIG. 2.

Experimental results of the spatial analysis: percentages of microsamples that were positive for the presence of 2,4-D degraders as a function of microsample size for the different column treatments. S, original soil; C, soils from control columns; IP1, IIP2, and IIIP3, soils after one, two, and three pulses of 2,4-D, respectively. The suffixes a, b, c, d, and e indicate replicates of the same treatment.

FIG. 3.

Theoretical distributions compatible with experimental data. Each experiment is represented by an area corresponding to the range of compatible theoretical distributions. Nonoverlapping areas indicate that theoretical distributions are significantly different (P ≤ 0.025). (A) Percentage of positive elementary units in the grid simulating the soil as a function of the diameter of soil patches colonized by 2,4-D degraders. The order of magnitude of 2,4-D degrader densities per gram of soil are indicated next to the results. (B) Number of colonized patches per gram of soil as a function of the diameter of soil patches colonized by 2,4-D degraders. The percentages of 2,4-D degraders are indicated next to the results. S, original soil; C, soils from control columns; IP1, IIP2, and IIIP3, soils after one, two, and three pulses of 2,4-D, respectively. The suffixes a, b, c, d, and e indicate replicates of the same treatment.