Bradyrhizobium centrolobii and Bradyrhizobium macuxiense sp. nov. isolated from Centrolobium paraense grown in soil of Amazonia, Brazil.

Thirteen Gram-negative, aerobic, motile with polar flagella, rod-shaped bacteria were isolated from root nodules of Centrolobium paraense Tul. grown in soils from the Amazon region of Brazil. Growth of strains was observed at temperature range 20-36 °C (optimal 28 °C), pH ranges 5-11 (optimal 6.0-7.0), and 0.1-0.5%NaCl (optimal 0.1-0.3%). Analysis of 16S rRNA gene placed the strains into two groups within Bradyrhizobium. Closest neighbouring species (98.8%) for group I was B. neotropicale while for group II were 12 species with more than 99% of similarity. Multi-locus sequence analysis (MLSA) with dnaK, glnII, recA, and rpoB confirmed B. neotropicale BR 10247T as the closest type strain for the group I and B. elkanii USDA 76T and B. pachyrhizi PAC 48T for group II. Average Nucleotide Identity (ANI) differentiated group I from the B. neotropicale BR 10247T (79.6%) and group II from B. elkanii USDA 76T and B. pachyrhizi PAC 48T (88.1% and 87.9%, respectively). Fatty acid profiles [majority C16:0 and Summed feature 8 (18:1ω6c/18:1ω7c) for both groups], DNA G + C content, and carbon compound utilization supported the placement of the novel strains in the genus Bradyrhizobium. Gene nodC and nifH of the new strains have in general low similarity with other Bradyrhizobium species. Both groups nodulated plants from the tribes Crotalarieae, Dalbergiae, Genisteae, and Phaseoleae. Based on the presented data, two novel species which the names Bradyrhizobium centrolobii and Bradyrhizobium macuxiense are proposed, with BR 10245T (=HAMBI 3597T) and BR 10303T (=HAMBI 3602T) as the respective-type strains.

Bradyrhizobium neotropicale sp. nov., isolated from effective nodules of Centrolobium paraense.

Root nodule bacteria were isolated from Centrolobium paraense Tul. grown in soils from the Amazon region, State of Roraima (Brazil). 16S rRNA gene sequence analysis of seven strains (BR 10247(T), BR 10296, BR 10297, BR 10298, BR 10299, BR 10300 and BR 10301) placed them in the genus Bradyrhizobium with the closest neighbours being the type strains of Bradyrhizobium paxllaeri (98.8 % similarity), Bradyrhizobium icense (98.8 %), Bradyrhizobium lablabi (98.7 %), Bradyrhizobium jicamae (98.6 %), Bradyrhizobium elkanii (98.6 %), Bradyrhizobium pachyrhizi (98.6 %) and Bradyrhizobium retamae (98.3 %). This high similarity, however, was not confirmed by the intergenic transcribed spacer (ITS) 16S-23S rRNA region sequence analysis nor by multi-locus sequence analysis. Phylogenetic analyses of five housekeeping genes (dnaK, glnII, gyrB, recA and rpoB) revealed Bradyrhizobium iriomotense EK05(T) ( = LMG 24129(T)) to be the most closely related type strain (95.7 % sequence similarity or less). Chemotaxonomic data, including fatty acid profiles [major components being C16 : 0 and summed feature 8 (18 : 1ω6c/18 : 1ω7c)], DNA G+C content, slow growth rate and carbon compound utilization patterns, supported the placement of the novel strains in the genus Bradyrhizobium. Results of DNA-DNA relatedness studies and physiological data (especially carbon source utilization) differentiated the strains from the closest recognized species of the genus Bradyrhizobium. Symbiosis-related genes for nodulation (nodC) and nitrogen fixation (nifH) placed the novel species in a new branch within the genus Bradyrhizobium. Based on the current data, these seven strains represent a novel species for which the name Bradyrhizobium neotropicale sp. nov. is proposed. The type strain is BR 10247(T) ( = HAMBI 3599(T)).

Bradyrhizobium ingae sp. nov., isolated from effective nodules of Inga laurina grown in Cerrado soil.

Root-nodule bacteria were isolated from Inga laurina (Sw.) Willd. growing in the Cerrado Amazon region, State of Roraima, Brazil. The 16S rRNA gene sequences of six strains (BR 10250(T), BR 10248, BR 10249, BR 10251, BR 10252 and BR 10253) showed low similarities with currently described species of the genus Bradyrhizobium. Phylogenetic analyses of sequences of five housekeeping genes (dnaK, glnII, gyrB, recA and rpoB) revealed Bradyrhizobium iriomotense EK05(T) to be the closest type strain (97.4% sequence similarity or less). Chemotaxonomic data, including fatty acid profiles [with the major components C16:0 and summed feature 8 (C18:1ω6c/C18:1ω7c)], the slow growth rate and carbon compound utilization patterns supported the assignment of our strains to the genus Bradyrhizobium. Results from DNA-DNA hybridizations and physiological traits differentiated our strains from the closest related species of the genus Bradyrhizobium with validly published names. Sequences of symbiosis-related genes for nodulation (nodC) and nitrogen fixation (nifH) grouped together with those of B. iriomotense EK05(T) and Bradyrhizobium sp. strains BR 6610 (used as a commercial inoculant for Inga marginata in Brazil) and TUXTLAS-10 (previously observed in Central America). Based on these data, the six strains represent a novel species, for which the name Bradyrhizobium ingae sp. nov. is proposed. The type strain is BR 10250(T) ( = HAMBI 3600(T)).

Bacteria isolated from soils of the western Amazon and from rehabilitated bauxite-mining areas have potential as plant growth promoters.

Several processes that promote plant growth were investigated in endophytic and symbiotic bacteria isolated from cowpea and siratro nodules and also in bacterial strains recommended for the inoculation of cowpea beans. The processes verified in 31 strains were: antagonism against phytopathogenic fungi, free-living biological nitrogen fixation, solubilization of insoluble phosphates and indole acetic acid (IAA) production. The resistance to antibiotics was also assessed. Sequencing of the partial 16S rRNA gene was performed and the strains were identified as belonging to different genera. Eight strains, including some identified as Burkholderia fungorum, fixed nitrogen in the free-living state. Eighteen strains exhibited potential to solubilize calcium phosphate, and 13 strains could solubilize aluminum phosphate. High levels of IAA production were recorded with L-tryptophan addition for the strain UFLA04-321 (42.3 µg mL⁻¹). Strains highly efficient in symbiosis with cowpea bean, including strains already approved as inoculants showed the ability to perform other processes that promote plant growth. Besides, these strains exhibited resistance to several antibiotics. The ability of the nitrogen-fixing bacteria to perform other processes and their adaptation to environmental conditions add value to these strains, which could lead to improved inoculants for plant growth and environmental quality.

Biotechnological potential of fungi from a mangrove ecosystem: Enzymes, salt tolerance and decolorization of a real textile effluent.

The mangrove is an ecosystem bounded by the line of the largest tide in size that occurs in climatic and subtropical regions. In this environment, microorganisms and their enzymes are involved in a series of transformations and nutrient cycling. To evaluate the biotechnological potential of fungi from a mangrove ecosystem, samples from mangrove trees were collected at the Paranaguá Estuarine Complex in Brazil and 40 fungal isolates were obtained, cultivated, and screened for hydrolytic and ligninolytic enzymes production, adaptation to salinity and genetic diversity. The results showed a predominance of hydrolytic enzymes and fungal tolerance to ≤ 50 g L-1 sodium chloride (NaCl) concentration, a sign of adaptive halophilia. Through morphological and molecular analyses, the isolates were identified as: Trichoderma atroveride, Microsphaeropsis arundinis, Epicoccum sp., Trichoderma sp., Gliocladium sp., Geotrichum sp. and Cryphonectria sp. The ligninolytic enzymatic potential of the fungi was evaluated in liquid cultures in the presence and absence of seawater and the highest activity of laccase among isolates was observed in the presence of seawater with M. arundinis (LB07), which produced 1,037 U L-1. Enzymatic extracts of M. arundinis fixed at 100 U L-1 of laccase partially decolorized a real textile effluent in a reaction without pH adjustment and chemical mediators. Considering that mangrove fungi are still few explored, the results bring an important contribution to the knowledge about these microorganisms, as their ability to adapt to saline conditions, biodegradation of pollutants, and enzymatic potential, which make them promising candidates in biotechnological processes.

Antioxidant, antibacterial and antitumoural activities of kraft lignin from hardwood fractionated by acid precipitation.

Kraft lignin, so far useful for energy generation, has been gathering considerable attention as an alternative material to replace fossil-based resources mainly due to its high phenolic content. However, the wide molecular weight distribution and chemical composition heterogeneity led to the development of fractionation methods. Herein, to narrow such characteristics we used eucalypt kraft lignin fractionated at pH's 9, 7, 5 and 3 by sequential acid precipitation. These lignin fractions were first characterised by simultaneous pyrolysis and trimethylsilylation (SPyT) with N-Methyl-N-(trimethylsilyl) trifluoroacetamide with posterior tests of antioxidant, antibacterial, and antitumour activities. We observed higher ratio of syringyl/guaiacyl groups and increase in antioxidant activity in those fractions with lower molecular weight (precipitated at lower pH's). Fractions precipitated at pH's 9 and 7 have shown an outstanding antibacterial activity against five bacteria. Moreover, fractions 7 and 5 presented at cytotoxicity tests higher ability to inhibit the growth of U87MG and T98G glioma cells, while only a slight inhibition of adult human fibroblasts (non-tumour cells) was detected.

Cupriavidus necator isolates are able to fix nitrogen in symbiosis with different legume species.

The aim of the present study was to identify a collection of 35 Cupriavidus isolates at the species level and to examine their capacity to nodulate and fix N(2). These isolates were previously obtained from the root nodules of two promiscuous trap species, Phaseolus vulgaris and Leucaena leucocephala, inoculated with soil samples collected near Sesbania virgata plants growing in Minas Gerais (Brazil) pastures. Phenotypic and genotypic methods applied for this study were SDS-PAGE of whole-cell proteins, and 16S rRNA and gyrB gene sequencing. To confirm the ability to nodulate and fix N(2), the presence of the nodC and nifH genes was also determined, and an experiment was carried out with two representative isolates in order to authenticate them as legume nodule symbionts. All 35 isolates belonged to the betaproteobacterium Cupriavidus necator, they possessed the nodC and nifH genes, and two representative isolates were able to nodulate five different promiscuous legume species: Mimosa caesalpiniaefolia, L. leucocephala, Macroptilium atropurpureum, P. vulgaris and Vigna unguiculata. This is the first study to demonstrate that C. necator can nodulate legume species.

Diazotrophic Burkholderia species isolated from the Amazon region exhibit phenotypical, functional and genetic diversity.

Forty-eight Burkholderia isolates from different land use systems in the Amazon region were compared to type strains of Burkholderia species for phenotypic and functional characteristics that can be used to promote plant growth. Most of these isolates (n=46) were obtained by using siratro (Macroptilium atropurpureum - 44) and common bean (Phaseolus vulgaris - 2) as the trap plant species; two isolates were obtained from nodules collected in the field from Indigofera suffruticosa and Pithecellobium sp. The evaluated characteristics were the following: colony characterisation on "79" medium, assimilation of different carbon sources, enzymatic activities, solubilisation of phosphates, nitrogenase activity and antifungal activity against Fusarium oxysporium f. sp. phaseoli. Whole cell protein profiles, 16S rRNA, gyrB, and recA gene sequencing and multilocus sequence typing were used to identify the isolates. The isolates showed different cultural and biochemical characteristics depending on the legume species from which they were obtained. Except for one isolate from I. suffruticosa, all isolates were able to solubilise calcium phosphate and present nitrogenase activity under free-living conditions. Only one isolate from common beans, showed antifungal activity. The forty four isolates from siratro nodules were identified as B. fungorum; isolates UFLA02-27 and UFLA02-28, obtained from common bean plants, were identified as B. contaminans; isolate INPA89A, isolated from Indigofera suffruticosa, was a close relative of B. caribensis but could not be assigned to an established species; isolate INPA42B, isolated from Pithecellobium sp., was identified as B. lata. This is the first report of nitrogenase activity in B. fungorum, B. lata and B. contaminans.