Streptomyces biostimulants: an effective sustainable approach to reduce inorganic N input and maintain high yield of wheat crop in different soil types.

AIMS: This study aimed to isolate and characterize endophytic plant growth-promoting (PGP) actinomycetes from the wild medicinal plant Zygophyllum album. METHODS AND RESULTS: Eight actinomycetes were isolated, identified, and screened for their PGP activities to improve the growth and production of wheat plants under low N-inputs. Based on 16S rRNA analysis, the isolated actinobacteria showed high diversity and had multiple in vitro PGP attributes. In pot experiments, Streptomyces sp. NGB-Act4 and NGB-Act6 demonstrated the highest significant PGP activities to enhance the growth of wheat plants under reduced N-inputs. Under various field conditions (high-fertility clay soils and low-fertility sandy soils), in combination with 50% N-dose, the two streptomycetes showed significant increases in grain N% and grain yield of the wheat crop compared with the 50% N-fertilized treatment. Irrespective of soil type, wheat plants inoculated with strain NGB-Act4 produced grain yield and grain N% significantly greater than or comparable to the full N-dose treatment. CONCLUSIONS: This is the first field report on the successful use of endophytic streptomycetes as an effective strategy to improve wheat yield and reduce the use of synthetic N fertilizers.

Native Rhizospheric and Endophytic Fungi as Sustainable Sources of Plant Growth Promoting Traits to Improve Wheat Growth under Low Nitrogen Input.

Wheat crops require effective nitrogen fertilization to produce high yields. Only half of chemical N2 fertilizers are absorbed into plants while the rest remains in the soil, causing environmental problems. Fungi could maximize nitrogen absorption, and from an environmental and biodiversity point of view, there is an urgent necessity for bioprospecting native fungi associated with wild plants growing in harsh environments, e.g., St. Katherine Protectorate (SKP) in the arid Sinai. Recovered taxa, either endophytic and/or rhizospheric, were screened for their plant growth-promoting (PGP) traits. Eighteen fungal isolates (15 rhizospheric and 3 endophytic) belonging to anamorphic ascomycetes were recovered from 9 different wild plants, and their PGP traits (indole-3-acetic acid [IAA] production, phosphate solubilization, siderophore production, and hydrolytic enzyme production) were measured. Rhizospheric isolate NGB-WS14 (Chaetosphaeronema achilleae) produced high levels of IAA (119.1 µg mL-1) in the presence of tryptophan, while NGB-WS 8 (Acrophialophora levis) produced high IAA levels (42.4 µg mL-1) in the absence of tryptophan. The highest phosphate-solubilizing activity (181.9 µg mL-1) was recorded by NGB-WFS2 (Penicillium chrysogenum). Endophytic isolate NGB-WFE16 (Fusarium petersiae) exhibited a high percentage level of Siderophore Unit (96.5% SU). All isolates showed variability in the secretion of extracellular hydrolytic enzymes. Remarkably, all isolates had antagonistic activity (55.6% to 87.3% suppression of pathogen growth) against the pathogenic taxon Alternaria alternata (SCUF00001378) in the dual-assay results. Out of the 18 isolates, 4 rhizospheric and 1 endophytic isolate showed significant increases in shoot dry weight and shoot nitrogen and chlorophyll content of wheat plants subjected to low inputs of chemical nitrogen (N) fertilizers (50% reduction) compared with the non-inoculated control in a pot experiment. Potent taxa were subjected to sequencing for molecular confirmation of phenotypic identification. The retrieved ITS sequences in this study have been deposited in GenBank under accession numbers from LC642736 to LC642740. This study considered the first report of endophytic fungi of Cheilanthes vellea, a wild plant with PGPF which improves wheat growth. These results recommend using PGPF as inoculants to alleviate low nitrogen fertilization.

Comparative Analysis of the Cultured and Total Bacterial Community in the Wheat Rhizosphere Microbiome Using Culture-Dependent and Culture-Independent Approaches.

Rhizosphere and root-associated bacteria are key components of crop production and sustainable agriculture. However, utilization of these beneficial bacteria is often limited by conventional culture techniques because a majority of soil microorganisms cannot be cultured using standard laboratory media. Therefore, the purpose of this study was to improve culturability and investigate the diversity of the bacterial communities from the wheat rhizosphere microbiome collected from three locations in Egypt with contrasting soil characteristics by using metagenomic analysis and improved culture-based methods. The improved strategies of the culture-dependent approach included replacing the agar in the medium with gellan gums and modifying its preparation by autoclaving the phosphate and gelling agents separately. Compared to the total operational taxonomic units (OTUs) observed from the metagenomic data sets derived from the three analyzed soils, 1.86 to 2.52% of the bacteria were recovered using the modified cultivation strategies, whereas less than 1% were obtained employing the standard cultivation protocols. Twenty-one percent of the cultivable isolates exhibited multiple plant growth-promoting (PGP) properties, including P solubilization activity and siderophore production. From the metagenomic analysis, the most abundant phyla were Proteobacteria, Actinobacteria, Chloroflexi, Bacteroidetes, and Firmicutes. Moreover, the relative abundance of the specific bacterial taxa was correlated with the soil characteristics, demonstrating the effect of the soil in modulating the plant rhizosphere microbiome. IMPORTANCE Bacteria colonizing the rhizosphere, a narrow zone of soil surrounding the root system, are known to have beneficial effects in improving the growth and stress tolerance of plants. However, most bacteria in natural environments, especially those in rhizosphere soils, are recalcitrant to cultivation using traditional techniques, and thus their roles in soil health and plant growth remain unexplored. Hence, investigating new culture media and culture conditions to bring "not-yet-cultured" species into cultivation and to identify new functions is still an important task for all microbiologists. To this end, we describe improved cultivation protocols that increase the number and diversity of cultured bacteria from the rhizosphere of wheat plants. Using such approaches will lead to new insights into culturing more beneficial bacteria that live in the plant rhizosphere, in so doing creating greater opportunities not only for field application but also for promoting sustainability.

Diverse Rhizobium strains isolated from root nodules of Trifolium alexandrinum in Egypt and symbiovars.

Berseem clover (T. alexandrinum) is the main forage legume crop used as animal feed in Egypt. Here, eighty rhizobial isolates were isolated from root nodules of berseem clover grown in different regions in Egypt and were grouped by RFLP-16S rRNA ribotyping. Representative isolates were characterized using phylogenetic analyses of the 16S rRNA, rpoB, glnA, pgi, and nodC genes. We also investigated the performance of these isolates using phenotypic tests and nitrogen fixation efficiency assays. The majority of strains (<90%) were closely related to Rhizobium aegyptiacum and Rhizobium aethiopicum and of the remaining strains, six belonged to the Rhizobium leguminosarum genospecies complex and only one strain was assigned to Agrobacterium fabacearum. Despite their heterogeneous chromosomal background, most of the strains shared nodC gene alleles corresponding to symbiovar trifolii. Some of the strains closely affiliated to R. aegyptiacum and R. aethiopicum had superior nodulation and nitrogen fixation capabilities in berseem clover, compared to the commercial inoculant (Okadein®) and N-added treatments. R. leguminosarum strain NGB-CR 17 that harbored a nodC allele typical of symbiovar viciae, was also able to form an effective symbiosis with clover. Two strains with nodC alleles of symbiovar trifolii, R. aegyptiacum strains NGB-CR 129 and 136, were capable of forming effective nodules in Phaseolus vulgaris in axenic greenhouse conditions. This adds the symbiovar trifolii which is well-established in the Egyptian soils to the list of symbiovars that form nodules in P. vulgaris.