Transcriptomic profiling of Bacillus amyloliquefaciens FZB42 in response to maize root exudates

Abstract

Background: Plant root exudates have been shown to play an important role in mediating interactions between plant growth-promoting rhizobacteria (PGPR) and their host plants. Most investigations were performed on Gram-negative rhizobacteria, while much less is known about Gram-positive rhizobacteria. To elucidate early responses of PGPR to root exudates, we investigated changes in the transcriptome of a Gram-positive PGPR to plant root exudates.

Results: Bacillus amyloliquefaciens FZB42 is a well-studied Gram-positive PGPR. To obtain a comprehensive overview of FZB42 gene expression in response to maize root exudates, microarray experiments were performed. A total of 302 genes representing 8.2% of the FZB42 transcriptome showed significantly altered expression levels in the presence of root exudates. The majority of the genes (261) was up-regulated after incubation of FZB42 with root exudates, whereas only 41 genes were down-regulated. Several groups of the genes which were strongly induced by the root exudates are involved in metabolic pathways relating to nutrient utilization, bacterial chemotaxis and motility, and non-ribosomal synthesis of antimicrobial peptides and polyketides.

Conclusions: Here we present a transcriptome analysis of the root-colonizing bacterium Bacillus amyloliquefaciens FZB42 in response to maize root exudates. The 302 genes identified as being differentially transcribed are proposed to be involved in interactions of Gram-positive bacteria with plants.

Figure 1

Composition and concentration of the maize root exudates. Exudates collected from the roots of maize seedlings were assayed by HPLC. Organic acids, amino acids, and carbohydrates were quantified and those which had a concentration of >0.1 μmol g-1(dry weight) were included in the graph. Proline, a known constituent of maize root exudate, was not detected since the derivatization reagent (OPA) used reacts only with primary amino groups.

Figure 2

Number of FZB42 genes altered in transcription in response to root exudates at different exudate concentrations and cell densities. Maize root exudates were supplemented in three concentrations (0.25 mg/ml, 0.5 mg/ml and 1.0 mg/ml) to FZB42 cultures and total RNA was prepared from the bacterial cells harvested at two optical densities (OD₆₀₀ = 1.0 and OD₆₀₀ = 3.0). Genes significantly altered in transcription (q ≤ 0.01 and fold change ≥1.5) by presence of root exudates are represented in the figure.

Figure 3

Overview of groups of the 302 genes altered in transcription by root exudates. A total of 302 genes were significantly altered (q ≤ 0.01 and fold change ≥1.5) in transcription by the maize root exudates. “Up” indicates genes that were up-regulated in presence root exudates, while “down” the ones that were down-regulated by the root exudates. The genes encoding a product with known or unknown function and those encoding a hypothetical protein were indicated. The number of genes of each section and their percentage is depicted.

Figure 4

Distribution in various functional categories of the genes altered in transcription by root exudates. Among the 302 genes altered in transcription by maize root exudates at OD3.0, those with known (189 genes) or putative (44 genes) products were classified according to their function. The percentage of each group is indicated.

Figure 5

Fold-change of differentially expressed genes selected for validation by Real-time PCR. The fold changes revealed by real-time PCR of the selected genes were determined using the software REST. Three repeats were performed for each gene. For comparison, the fold changes obtained in microarray analysis were shown in parenthesis below each specific gene. The boxes represent the distance between the 25th and the 75th percentile. The lines in the boxes represent the median gene expression. Whiskers represent the minimum and maximum observations.

Figure 6

A subset of the up-regulated genes with known function in response to maize root exudates. The significantly up-regulated genes by the root exudates were mapped in the KEGG pathway and the diagram was accordingly adapted. The products encoded by the up-regulated genes were highlighted in red, whilst the down-regulated YadH was highlighted in green. CM stands for cell membrane.