Effects of Bacillus subtilis T6-1 on the Rhizosphere Microbial Community Structure of Continuous Cropping Poplar

Abstract

The continuous cropping obstacles in poplar cultivation cause declines in wood yield and serious soil-borne diseases, mainly because of structural alterations in the microbial community and the aggregation of pathogenic fungi. Bacillus subtilis T6-1, isolated from poplar rhizospheric soil, has strong antagonistic effects on poplar pathogens. We aimed to investigate the effects of B. subtilis T6-1 on the structure of the microbial community in the poplar rhizosphere. Poplar seedlings were replanted in three successive generations of soil. The diameter at breast height, plant height, and the number of culturable bacteria of the poplars inoculated with T6-1 exceeded those in the non-inoculated control group. qPCR analysis revealed that the total abundance of T6-1 bacteria in the treated poplars was remarkably higher in contrast to that in the control group. Illumina MiSeq sequencing was employed to track the alterations in diversity and structure of the total microbial community in the poplar rhizosphere inoculated with B. subtilis T6-1. Fungal diversity and abundance in the T6-1 rhizosphere were remarkably lower in contrast with those in the control rhizosphere. The proportion of Bacillus sp. in the total bacterial community in the T6-1 and control groups was 3.04% and 2.38%, respectively, while those of the Rhizoctonia sp. was 2.02% and 5.82%, respectively. In conclusion, B. subtilis T6-1 has the potential to serve as a microbial agent, enhancing the structure of the rhizosphere microbial community as well as promoting tree growth in poplar cultivation.

Keywords: Bacillus subtilis; continuous cropping obstacles; microbiology community structure; poplar; rhizosphere.

Figure 1

Rare-faction curves of (a) communities of bacteria and (b) fungi. The rare-faction curves exhibiting influences of a dissimilarity cut-off of 3% on OTUs numbers uncovered in the study groups. The T6-1 group consisted of poplar trees, which were exposed to bacterial zymotic fluid with T6-1. The CK group consisted of poplar trees, which were exposed to an equivalent volume of sterile zymotic fluid. We analyzed three replicates for every treatment.

Figure 2

Shannon curves of communities of (a) bacteria and (b) fungi. Shannon curves, illustrating the influences of a dissimilarity cut-off of 3% on OTUs numbers uncovered in study groups. The T6-1 group consisted of poplar trees, which were exposed to bacterial zymotic fluid with T6-1. The CK group consisted of poplar trees, which were exposed to an equivalent volume of sterile zymotic fluid. We analyzed three replicates for every treatment.

Figure 3

Structures of communities of bacteria and fungi in study groups (a) Phylum level bacterial relative abundance (b) Genus level bacterial relative abundance (c) Phylum level fungal relative abundance (d) Genus level fungal relative abundance. Stacked bar graphs illustrating major genera relative abundances. The T6-1 group consisted of poplar trees, which were exposed to bacterial zymotic fluid with T6-1. The CK group consisted of poplar trees, which were exposed to an equivalent volume of sterile zymotic fluid. We analyzed three replicates for every treatment. The non-metric multidimensional scaling (NMDS) plot on OTU level could be found in Supplementary Figure S1.

Figure 4

Bacterial and fungal distribution hierarchical clustering heatmaps (a) Phylum level bacterial relative abundance (b) Genus level bacterial relative abundance (c) Phylum level fungal relative abundance. (d) Genus level fungal relative abundance. The heatmaps illustrating distribution of diverse communities from the two groups at the genus level. The T6-1 group consisted of poplar trees, which were exposed to bacterial zymotic fluid containing T6-1. The CK group consisted of poplar trees, which were exposed to an equivalent volume of sterile zymotic fluid. We analyzed three replicates for every treatment.

Figure 5

Venn diagram exhibiting unique, as well as shared (a) bacterial OTUs (b) fungal OTUs for the two treatments. The T6-1 group included poplar trees, which were exposed to bacterial zymotic fluid with T6-1. The CK group consisted of poplar trees, which were exposed to an equivalent volume of sterile zymotic fluid. We analyzed three replicates for every treatment.