Natural immunity stimulation using ELICE16INDURES® plant conditioner in field culture of soybean

Abstract

Recently, climate change has had an increasing impact on the world. Innate defense mechanisms operating in plants - such as PAMP-triggered Immunity (PTI) - help to reduce the adverse effects caused by various abiotic and biotic stressors. In this study, the effects of ELICE16INDURES® plant conditioner for organic farming, developed by the Research Institute for Medicinal Plants and Herbs Ltd. Budakalász Hungary, were studied in a soybean population in Northern Hungary. The active compounds and ingredients of this product were selected in such a way as to facilitate the triggering of general plant immunity without the presence and harmful effects of pathogens, thereby strengthening the healthy plant population and preparing it for possible stress effects. In practice, treatments of this agent were applied at two different time points and two concentrations. The conditioning effect was well demonstrated by using agro-drone and ENDVI determination in the soybean field. The genetic background of healthier plants was investigated by NGS sequencing, and by the expression levels of genes encoding enzymes involved in the catalysis of metabolic pathways regulating PTI. The genome-wide transcriptional profiling resulted in 13 contigs related to PAMP-triggered immunity and activated as a result of the treatments. Further analyses showed 16 additional PTI-related contigs whose gene expression changed positively as a result of the treatments. The gene expression values of genes encoded in these contigs were determined by in silico mRNA quantification and validated by RT-qPCR. Both - relatively low and high treatments - showed an increase in gene expression of key genes involving AOC, IFS, MAPK4, MEKK, and GST. Transcriptomic results indicated that the biosyntheses of jasmonic acid (JA), salicylic acid (SA), phenylpropanoid, flavonoid, phytoalexin, and cellular detoxification processes were triggered in the appropriate molecular steps and suggested that plant immune reactions may be activated also artificially, and innate immunity can be enhanced with proper plant biostimulants.

Keywords: Defense mechanism; ELICE16INDURES®; Expression profiling; Glycine max; Soybean.

Graphical abstract

Fig. 1

Soybean plots where data were collected by drone recording. “A” denotes untreated control (1), “B” marks treated plots where 4: sprayed with low dosage (20 g/ha), 8: sprayed with high dosage (240 g/ha).

Fig. 2

The biological role of upregulated, defense-related genes showing differentiated gene expression as a result of treatments. Bar chart representation of the biological roles of upregulated defense-related genes showing differential gene expression as a result of treatments. The figure shows the biological processes of the Gene Ontology (GO) term and summarizes the annotated genes of each group. Data reflect the expression differences between individual genes obtained by Fisher's Exact Test.

Fig. 3

Based on the KEGG database, one of the genes was shown to be upregulated by the plant conditioning treatment, as the 7-IOMT enzyme, is located in the isoflavonoid biosynthesis pathway. The figure shows a part of the KEGG pathway of isoflavonoid biosynthesis (Abbreviations: 7-IOMT: isoflavone 7-O-methyltransferase, HI4OMT: 2-hydroxyisoflavanone 4′-O -methyltransferase).

Fig. 4

Expression analyses of GOIs in silico. Genes showing upregulation and related to defense-responses in the samples collected at two sampling times of the two treatments. Mean values are plotted and the standard deviation of the four biological replicates is shown. Data show increases in gene expression values in percentages compared to the control conditions.

Fig. 5

RT-qPCR amplification of GOIs. Relative gene expression was determined for GAPDH and TUBB3 reference genes. Values are mean ± SEM (n = 4). p < 0.05 vs. control, p < 0.05 vs. treated group.

Fig. 6

Representation of the experimental design. Timeline of samples collected in field-grown soybean. Numbers denote samples as follows: 525 means samples, collected on 25, May 2020; 710 means those, collected on 10 July 2020; ELICE16INDURES® treatments of 1–0 g/ha (control); 4–20 g/ha and 8–240 g/ha. Sample marks were indicated with NCBI data deposition numbers. Combined assembly means de novo Glycine max transcriptome dataset [22].