Exopolysaccharide production in Ensifer meliloti laboratory and native strains and their effects on alfalfa inoculation.

Bacterial surface molecules have an important role in the rhizobia-legume symbiosis. Ensifer meliloti (previously, Sinorhizobium meliloti), a symbiotic Gram-negative rhizobacterium, produces two different exopolysaccharides (EPSs), termed EPS I (succinoglycan) and EPS II (galactoglucan), with different functions in the symbiotic process. Accordingly, we undertook a study comparing the potential differences in alfalfa nodulation by E. meliloti strains with differences in their EPSs production. Strains recommended for inoculation as well as laboratory strains and native strains isolated from alfalfa fields were investigated. This study concentrated on EPS-II production, which results in mucoid colonies that are dependent on the presence of an intact expR gene. The results revealed that although the studied strains exhibited different phenotypes, the differences did not affect alfalfa nodulation itself. However, subtle changes in timing and efficacy to the effects of inoculation with the different strains may result because of other as-yet unknown factors. Thus, additional research is needed to determine the most effective inoculant strains and the best conditions for improving alfalfa production under agricultural conditions.

Draft Genome of Burkholderia cenocepacia TAtl-371, a Strain from the Burkholderia cepacia Complex Retains Antagonism in Different Carbon and Nitrogen Sources.

Burkholderia cenocepacia TAtl-371 was isolated from the rhizosphere of a tomato plant growing in Atlatlahucan, Morelos, Mexico. This strain exhibited a broad antimicrobial spectrum against bacteria, yeast, and fungi. Here, we report and describe the improved, high-quality permanent draft genome of B. cenocepacia TAtl-371, which was sequenced using a combination of PacBio RS and PacBio RS II sequencing methods. The 7,496,106 bp genome of the TAtl-371 strain is arranged in three scaffolds, contains 6722 protein-coding genes, and 99 RNA only-encoding genes. Genome analysis revealed genes related to biosynthesis of antimicrobials such as non-ribosomal peptides, siderophores, chitinases, and bacteriocins. Moreover, analysis of bacterial growth on different carbon and nitrogen sources shows that the strain retains its antimicrobial ability.

Reviewing and renewing the use of beneficial root and soil bacteria for plant growth and sustainability in nutrient-poor, arid soils.

A rapidly increasing human population coupled with climate change and several decades of over-reliance on synthetic fertilizers has led to two pressing global challenges: food insecurity and land degradation. Therefore, it is crucial that practices enabling both soil and plant health as well as sustainability be even more actively pursued. Sustainability and soil fertility encompass practices such as improving plant productivity in poor and arid soils, maintaining soil health, and minimizing harmful impacts on ecosystems brought about by poor soil management, including run-off of agricultural chemicals and other contaminants into waterways. Plant growth promoting bacteria (PGPB) can improve food production in numerous ways: by facilitating resource acquisition of macro- and micronutrients (especially N and P), modulating phytohormone levels, antagonizing pathogenic agents and maintaining soil fertility. The PGPB comprise different functional and taxonomic groups of bacteria belonging to multiple phyla, including Proteobacteria, Firmicutes, Bacteroidetes, and Actinobacteria, among others. This review summarizes many of the mechanisms and methods these beneficial soil bacteria use to promote plant health and asks whether they can be further developed into effective, potentially commercially available plant stimulants that substantially reduce or replace various harmful practices involved in food production and ecosystem stability. Our goal is to describe the various mechanisms involved in beneficial plant-microbe interactions and how they can help us attain sustainability.

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.

Inoculation With a Microbe Isolated From the Negev Desert Enhances Corn Growth.

Corn (Zea mays L.) is not only an important food source, but also has numerous uses, including for biofuels, fillers for cosmetics, glues, and so on. The amount of corn grown in the U.S. has significantly increased since the 1960's and with it, the demand for synthetic fertilizers and pesticides/fungicides to enhance its production. However, the downside of the continuous use of these products, especially N and P fertilizers, has been an increase in N2O emissions and other greenhouse gases into the atmosphere as well as run-off into waterways that fuel pollution and algal blooms. These approaches to agriculture, especially if exacerbated by climate change, will result in decreased soil health as well as human health. We searched for microbes from arid, native environments that are not being used for agriculture because we reasoned that indigenous microbes from such soils could promote plant growth and help restore degraded soils. Employing cultivation-dependent methods to isolate bacteria from the Negev Desert in Israel, we tested the effects of several microbial isolates on corn in both greenhouse and small field studies. One strain, Dietzia cinnamea 55, originally identified as Planomicrobium chinense, significantly enhanced corn growth over the uninoculated control in both greenhouse and outside garden experiments. We sequenced and analyzed the genome of this bacterial species to elucidate some of the mechanisms whereby D. cinnamea 55 promoted plant growth. In addition, to ensure the biosafety of this previously unknown plant growth promoting bacterial (PGPB) strain as a potential bioinoculant, we tested the survival and growth of Caenorhabditis elegans and Galleria mellonella (two animal virulence tests) as well as plants in response to D. cinnamea 55 inoculation. We also looked for genes for potential virulence determinants as well as for growth promotion.

Antifungal Activity of Bacillus Species Against Fusarium and Analysis of the Potential Mechanisms Used in Biocontrol.

Fusarium is a complex genus of ascomycete fungi that consists of plant pathogens of agricultural relevance. Controlling Fusarium infection in crops that leads to substantial yield losses is challenging. These economic losses along with environmental and human health concerns over the usage of chemicals in attaining disease control are shifting focus toward the use of biocontrol agents for effective control of phytopathogenic Fusarium spp. In the present study, an analysis of the plant-growth promoting (PGP) and biocontrol attributes of four bacilli (Bacillus simplex 30N-5, B. simplex 11, B. simplex 237, and B. subtilis 30VD-1) has been conducted. The production of cellulase, xylanase, pectinase, and chitinase in functional assays was studied, followed by in silico gene analysis of the PGP-related and biocontrol-associated genes. Of all the bacilli included in this study, B. subtilis 30VD-1 (30VD-1) demonstrated the most effective antagonism against Fusarium spp. under in vitro conditions. Additionally, 100 µg/ml of the crude 1-butanol extract of 30VD-1's cell-free culture filtrate caused about 40% inhibition in radial growth of Fusarium spp. Pea seed bacterization with 30VD-1 led to considerable reduction in wilt severity in plants with about 35% increase in dry plant biomass over uninoculated plants growing in Fusarium-infested soil. Phase contrast microscopy demonstrated distortions and abnormal swellings in F. oxysporum hyphae on co-culturing with 30VD-1. The results suggest a multivariate mode of antagonism of 30VD-1 against phytopathogenic Fusarium spp., by producing chitinase, volatiles, and other antifungal molecules, the characterization of which is underway.

Whole Genome Analyses Suggests that Burkholderia sensu lato Contains Two Additional Novel Genera (Mycetohabitans gen. nov., and Trinickia gen. nov.): Implications for the Evolution of Diazotrophy and Nodulation in the Burkholderiaceae.

Burkholderia sensu lato is a large and complex group, containing pathogenic, phytopathogenic, symbiotic and non-symbiotic strains from a very wide range of environmental (soil, water, plants, fungi) and clinical (animal, human) habitats. Its taxonomy has been evaluated several times through the analysis of 16S rRNA sequences, concantenated 4⁻7 housekeeping gene sequences, and lately by genome sequences. Currently, the division of this group into Burkholderia, Caballeronia, Paraburkholderia, and Robbsia is strongly supported by genome analysis. These new genera broadly correspond to the various habitats/lifestyles of Burkholderia s.l., e.g., all the plant beneficial and environmental (PBE) strains are included in Paraburkholderia (which also includes all the N2-fixing legume symbionts) and Caballeronia, while most of the human and animal pathogens are retained in Burkholderia sensu stricto. However, none of these genera can accommodate two important groups of species. One of these includes the closely related Paraburkholderia rhizoxinica and Paraburkholderia endofungorum, which are both symbionts of the fungal phytopathogen Rhizopus microsporus. The second group comprises the Mimosa-nodulating bacterium Paraburkholderia symbiotica, the phytopathogen Paraburkholderia caryophylli, and the soil bacteria Burkholderia dabaoshanensis and Paraburkholderia soli. In order to clarify their positions within Burkholderia sensu lato, a phylogenomic approach based on a maximum likelihood analysis of conserved genes from more than 100 Burkholderia sensu lato species was carried out. Additionally, the average nucleotide identity (ANI) and amino acid identity (AAI) were calculated. The data strongly supported the existence of two distinct and unique clades, which in fact sustain the description of two novel genera Mycetohabitans gen. nov. and Trinickia gen. nov. The newly proposed combinations are Mycetohabitans endofungorum comb. nov., Mycetohabitansrhizoxinica comb. nov., Trinickia caryophylli comb. nov., Trinickiadabaoshanensis comb. nov., Trinickia soli comb. nov., and Trinickiasymbiotica comb. nov. Given that the division between the genera that comprise Burkholderia s.l. in terms of their lifestyles is often complex, differential characteristics of the genomes of these new combinations were investigated. In addition, two important lifestyle-determining traits-diazotrophy and/or symbiotic nodulation, and pathogenesis-were analyzed in depth i.e., the phylogenetic positions of nitrogen fixation and nodulation genes in Trinickia via-à-vis other Burkholderiaceae were determined, and the possibility of pathogenesis in Mycetohabitans and Trinickia was tested by performing infection experiments on plants and the nematode Caenorhabditis elegans. It is concluded that (1) T. symbiotica nif and nod genes fit within the wider Mimosa-nodulating Burkholderiaceae but appear in separate clades and that T. caryophyllinif genes are basal to the free-living Burkholderia s.l. strains, while with regard to pathogenesis (2) none of the Mycetohabitans and Trinickia strains tested are likely to be pathogenic, except for the known phytopathogen T. caryophylli.

Draft genome of Paraburkholderia caballeronis TNe-841T, a free-living, nitrogen-fixing, tomato plant-associated bacterium.

10.1601/nm.26956 caballeronis is a plant-associated bacterium. Strain TNe-841T was isolated from the rhizosphere of tomato (Solanum lycopersicum L. var. lycopersicum) growing in Nepantla Mexico State. Initially this bacterium was found to effectively nodulate Phaseolus vulgaris L. However, from an analysis of the genome of strain TNe-841T and from repeat inoculation experiments, we found that this strain did not nodulate bean and also lacked nodulation genes, suggesting that the genes were lost. The genome consists of 7,115,141 bp with a G + C content of 67.01%. The sequence includes 6251 protein-coding genes and 87 RNA genes.