Co-analysis of cucumber rhizosphere metabolites and microbial PLFAs under excessive fertilization in solar greenhouse

Abstract

Fertilizer application is the most common measure in agricultural production, which can promote the productivity of crops such as cucumbers, but the problem of excessive fertilization occurs frequently in solar greenhouses. However, the effects of fertilization levels on cucumber rhizosphere soil microbes and metabolites and their relationships are still unclear. In order to determine how fertilization levels affect the rhizosphere microenvironment, we set up four treatments in the solar greenhouse: no-fertilization (N0P0K0), normal fertilization (N1P1K1), slight excessive fertilization (N2P2K2), and extreme excessive fertilization (N3P3K3). The results showed that fertilization treatments significantly increased cucumber yield compared to no-fertilization, but, the yield of N3P3K3 was significantly lower than that of N1P1K1 and N2P2K2. Fertilization levels had significant effects on rhizosphere microorganisms, and pH, NH₄ ⁺-N and AP were the main environmental factors that affected the changes in microbial communities. The total PLFAs, the percentages of fungi and arbuscular mycorrhizal fungi (AMF) were significantly reduced and bacteria percentage was significantly increased in N3P3K3 compared to other fertilization treatments. Differential metabolites under different fertilization levels were mainly organic acids, esters and sugars. Soil phenols with autotoxic effect under fertilization treatments were higher than that of N0P0K0. In addition, compared with soil organic acids and alkanes of N0P0K0, N2P2K2 was significantly increased, and N3P3K3 was not significantly different. This suggested that cucumber could maintain microbial communities by secreting beneficial metabolites under slight excessive fertilization (N2P2K2). But under extremely excessive fertilization (N3P3K3), the self-regulating ability of cucumber plants and rhizosphere soil was insufficient to cope with high salt stress. Furthermore, co-occurrence network showed that 16:1ω5c (AMF) was positively correlated with 2-palmitoylglycerol, hentriacontane, 11-octadecenoic acid, decane,4-methyl- and d-trehalose, and negatively correlated with 9-octadecenoic acid at different fertilization levels. This indicated that the beneficial microorganisms in the cucumber rhizosphere soil promoted with beneficial metabolites and antagonized with harmful metabolites. But with the deepening of overfertilization, the content of beneficial microorganisms and metabolites decreased. The study provided new insights into the interaction of plant rhizosphere soil metabolites and soil microbiomes under the different fertilization levels.

Keywords: cucumber; excessive chemical fertilizer; metabolites; microbials PLFAs; rhizosphere; solar greenhouse.

Figure 1

The effects of different fertilization levels on (A) plant images, (B) cucumber yields, (C) leaf photosynthetic pigments, and (D) plant dry weights. Values are means ± SEM (n = 5). Different letters above the bars indicate statistically significant differences at p < 0.05.

Figure 2

The effects of different fertilization levels on soil enzymes activities. Values are means ± SEM (n = 5). Different letters above the bars indicate statistically significant differences at p < 0.05.

Figure 3

The effects of different fertilization levels on soil microbial communities. (A) Heatmap of relative abundance of microbial PLFAs, reddish color indicates increased metabolites content while bluish color indicates decreased metabolites content. (B) Redundancy analysis (RDA) of the rhizosphere microbial PLFAs with soil chemical properties, Significant variables via forward selection are labeled with asterisk (**, and * represent p < 0.01, and 0.05, respectively).

Figure 4

The partial least squares-discriminant analysis (PLS-DA) of soil metabolites under different fertilization levels.

Figure 5

(A) Relative abundance of classified metabolites under different fertilization levels. Values are means ± SEM (n = 5). Different letters above the bars indicate statistically significant differences at p < 0.05. (B) Heatmap of differential metabolites under different fertilization levels, reddish color indicates increased metabolites content while bluish color indicates decreased metabolites content, * represent differential metabolites for three fertilization treatments compared to no-fertilization (VIP > 1 and p < 0.05).

Figure 6

Co-occurrence network of the microbial PLFAs and differential metabolites under different fertilization levels. A connection indicates a strong correlation (spearman correlation analysis, p < 0.05), the stronger the spearman correlation, the thicker the line. The size of each node is proportional to the number of connections, the blue and red lines indicate negative and positive relationships, respectively.