Molecular Docking of Lac_CB10: Highlighting the Great Potential for Bioremediation of Recalcitrant Chemical Compounds by One Predicted Bacteroidetes CopA-Laccase.

Laccases are multicopper oxidases (MCOs) with a broad application spectrum, particularly in second-generation ethanol biotechnology and the bioremediation of xenobiotics and other highly recalcitrant compounds. Synthetic pesticides are xenobiotics with long environmental persistence, and the search for their effective bioremediation has mobilized the scientific community. Antibiotics, in turn, can pose severe risks for the emergence of multidrug-resistant microorganisms, as their frequent use for medical and veterinary purposes can generate constant selective pressure on the microbiota of urban and agricultural effluents. In the search for more efficient industrial processes, some bacterial laccases stand out for their tolerance to extreme physicochemical conditions and their fast generation cycles. Accordingly, to expand the range of effective approaches for the bioremediation of environmentally important compounds, the prospection of bacterial laccases was carried out from a custom genomic database. The best hit found in the genome of Chitinophaga sp. CB10, a Bacteroidetes isolate obtained from a biomass-degrading bacterial consortium, was subjected to in silico prediction, molecular docking, and molecular dynamics simulation analyses. The putative laccase CB10_180.4889 (Lac_CB10), composed of 728 amino acids, with theoretical molecular mass values of approximately 84 kDa and a pI of 6.51, was predicted to be a new CopA with three cupredoxin domains and four conserved motifs linking MCOs to copper sites that assist in catalytic reactions. Molecular docking studies revealed that Lac_CB10 had a high affinity for the molecules evaluated, and the affinity profiles with multiple catalytic pockets predicted the following order of decreasing thermodynamically favorable values: tetracycline (-8 kcal/mol) > ABTS (-6.9 kcal/mol) > sulfisoxazole (-6.7 kcal/mol) > benzidine (-6.4 kcal/mol) > trimethoprim (-6.1 kcal/mol) > 2,4-dichlorophenol (-5.9 kcal/mol) mol. Finally, the molecular dynamics analysis suggests that Lac_CB10 is more likely to be effective against sulfisoxazole-like compounds, as the sulfisoxazole-Lac_CB10 complex exhibited RMSD values lower than 0.2 nm, and sulfisoxazole remained bound to the binding site for the entire 100 ns evaluation period. These findings corroborate that LacCB10 has a high potential for the bioremediation of this molecule.

Reclassification of the taxonomic status of SEMIA3007 isolated in Mexico B-11A Mex as Rhizobium leguminosarum bv. viceae by bioinformatic tools.

BACKGROUND: Evidence based on genomic sequences is extremely important to confirm the phylogenetic relationships within the Rhizobium group. SEMIA3007 was analyzed within the Mesorhizobium groups to define the underlying causes of taxonomic identification. We previously used biochemical tests and phenotypic taxonomic methods to identify bacteria, which can lead to erroneous classification. An improved understanding of bacterial strains such as the Mesorhizobium genus would increase our knowledge of classification and evolution of these species. RESULTS: In this study, we sequenced the complete genome of SEMIA3007 and compared it with five other Mesorhizobium and two Rhizobium genomes. The genomes of isolated SEMIA3007 showed several orthologs with M. huakuii, M. erdmanii and M. loti. We identified SEMIA3007 as a Mesorhizobium by comparing the 16S rRNA gene and the complete genome. CONCLUSION: Our ortholog, 16S rRNA gene and average nucleotide identity values (ANI) analysis all demonstrate SEMIA3007 is not Rhizobium leguminosarum bv. viceae. The results of the phylogenetic analysis clearly show SEMIA3007 is part of the Mesorhizobium group and suggest a reclassification is warranted.

Lignin degradation by a novel thermophilic and alkaline yellow laccase from Chitinophaga sp.

Laccases (EC 1.10.3.2) are oxidoreductases that belong to the multicopper oxidase subfamily and are classified as yellow/white or blue according to their absorption spectrum. Yellow laccases are more useful for industrial processes since they oxidize nonphenolic compounds in the absence of a redox mediator and stand out for being more stable and functional under extreme conditions. This study aimed to characterize a new laccase that was predicted to be present in the genome of Chitinophaga sp. CB10 - Lac_CB10. Lac_CB10, with a molecular mass of 100.06 kDa, was purified and characterized via biochemical assays using guaiacol as a substrate. The enzyme demonstrated extremophilic characteristics, exhibiting relative activity under alkaline conditions (CAPS buffer pH 10.5) and thermophilic conditions (80-90°C), as well as maintaining its activity above 50% for 5 h at 80°C and 90°C. Furthermore, Lac_CB10 presented a spectral profile typical of yellow laccases, exhibiting only one absorbance peak at 300 nm (at the T2/T3 site) and no peak at 600 nm (at the T1 site). When lignin was degraded using copper as an inducer, 52.27% of the material was degraded within 32 h. These results highlight the potential of this enzyme, which is a novel yellow laccase with thermophilic and alkaline activity and the ability to act on lignin. This enzyme could be a valuable addition to the biorefinery process. In addition, this approach has high potential for industrial application and in the bioremediation of contaminated environments since these processes often occur at extreme temperatures and pH values. IMPORTANCE: The characterization of the novel yellow laccase, Lac_CB10, derived from Chitinophaga sp. CB10, represents a significant advancement with broad implications. This enzyme displays exceptional stability and functionality under extreme conditions, operating effectively under both alkaline (pH 10.5) and thermophilic (80-90°C) environments. Its capability to maintain considerable activity over extended periods, even at high temperatures, showcases its potential for various industrial applications. Moreover, its distinctive ability to efficiently degrade lignin-demonstrated by a significant 52.27% degradation within 32 h-signifies a promising avenue for biorefinery processes. This newfound laccase's characteristics position it as a crucial asset in the realm of bioremediation, particularly in scenarios involving contamination at extreme pH and temperature levels. The study's findings highlight the enzyme's capacity to address challenges in industrial processes and environmental cleanup, signifying its vital role in advancing biotechnological solutions.

Unveiling genomic features linked to traits of plant growth-promoting bacterial communities from sugarcane.

Microorganisms are ubiquitous, and those inhabiting plants have been the subject of several studies. Plant-associated bacteria exhibit various biological mechanisms that enable them to colonize host plants and, in some cases, enhance their fitness. In this study, we describe the genomic features predicted to be associated with plant growth-promoting traits in six bacterial communities isolated from sugarcane. The use of highly accurate single-molecule real-time sequencing technology for metagenomic samples from these bacterial communities allowed us to recover 17 genomes. The taxonomic assignments for the binned genomes were performed, revealing taxa distributed across three main phyla: Bacillota, Bacteroidota, and Pseudomonadota, with the latter being the most representative. Subsequently, we functionally annotated the metagenome-assembled genomes (MAGs) to characterize their metabolic pathways related to plant growth-promoting traits. Our study successfully identified the enrichment of important functions related to phosphate and potassium acquisition, modulation of phytohormones, and mechanisms for coping with abiotic stress. These findings could be linked to the robust colonization of these sugarcane endophytes.

Draft genome sequence of the nitrogen-fixing symbiotic bacterium Bradyrhizobium elkanii 587.

The draft sequence of the genome of Bradyrhizobium elkanii 587 is presented. This was obtained using Illumina Next-Gen DNA sequencing combined with Sanger sequencing. Genes for the pathways involved in biological nitrogen fixation (the nif gene cluster), nod genes including nodABC, and genes for the type III protein secretion system (T3SS) are present.

Genetic diversity and population differentiation of the causal agent of citrus black spot in Brazil.

One of the most important diseases that affect sweet orange orchards in Brazil is the Citrus Black Spot that is caused by the fungus Guignardia citricarpa. This disease causes irreparable losses due to the premature falling of fruit, as well as its severe effects on the epidermis of ripe fruit that renders them unacceptable at the fresh fruit markets. Despite the fact that the fungus and the disease are well studied, little is known about the genetic diversity and the structure of the fungi populations in Brazilian orchards. The objective of this work was study the genetic diversity and population differentiation of G. citricarpa associated with four sweet orange varieties in two geographic locations using DNA sequence of ITS1-5.8S-ITS2 region from fungi isolates. We observed that different populations are closely related and present little genetic structure according to varieties and geographic places with the highest genetic diversity distributed among isolates of the same populations. The same haplotypes were sampled in different populations from the same and different orange varieties and from similar and different origins. If new and pathogenic fungi would become resistant to fungicides, the observed genetic structure could rapidly spread this new form from one population to others.

Negative effects on the development of Chrysodeixis includens and Spodoptera cosmioides fed by peanut plants inoculated with entomopathogenic fungi.

Recent studies have shown that entomopathogenic fungi, as endophytes, can have beneficial effects on plants, protecting them from defoliating insects. The potential of endophytic association by entomopathogenic fungi with the peanut crop has been little explored. In our study, we conducted experiments by inoculation of peanut seeds through a soil drench method with nine strains/species of entomopathogenic fungi of the genera Metarhizium, Beauveria and Cordyceps, subsequently these plants were consumed by two larval pests, Chrysodeixis includens and Spodoptera cosmioides. The parameters of larval growth rates, mortality, foliar consumption and larval period were observed during the development of larvae. In addition, the endophytic capacity of these fungi in peanut plants and their persistence in soil were investigated. In two replicate greenhouse trials for each larva, peanut plants were inoculated with fungi by the soil-drench method. We evaluated the performance of C. includens and S. cosmioides feeding on inoculated peanut plants starting at the 2nd larval instar. The larval and pupal weights of C. includens and S. cosmioides were significantly different among the fungal treatment groups, where insects feeding on control plants exhibited higher larval and pupal weights than insects feeding on treated plants. The differences in larval period showed that Control larvae pupated faster than the larvae fed on fungal-inoculated plants, fungal treatments had a larval period of 3 to 5 days more than the control. The mortality rates of C. includens and S. cosmioides were significantly different among the fungal treatment groups, insects fed on Control plants exhibited higher survival than insects fed on fungal-inoculated plants. The persistence of all Metarhizium fungi was higher in the soil compared to other fungi, and only Metarhizium and B. bassiana IBCB215 emerged from the phyllosphere of peanut plants. Although the fungus Cordyceps presented the worst performance among the fungal treatments. Overall, our results demonstrate the negative effects on the development of C. includens and S. cosmioides that were fed on fungal-inoculated peanut plants, the best results recorded were for Metarhizium strains and the fungus B. bassiana IBCB215.

A putative twin-arginine translocation system in the phytopathogenic bacterium Xylella fastidiosa.

The twin-arginine translocation (Tat) pathway of the xylem-limited phytopathogenic bacterium Xylella fastidiosa strain 9a5c, responsible for citrus variegated chlorosis, was explored. The presence of tatA, tatB, and tatC in the X. fastidiosa genome together with a list of proteins harboring 2 consecutive arginines in their signal peptides suggested the presence of a Tat pathway. The functional Tat dependence of X. fastidiosa OpgD was examined. Native or mutated signal peptides were fused to the ß-lactamase. Expression of fusion with intact signal peptides mediated high resistance to ampicillin in Escherichia coli tat+ but not in the E. coli tat null mutant. The replacement of the 2 arginines by 2 lysines prevented the export of ß-lactamase in E. coli tat+, demonstrating that X. fastidiosa OpgD carries a signal peptide capable of engaging the E. coli Tat machinery. RT-PCR analysis revealed that the tat genes are transcribed as a single operon. tatA, tatB, and tatC genes were cloned. Complementation assays in E. coli devoid of all Tat or TatC components were unsuccessful, whereas X. fastidiosa Tat components led to a functional Tat translocase in E. coli TatB-deficient strain. Additional experiments implicated that X. fastidiosa TatB component could form a functional heterologous complex with the E. coli TatC component.

Characterization of a new bifunctional endo-1,4-ß-xylanase/esterase found in the rumen metagenome.

Metagenomic data mining of the Nellore cattle rumen microbiota identified a new bifunctional enzyme, endo-1,4-ß-xylanase/esterase, which was subsequently overexpressed in E. coli BL21 (DE3). This enzyme was stable at pH intervals of 5 to 6.5 and temperatures between 30 and 45 °C, and under the test conditions, it had a Vmax of 30.959 ± 2.334 µmol/min/mg, Km of 3.6 ± 0.6 mM and kcat of 2.323 ± 175 s-1. Additionally, the results showed that the enzyme is tolerant to NaCl and organic solvents and therefore is suitable for industrial environments. Xylanases are widely applicable, and the synergistic activity of endo-1,4-ß-xylanase/esterase in a single molecule will improve the degradation efficiency of heteroxylans via the creation of xylanase binding sites. Therefore, this new molecule has the potential for use in lignocellulosic biomass processing and as an animal feed food additive and could improve xylooligosaccharide production efficiency.

Bagasse minority pathway expression: Real time study of GH2 ß-mannosidases from bacteroidetes.

After being isolated from a sugarcane pile, the bacterium Chitinophaga sp. CB10 demonstrated to be a rich source of carbohydrases, with 350 predicted CAZyme domains. CB10 was able to grow on carbohydrates of different structural complexities: glucose, carboxymethylcellulose, corn starch, galactomannan, Aloe vera gum and sugarcane bagasse. The sugarcane bagasse is a rich source of complex polymers, and the diversity of metabolites released by its enzymatic hydrolysis has an important role for green chemistry, including minority pathways such as the degradation of mannan conjugates. In this sense, CB10 demonstrated considerable levels of gene expression for mannanases, and was stable for a period of 96-144 hours in the presence of sugarcane bagasse as sole carbon source. The bacterium showed respectively 4.8x and 5.6x expression levels for two genes predicted for GH2 ß-mannosidase: one located within a gene cluster identified as "polysaccharide utilization loci" (PUL), and another a classic ß-mannosidase. These enzymes shared less than 45% of identity with enzymes characterized from the genus Chitinophaga belonging to the phylum Bacteroidetes. The degree of novelty-as demonstrated by the low identity with previously characterized enzymes; the remarkable capability to grow in different substrates; mannanase activity, evidenced by the release of residual oligosaccharides in the cultivation with galactomannan (HPLC-RID, 12.3 mMol); associated to the ability of mannanases expression in a low concentration of inductor conditions (sugarcane bagasse, 0.2%) indicate the high potential for the application of CB10 as a source of enzymes in the production of oligosaccharides from biomass. This capacity might prove to be very valuable for the biorefinery process of pre-biotic precursors and other functional oligosaccharides focused on the food and pharmaceutical industries.

Bacterial communities in mining soils and surrounding areas under regeneration process in a former ore mine.

Human activities on the Earth's surface change the landscape of natural ecosystems. Mining practices are one of the most severe human activities, drastically altering the chemical, physical and biological properties of the soil environment. Bacterial communities in soil play an important role in the maintenance of ecological relationships. This work shows bacterial diversity, metabolic repertoire and physiological behavior in five ecosystems samples with different levels of impact. These ecosystems belong to a historical area in Iron Quadrangle, Minas Gerais, Brazil, which suffered mining activities until its total depletion without recovery since today. The results revealed Proteobacteria as the most predominant phylum followed by Acidobacteria, Verrucomicrobia, Planctomycetes, and Bacteroidetes. Soils that have not undergone anthropological actions exhibit an increase ability to degrade carbon sources. The richest soil with the high diversity was found in ecosystems that have suffered anthropogenic action. Our study shows profile of diversity inferring metabolic profile, which may elucidate the mechanisms underlying changes in community structure in situ mining sites in Brazil. Our data comes from contributing to know the bacterial diversity, relationship between these bacteria and can explore strategies for natural bioremediation in mining areas or adjacent areas under regeneration process in iron mining areas.

Bg10: A Novel Metagenomics Alcohol-Tolerant and Glucose-Stimulated GH1 ß-Glucosidase Suitable for Lactose-Free Milk Preparation.

New ß-glucosidases with product (glucose) or ethanol tolerances are greatly desired to make industrial processes more marketable and efficient. Therefore, this report describes the in silico/vitro characterization of Bg10, a metagenomically derived homodimeric ß-glucosidase that exhibited a Vmax of 10.81 ± 0.43 µM min-1, Kcat of 175.1± 6.91 min-1, and Km of 0.49 ± 0.12 mM at a neutral pH and 37°C when pNP-ß-D-glucopyranoside was used as the substrate, and the enzyme retained greater than 80% activity within the respective pH and temperature ranges of 6.5 to 8.0 and 35 to 40°C. The enzyme was stimulated by its product, glucose; consequently, the Bg10 activity against 50 and 100 mM of glucose were increased by 36.8% and 22%, respectively, while half of the activity was retained at 350 mM. Moreover, the Bg10 was able to hydrolyse 55% (milk sample) and 100% (purified sugar) of the lactose at low (6°C) and optimum (37°C) temperatures, respectively, suggesting the possibility of further optimization of the reaction for lactose-free dairy production. In addition, the enzyme was able to fully hydrolyse 40 mM of cellobiose at one hour and was tolerant to ethanol up to concentrations of 500 mM (86% of activity), while a 1 M concentration still resulted in 41% residual activity, which could be interesting for biofuel production.

Influence of Vinasse Application in the Structure and Composition of the Bacterial Community of the Soil under Sugarcane Cultivation.

Although the use of vinasse as a waste helps replenish soil nutrients and improves the quality of the sugarcane crop, it is known that vinasse residues alter the diversity of bacteria naturally present in the soil. The actual impacts of vinasse application on the selection of bacterial taxa are not understood because no studies have addressed this phenomenon directly. Analysis of 16S rRNA gene clone sequences from four soil types showed that the soil planted with sugarcane and fertilized with vinasse has a high diversity of bacteria compared to other biomes, where Acidobacteria were the second most abundant phylum. Although the composition and structure of bacterial communities differ significantly in the four environments (Libshuff's test), forest soils and soil planted with sugarcane without vinasse fertilizer were similar to each other because they share at least 28 OTUs related to Rhizobiales, which are important agents involved in nitrogen fixation. OTUs belonging to Actinomycetales were detected more often in the soil that had vinasse applied, indicating that these groups are more favored by this type of land management.

Optimized medium culture for Acidobacteria subdivision 1 strains.

Members of subdivision 1 of the phylum Acidobacteria were grown at different pH values in a new medium formulation named PSYL 5, which includes sucrose as a carbon source and other compounds (such as KH2PO4 and MgSO4.7H2O). Growth rate was nearly constant at pH 5.0 and declined at pH 3-4 and 6-7. However, it was found that effects involving good carbon/nitrogen ratios and pH on the growth of the members of Acidobacteria subdivision 1 were significant, and the strongest effect of these conditions was at pH 5.0. In addition, incubation time of 48, 72, 96 and 120 h was shorter than that described previously for members of Acidobacteria subdivision 1 on solid laboratory media.

Remoção de herbicida atrazina por meio de filtros de carvão ativado granular associados com microrganismos no tratamento de água para abastecimento / Removal of atrazine herbicide through granular activated carbon filters associated with microorganisms in drinking water treatment

RESUMO A atrazina é um herbicida sintético comumente utilizado no controle de ervas gramíneas daninhas e folhagens em lavouras, e é um dos principais contaminantes dos solos e dos ecossistemas aquáticos. Muitos métodos têm sido sugeridos para remover herbicidas da água potável. Contudo, esses métodos são custosos, muitos têm problemas de desempenho, produzem diversos subprodutos intermediários tóxicos, prejudiciais e perigosos. Entretanto, a atrazina é susceptível à degradação por microrganismos presentes na água, no sedimento e no esgoto. Considerando esses aspectos, o objetivo principal do estudo foi investigar a biodegradação e a filtração por meio de filtros de carvão com atividade biológica (CAB) para remoção da atrazina, e sua identificação filogenética associada a esses microrganismos. Os resultados demonstraram que a atrazina foi degradada por microrganismos presentes no biofilme, com remoção superior a 89% nos filtros CAB. Os microrganismos encontrados integram-se ao grupo das bactérias, composto dos gêneros Acinetobacter,Bacillus, Exiguobacterium e Pseudomonas. Este estudo nos permite inferir sobre a capacidade de biodegradação da atrazina por bactérias presentes nos filtros CAB, a capacidade de remover herbicidas por meio desse sistema de filtros e a possível utilização dessa tecnologia como alternativa para o controle e a remoção dessa substância no tratamento de água.

ABSTRACT Atrazine is a synthetic herbicide commonly used to control weeds and foliage in crops, and is a major contaminants of soil and water ecosystems. Many methods have been suggested to remove herbicides from drinking water. However, these methods are very costly, many have performance problems, produce a lot of toxic intermediates which are very harmful and dangerous. However, atrazine is susceptible to degradation by microorganism present in water, sediment, and sewage effluents. Considering these aspects, the main objective of the study was to investigate the biodegradation and filtration for using biological activated carbon (BAC) filters to remove atrazine, and their phylogenetic identification associated with these microorganisms. The results showed that atrazine was biodegraded by microorganism present in the biofilm, with removal over 80% in BAC filters. The microorganisms found integrate to the group of bacteria, composed by the genera Acinetobacter, Bacillus, Exiguobacterium, and Pseudomonas. This study allows us to infer the ability to biodegrade atrazine by bacteria present in BAC filters and capacity to remove herbicides by BAC filters, and the possible use of this technology as an alternative for the control and removal of this substance in water treatment.

Intense Exercise and Aerobic Conditioning Associated with Chromium or L-Carnitine Supplementation Modified the Fecal Microbiota of Fillies.

Recent studies performed in humans and rats have reported that exercise can alter the intestinal microbiota. Athletic horses perform intense exercise regularly, but studies characterizing horse microbiome during aerobic conditioning programs are still limited. Evidence has indicated that this microbial community is involved in the metabolic homeostasis of the host. Research on ergogenic substances using new sequencing technologies have been limited to the intestinal microbiota and there is a considerable demand for scientific studies that verify the effectiveness of these supplements in horses. L-carnitine and chromium are potentially ergogenic substances for athletic humans and horses since they are possibly able to modify the metabolism of carbohydrates and lipids. This study aimed to assess the impact of acute exercise and aerobic conditioning, associated either with L-carnitine or chromium supplementation, on the intestinal microbiota of fillies. Twelve "Mangalarga Marchador" fillies in the incipient fitness stage were distributed into four groups: control (no exercise), exercise, L-carnitine (10g/day) and chelated chromium (10mg/day). In order to investigate the impact of acute exercise or aerobic conditioning on fecal microbiota all fillies undergoing the conditioning program were analyzed as a separate treatment. The fillies underwent two incremental exercise tests before and after training on a treadmill for 42 days at 70-80% of the lactate threshold intensity. Fecal samples were obtained before and 48 h after acute exercise (incremental exercise test). Bacterial populations were characterized by sequencing the V4 region of the 16S rRNA gene using the MiSeq Illumina platform, and 5,224,389 sequences were obtained from 48 samples. The results showed that, overall, the two most abundant phyla were Firmicutes (50.22%) followed by Verrucomicrobia (15.13%). The taxa with the highest relative abundances were unclassified Clostridiales (17.06%) and "5 genus incertae sedis" from the phylum Verrucomicrobia (12.98%). There was a decrease in the phylum Chlamydiae and in the genus Mycobacterium after the second incremental exercise test. Intense exercise changed the community's structure and aerobic conditioning was associated with changes in the composition and structure of the intestinal bacterial population of fillies. The intra-group comparison showed that chromium or L-carnitine induced moderate changes in the fecal microbiota of fillies, but the microbiota did not differ from the control group, which was exercised with no supplementation. Fecal pH correlated positively with Simpson's index, while plasma pH correlated negatively. Our results show that exercise and aerobic conditioning can change in the microbiota and provide a basis for further studies enrolling a larger number of horses at different fitness levels to better understand the effects of exercise and training on the intestinal microbiota of horses.

Est16, a New Esterase Isolated from a Metagenomic Library of a Microbial Consortium Specializing in Diesel Oil Degradation.

Lipolytic enzymes have attracted attention from a global market because they show enormous biotechnological potential for applications such as detergent production, leather processing, cosmetics production, and use in perfumes and biodiesel. Due to the intense demand for biocatalysts, a metagenomic approach provides methods of identifying new enzymes. In this study, an esterase designated as Est16 was selected from 4224 clones of a fosmid metagenomic library, revealing an 87% amino acid identity with an esterase/lipase (accession number ADM63076.1) from an uncultured bacterium. Phylogenetic studies showed that the enzyme belongs to family V of bacterial lipolytic enzymes and has sequence and structural similarities with an aryl-esterase from Pseudomonas fluorescens and a patented Anti-Kazlauskas lipase (patent number US20050153404). The protein was expressed and purified as a highly soluble, thermally stable enzyme that showed a preference for basic pH. Est16 exhibited activity toward a wide range of substrates and the highest catalytic efficiency against p-nitrophenyl butyrate and p-nitrophenyl valerate. Est16 also showed tolerance to the presence of organic solvents, detergents and metals. Based on molecular modeling, we showed that the large alpha-beta domain is conserved in the patented enzymes but not the substrate pocket. Here, it was demonstrated that a metagenomic approach is suitable for discovering the lipolytic enzyme diversity and that Est16 has the biotechnological potential for use in industrial processes.

Chemical and rheological properties of exopolysaccharides produced by four isolates of rhizobia.

The rheological, physicochemical properties, emulsification and stability of exopolysaccharides (EPSs) from four rhizobia isolates (LBMP-C01, LBMP-C02, LBMP-C03 and LBMP-C04) were studied. The EPS yields of isolates under these experimental conditions were in the range of 1.5-6.63gL(-1). The LBMP-C04 isolate, which presented the highest EPS production (6.63gL(-1)), was isolated from Arachis pintoi and was identified as a Rhizobium sp. strain that could be explored as a possible potential source for the production of extracellular heteropolysaccharides. All polymers showed a pseudoplastic non-Newtonian fluid behavior or shear thinning property in aqueous solutions. Among the four EPS tested against hydrocarbons, EPS LBMP-C01 was found to be more effective against hexane, olive and soybean oils (89.94%, 82.75% and 81.15%, respectively). Importantly, we found that changes in pH (2-11) and salinity (0-30%) influenced the emulsification of diesel oil by the EPSs. EPSLBMP-C04 presented optimal emulsification capacity at pH 10 (E24=53%) and 30% salinity (E24=27%). These findings contribute to the understanding of the influence of the chemical composition, physical properties and biotechnology applications of rhizobial EPS solutions their bioemulsifying properties.

Genomics-based design of defined growth media for the plant pathogen Xylella fastidiosa.

Based on the genetic analysis of the phytopathogen Xylella fastidiosa genome, five media with defined composition were developed and the growth abilities of this fastidious prokaryote were evaluated in liquid media and on solid plates. All media had a common salt composition and included the same amounts of glucose and vitamins but differed in their amino acid content. XDM(1) medium contained amino acids threonine, serine, glycine, alanine, aspartic acid and glutamic acid, for which complete degradation pathways occur in X. fastidiosa; XDM(2) included serine and methionine, amino acids for which biosynthetic enzymes are absent, plus asparagine and glutamine, which are abundant in the xylem sap; XDM(3) had the same composition as XDM(2) but with asparagine replaced by aspartic acid due to the presence of complete degradation pathway for aspartic acid; XDM(4) was a minimal medium with glutamine as a sole nitrogen source; XDM(5) had the same composition as XDM(4), plus methionine. The liquid and solidified XDM(2) and XDM(3) media were the most effective for the growth of X. fastidiosa. This work opens the opportunity for the in silico design of bacterial defined media once their genome is sequenced.

Evaluation of the biotechnological potential of Rhizobium tropici strains for exopolysaccharide production.

Rhizobium tropici, a member of the Rhizobiaceae family, has the ability to synthesize and secrete extracellular polysaccharides (EPS). Rhizobial EPS have attracted much attention from the scientific and industrial communities. Rhizobial isolates and R. tropici mutants that produced higher levels of EPS than the wild-type strain SEMIA4080 were used in the present study. The results suggested a heteropolymer structure for these EPS composed by glucose and galactose as prevailing monomer unit. All EPS samples exhibited a typical non-Newtonian and pseudoplastic fluid flow, and the aqueous solutions apparent viscosities increased in a concentration-dependent manner. These results serve as a foundation for further studies aimed at enhancing interest in the application of the MUTZC3, JAB1 and JAB6 strains with high EPS production and viscosity can be exploited for the large-scale commercial production of Rhizobial polysaccharides.