Cell-free microbial culture filtrates as candidate biostimulants to enhance plant growth and yield and activate soil- and plant-associated beneficial microbiota.

In this work we compiled information on current and emerging microbial-based fertilization practices, especially the use of cell-free microbial culture filtrates (CFs), to promote plant growth, yield and stress tolerance, and their effects on plant-associated beneficial microbiota. In addition, we identified limitations to bring microbial CFs to the market as biostimulants. In nature, plants act as metaorganisms, hosting microorganisms that communicate with the plants by exchanging semiochemicals through the phytosphere. Such symbiotic interactions are of high importance not only for plant yield and quality, but also for functioning of the soil microbiota. One environmentally sustainable practice to increasing crop productivity and/or protecting plants from (a)biotic stresses while reducing the excessive and inappropriate application of agrochemicals is based on the use of inoculants of beneficial microorganisms. However, this technology has a number of limitations, including inconsistencies in the field, specific growth requirements and host compatibility. Beneficial microorganisms release diffusible substances that promote plant growth and enhance yield and stress tolerance. Recently, evidence has been provided that this capacity also extends to phytopathogens. Consistently, soil application of microbial cell-free culture filtrates (CFs) has been found to promote growth and enhance the yield of horticultural crops. Recent studies have shown that the response of plants to soil application of microbial CFs is associated with strong proliferation of the resident beneficial soil microbiota. Therefore, the use of microbial CFs to enhance both crop yield and stress tolerance, and to activate beneficial soil microbiota could be a safe, efficient and environmentally friendly approach to minimize shortfalls related to the technology of microbial inoculation. In this review, we compile information on microbial CFs and the main constituents (especially volatile compounds) that promote plant growth, yield and stress tolerance, and their effects on plant-associated beneficial microbiota. In addition, we identify challenges and limitations for their use as biostimulants to bring them to the market and we propose remedial actions and give suggestions for future work.

Glutamate dehydrogenase is essential in the acclimation of Virgilia divaricata, a legume indigenous to the nutrient-poor Mediterranean-type ecosystems of the Cape Fynbos.

Glutamate dehydrogenase (NAD(H)- GDH, EC 1.4.1.2) is an important enzyme in nitrogen (N) metabolism. It serves as a link between C and N metabolism, in its role of assimilating ammonia into glutamine or deaminating glutamate into 2-oxoglutarate and ammonia. GDH may also have a key in the N assimilation of legumes growing in P-poor soils. Virgilia divaricata is such a legume, growing in the nutrient limited soils of the mediterranean-type Cape fynbos ecosystem. In order to understand the role of GDH in the nitrogen nutrition of V. divaricata, the aim of this study was to identify the GDH gene transcripts, their relative expressions and enzyme activity in P-stressed roots and nodules during N metabolism. During P deficiency there was a reduction in total plant biomass as well as total plant P concentration. The analysis of the GDH cDNA sequences in V. divaricata revealed the presence of GHD1 and GHD2 subunits, these corresponding to the GDH1, GDH-B and GDH3 genes of legumes and non-legume plants. The relative expression of GDH1 and GDH2 genes in the roots and nodules, indicates that two the subunits were differently regulated depending on the organ type, rather than P supply. Although both transcripts appeared to be ubiquitously expressed in the roots and nodules, the GDH2 transcript evidently predominated over those of GDH1. Furthermore, the higher expression of both GDH transcripts in the roots than nodules, suggests that roots are more reliant on on GDH in P-poor soils, than nodules. With regards to GHD activity, both aminating and deaminating GDH activities were differently affected by P deficiency in roots and nodules. This may function to assimilate N and regulate internal C and N in the roots and nodules. The variation in GDH1 and GDH2 transcript expression and GDH enzyme activities, indicate that the enzyme may be regulated by post-translational modification, instead of by gene expression during P deficiency in V. divaricata.

Suppression of allene oxide synthase 3 in potato increases degree of arbuscular mycorrhizal fungal colonization.

Arbuscular mycorrhizal (AM) is a mutually beneficial interaction among higher plants and soil fungi of the phylum Glomeromycota. Numerous studies have pointed that jasmonic acid plays an important role in the development of the intraradical fungus. This compound belongs to a group of biologically active compounds known as oxylipins which are derived from the oxidative metabolism of polyunsaturated fatty acids. Studies of the regulatory role played by oxylipins in AM colonization have generally focused on jasmonates, while few studies exist on the 9-LOX pathway of oxylipins during AM formation. Here, the cDNA of Allene oxide synthase 3 (AOS3), a key enzyme in the 9-LOX pathway, was used in the RNA interference (RNAi) system to transform potato plants in order to suppress its expression. Results show increases in AOS3 gene expression and 9-LOX products in roots of wild type potato mycorrhizal plants. The suppression of AOS3 gene expression increases the percentage of root with mycorrhizal colonization at early stages of AM formation. AOS3 RNA interference lead to an induction of LOXA and 13-LOX genes, a reduction in AOS3 derived 9-LOX oxylipin compounds and an increase in jasmonic acid content, suggesting compensation between 9 and 13-LOX pathways. The results in a whole support the hypothesis of a regulatory role for the 9-LOX oxylipin pathway during mycorrhization.

Variations in the mycorrhization characteristics in roots of wild-type and ABA-deficient tomato are accompanied by specific transcriptomic alterations.

Abscissic acid (ABA) determines mycorrhiza functionality and arbuscule development. In this study, we performed transcriptome analysis in response to different mycorrhization status according to the ABA content in the root to identify genes that may play a role in arbuscule functionality. Affymetrix Tomato GeneChip (approximately 10,000 probes) allowed us to detect and compare the transcriptional root profiling of tomato (Solanum lycopersicum) wild-type and ABA-deficient sitiens plants colonized by Glomus intraradices. A number of identified genes in tomato belong to a category of genes already described as "mycorrhizal core-set" in other host plants. The impairment in arbuscular mycorrhiza (AM) formation in ABA-deficient mutants was associated with upregulation of genes related to defense and cell wall modification, whereas functional mycorrhization in wild-type plants was associated with activation of genes related to isoprenoid metabolism. The oxylipin pathway was activated in tomato mycorrhizal roots at late stages of interaction, and was related to the control of fungal spread in roots, not with the establishment of the symbiosis. Induction of selected genes, representing a range of biological functions and representative of the three sets of genes specifically upregulated in the different plant phenotype, was confirmed by quantitative reverse-transcription polymerase chain reaction, and their response to phythohormone treatment was tested, showing that ethylene and jasmonic acid are key regulators of gene expression during AM development. Comparative analysis of mycorrhiza upregulated functional categories revealed significant changes in gene expression associated with the different mycorrhization status according to the ABA content in the roots.