Impact of Plant Secondary Metabolites on the Soil Microbiota

Abstract

This study revealed the impact of 2-benzoxazolinone (BOA), gramine, and quercetin on the microbiota in the bulk soil (Cologne agricultural soil) and investigated direct interactions of the plant secondary metabolites and their degradation products with soil bacteria and other plants. Only bacterial DNA from the soil could be analyzed because fungal and oomycetal DNA was below the detection limit. The absence of fungi and oomycetes from Cologne agricultural soil can be explained by the lack of initial carbon content as the soil had not been used for agriculture for over 15 years. The changes in the bacterial microbiota were followed by next-generation sequencing of soil DNA. In addition, surviving bacterial strains were isolated and characterized after treatment with BOA, gramine, or quercetin.<br /> BOA mainly exhibited its function by deterring potentially unfavorable bacterial strains in the bulk soil microbiome resulting in the decrease in 11 amplicon sequence variants (ASVs) and an increase in 10 ASVs. While no bacterial strains were isolated that could degrade BOA, the known BOA-derived detoxification substances N-(2-hydroxy-5-nitrophenyl) acetamide (5-N-AAP), and the BOA-OH isomers BOA-4-OH, BOA-5-OH, BOA-6-OH, and BOA-7-OH revealed their allelopathic character in interactions with A. thaliana and Z. mays. The presence of 5-N-AAP caused an increased expression of geranyllinalool synthase, which is known to produce geranyllinalool, a potential defense compound interfering with sphingolipid metabolism in herbivores. Treatment of Z. mays roots with BOA-OH isomers on the other hand initiated the expression of ROS-related genes indicating the necessity of superoxide radical detoxification of the BOA-derived products. <br /> Gramine exhibited its function in shaping the soil microbiota by mainly attracting potentially beneficial bacteria. While only five ASVs were decreased after gramine treatment, a total of 35 ASVs were increased in abundance. Therefore, gramine may serve as an additional carbon source for the bacteria in the soil. Indeed, one Arthrobacter strain was able to metabolize gramine via the degradation products indole-3-carboxaldehyde (I3A) and indole-3-carboxylic acid (I3C). The auxin-like compound I3A was also extracted two days after gramine application to bulk soil, and it is therefore available for interactions with other organisms including plants. Co-cultivation of A. thaliana with I3A resulted in an improvement in plant growth demonstrating its beneficial properties. <br /> The treatment of bulk soil with the flavonoid quercetin resulted in an increase of the number of 35 ASVs but also a decrease of 17 ASVs. Therefore, quercetin might function by attracting beneficial bacteria, but it is also capable of shaping the soil microbiota by deterring potentially pathogenic strains. Two Arthrobacter strains were identified that were able to degrade quercetin via protocatechuic acid and to use quercetin as a carbon source. Protocatechuic acid was thus identified as the toxic breakdown product of quercetin. Fatty acid analysis of bacterial strains that were able to degrade quercetin, revealed a decrease of the main fatty acid 15:0 anteiso with a subsequent increase of longer-chained fatty acids. This alteration in the fatty acid composition is presumably associated with a decrease in membrane rigidity. A less rigid bacterial membrane might be necessary for the bacterial strains to cope with the quercetin degradation product protocatechuic acid, which is known to cause oxidative stress in other organisms. <br /> This work thus demonstrates the complex roles of plant secondary metabolites during interactions with soil bacteria and plants. Further studies are required to decipher the functions of the metabolites on the different organisms.