Bioremediation potential of Pseudomonas genus isolates from residual water, capable of tolerating lead through mechanisms of exopolysaccharide production and biosorption.

The mechanisms of tolerance to heavy metals used by some microorganisms identified by bioprospection processes are useful for the development and implementation of bioremediation strategies for contaminated environments with high toxic load caused by heavy metals. A total of seven native microbial isolates were obtained from wastewater bodies from an industrial zone in the municipality of Girardota, Antioquia, Colombia. Subsequently, they were selected to evaluate their lead tolerance capacity at different concentrations. In addition, some parameters were determined, such as the capacity to produce exopolysaccharides and their biosorption to understand potential mechanisms associated to lead tolerance. According to the biocehemical test (Vitek) and the molecular analysis of sequences of 16S rDNA, bacterial were identified as Pseudomonas aeruginosa, Pseudomonas nitroreducens, and Pseudomonas alcaligenes. We determined that the seven isolates had the capacity to tolerate concentrations higher than 50 mg/ml of lead, and that the concentration and exposure time (40 h) to this metal significantly affect the Pseudomonas spp. isolates. Statistically significant differences were detected (p < 0.05) in the production of the exopolysaccharide (EPS) among the isolates. P. aeruginosa (P16) was the strain with the maximum absorbance exopolysaccharide measured. We evidenced that P. aeruginosa (P14) and P. nitroreducens (P20) have 80% capacity to biosorber lead using live mass (minimum range from 80.9% to 87%). It is suggested that the tolerance to lead exhibited by the environmental isolates of Pseudomonas spp. can be attributed to the production of exopolysaccharides and biosorption, which are protection factors for its survival in contaminated places. Finally, it was determined that the adsorption measured from dead biomass was significant (p < 0.05) from 40 h of exposure to metal (Average 182.2 ± 7). We generated new knowledge about the potential use of the Pseudomonas spp. genus to bioremediate affluent contaminated with heavy metals.

Phylogenetic diversity of 200+ isolates of the ectomycorrhizal fungus Cenococcum geophilum associated with Populus trichocarpa soils in the Pacific Northwest, USA and comparison to globally distributed representatives.

The ectomycorrhizal fungal symbiont Cenococcum geophilum is of high interest as it is globally distributed, associates with many plant species, and has resistance to multiple environmental stressors. C. geophilum is only known from asexual states but is often considered a cryptic species complex, since extreme phylogenetic divergence is often observed within nearly morphologically identical strains. Alternatively, C. geophilum may represent a highly diverse single species, which would suggest cryptic but frequent recombination. Here we describe a new isolate collection of 229 C. geophilum isolates from soils under Populus trichocarpa at 123 collection sites spanning a ~283 mile north-south transect in Western Washington and Oregon, USA (PNW). To further understanding of the phylogenetic relationships within C. geophilum, we performed maximum likelihood and Bayesian phylogenetic analyses to assess divergence within the PNW isolate collection, as well as a global phylogenetic analysis of 789 isolates with publicly available data from the United States, Japan, and European countries. Phylogenetic analyses of the PNW isolates revealed three distinct phylogenetic groups, with 15 clades that strongly resolved at >80% bootstrap support based on a GAPDH phylogeny and one clade segregating strongly in two principle component analyses. The abundance and representation of PNW isolate clades varied greatly across the North-South range, including a monophyletic group of isolates that spanned nearly the entire gradient at ~250 miles. A direct comparison between the GAPDH and ITS rRNA gene region phylogenies, combined with additional analyses revealed stark incongruence between the ITS and GAPDH gene regions, consistent with intra-species recombination between PNW isolates. In the global isolate collection phylogeny, 34 clades were strongly resolved using Maximum Likelihood and Bayesian approaches (at >80% MLBS and >0.90 BPP respectively), with some clades having intra- and intercontinental distributions. Together these data are highly suggestive of divergence within multiple cryptic species, however additional analyses such as higher resolution genotype-by-sequencing approaches are needed to distinguish potential species boundaries and the mode and tempo of recombination patterns.

Genetic Selection for Small Molecule Production in Competitive Microfluidic Droplets.

Biosensors can be used to screen or select for small molecule production in engineered microbes. However, mutations to the biosensor that interfere with accurate signal transduction are common, producing an excess of false positives. Strategies have been developed to avoid this limitation by physically separating the production pathway and biosensor, but these approaches have only been applied to screens, not selections. We have developed a novel biosensor-mediated selection strategy using competition between cocultured bacteria. When applied to the biosynthesis of cis,cis-muconate, we show that this strategy yields a selective advantage to producer strains that outweighs the costs of production. By encapsulating the competitive cocultures into microfluidic droplets, we successfully enriched the muconate-producing strains from a large population of control nonproducers. Facile selections for small molecule production will increase testing throughput for engineered microbes and allow for the rapid optimization of novel metabolic pathways.

Abiotic Stresses Shift Belowground Populus-Associated Bacteria Toward a Core Stress Microbiome.

Adverse growth conditions can lead to decreased plant growth, productivity, and survival, resulting in poor yields or failure of crops and biofeedstocks. In some cases, the microbial community associated with plants has been shown to alleviate plant stress and increase plant growth under suboptimal growing conditions. A systematic understanding of how the microbial community changes under these conditions is required to understand the contribution of the microbiome to water utilization, nutrient uptake, and ultimately yield. Using a microbiome inoculation strategy, we studied how the belowground microbiome of Populus deltoides changes in response to diverse environmental conditions, including water limitation, light limitation (shading), and metal toxicity. While plant responses to treatments in terms of growth, photosynthesis, gene expression and metabolite profiles were varied, we identified a core set of bacterial genera that change in abundance in response to host stress. The results of this study indicate substantial structure in the plant microbiome community and identify potential drivers of the phytobiome response to stress. IMPORTANCE The identification of a common "stress microbiome" indicates tightly controlled relationships between the plant host and bacterial associates and a conserved structure in bacterial communities associated with poplar trees under different growth conditions. The ability of the microbiome to buffer the plant from extreme environmental conditions coupled with the conserved stress microbiome observed in this study suggests an opportunity for future efforts aimed at predictably modulating the microbiome to optimize plant growth.

Synthesis of 1,4-dihydro-benzo[d][1,3]oxazin-2-ones from phthalides via an aminolysis-Hofmann rearrangement protocol.

A simple two-step procedure for the conversion of readily available phthalides to the corresponding benzoxazinones was developed. Initial ring-opening aminolysis to form a primary 2-hydroxymethylbenzamide, followed by reaction with bis(trifluoroacetoxy)iodobenzene (BTI) conveniently provided a variety of 4-substituted benzoxazinones.