Cláudio Roberto Fonsêca Sousa Soares

New insights into 1-aminocyclopropane-1-carboxylate (ACC) deaminase phylogeny, evolution and ecological significance.

The main objective of this work is the study of the phylogeny, evolution and ecological importance of the enzyme 1-aminocyclopropane-1-carboxylate (ACC) deaminase, the activity of which represents one of the most important and studied mechanisms used by plant growth–promoting microorganisms. The ACC deaminase gene and its regulatory elements presence in completely sequenced organisms was verified by multiple searches in diverse databases, and based on the data obtained a comprehensive analysis was conducted. Strain habitat, origin and ACC deaminase activity were taken into account when analyzing the results. In order to unveil ACC deaminase origin, evolution and relationships with other closely related pyridoxal phosphate (PLP) dependent enzymes a phylogenetic analysis was also performed. The data obtained show that ACC deaminase is mostly prevalent in some Bacteria, Fungi and members of Stramenopiles. Contrary to previous reports, we show that ACC deaminase genes are predominantly vertically inherited in various bacterial and fungal classes. Still, results suggest a considerable degree of horizontal gene transfer events, including interkingdom transfer events. A model for ACC deaminase origin and evolution is also proposed. This study also confirms the previous reports suggesting that the Lrp-like regulatory protein AcdR is a common mechanism regulating ACC deaminase expression in Proteobacteria, however, we also show that other regulatory mechanisms may be present in some Proteobacteria and other bacterial phyla. In this study we provide a more complete view of the role for ACC deaminase than was previously available. The results show that ACC deaminase may not only be related to plant growth promotion abilities, but may also play multiple roles in microorganisms developmental processes. Hence, exploring the origin and functioning of this enzyme may be the key in a variety of important agricultural and biotechnological applications.

Biological nitrogen fixation and phosphate solubilization by bacteria isolated from tropical soils

IntroductionIn addition to fixing atmospheric nitrogen, some bacterial isolates can also solubilize insoluble phosphates, further contributing to plant growth.AimsThe objectives of this study were the following: isolate, select, and identify nodulating bacteria in the cowpea that are efficient not only in biological nitrogen fixation (BNF) but also in the solubilization of insoluble inorganic phosphates; identify and quantify the organic acids produced; and establish the relationship between those acids and the solubilizing capacity.MethodsThe bacteria were captured from two soils containing high concentrations of insoluble phosphorus from the cities of Lavras and Patos de Minas, using the cowpea [Vigna unguiculata (L.) Walp.] as bait. We obtained 78 strains, which were characterized according to their cultural attributes in culture medium 79 with the strains UFLA 03-84, INPA 03-11B, and BR3267 (approved by the Ministry of Livestock and Supply Agriculture—MAPA, as inoculants for the cowpea) and Burkholderia cepacia (LMG1222T), which was used as a positive control for phosphate solubilization. Strains that were selected for their efficiency in both processes were identified by 16S rDNA sequence analysis. We evaluated the symbiotic efficiency (BNF) in a greenhouse and the solubilization efficiency of CaHPO4, Al(H2PO4)3, and FePO4.2H2O in solid and liquid GELP media. Strains that excelled at the solubilization of these phosphate sources were also evaluated for the production of the following organic acids: oxalic, citric, gluconic, lactic, succinic, and propionic.ResultsThe presence of Acinetobacter, Bacillus, Firmicutes, Microbacterium, Paenibacillus, and Rhizobium was detected by 16S rDNA sequencing and analysis. Bacterial strains obtained from cowpea nodules varied greatly in the efficiency of their BNF and phosphate solubilization processes, especially in the strains UFLA 03-09, UFLA 03-10, UFLA 03-12, and UFLA 03-13, which were more efficient in both processes. More strains were able to solubilize insoluble inorganic calcium and iron phosphates in liquid medium than in solid medium. The production of organic acids was related to the solubilization of CaHPO4 and FePO4.2H2O for some strains, and the type and concentration of the acid influenced this process.ConclusionsThese are the first results obtained with bacterial isolates from tropical soils in which the production of organic acids was detected and quantified to examine the solubilization of insoluble inorganic phosphates.

Fitotoxidez de cádmio para Eucalyptus maculata e E. urophylla em solução nutritiva

The effects of increasing concentration of Cd in nutrient solution on growth, mineral uptake and phytotoxicity symptoms in Eucalyptus maculata and E. urophylla seedlings were studied under greenhouse conditions. Seedlings were kept for five weeks in 2 L pots containing Clarks nutrient solution amended with 0, 45, 90, 135 and 180 mM as CdSO4. After one week of exposure to Cd treatments seedlings of E. maculata exhibited nerval reddish spots, interveinal leaf chlorosis, leaf necrosis, drought and root darkening, whereas E. urophylla showed additionally apical dieback and leaf fall. The critical dose to reduce shoot dry matter by 10% was low; 2.4 mM and 1.5 mM of Cd to E. maculata and E. urophylla, respectively, while toxicity critical concentration in the shoots were 14.5 and 10.8 mg kg-1 dry matter for these species, respectively.These results indicate that E. maculata is probably less sensitive to Cd than E. urophylla. It was also found that Cd reduced translocation of Cu by 24 and 43% as compared to control in E. maculata and E. urophylla, whereas translocation of Fe was reduced from 36% on average for the two species in the control to 12% at 180 mM of Cd. Shoot concentrations of Ca and Mg was also reduced in both species, reaching levels below those considered adequate for E. urophylla. Increasing concentration of Cd in nutrient solution reduced Ca and Mg contents in shoots of species, reaching Mg contents below those of the range considered suitable for E. urophylla. Relationships of Cd phytotoxicity in Eucalyptus with Cu and Fe translocation as well as with Mg foliar contents were shown.

Arbuscular mycorrhizal fungi in phytoremediation of contaminated areas by trace elements: mechanisms and major benefits of their applications

In recent decades, the concentration of trace elements has increased in soil and water, mainly by industrialization and urbanization. Recovery of contaminated areas is generally complex. In that respect, microorganisms can be of vital importance by making significant contributions towards the establishment of plants and the stabilization of impacted areas. Among the available strategies for environmental recovery, bioremediation and phytoremediation outstand. Arbuscular mycorrhizal fungi (AMF) are considered the most important type of mycorrhizae for phytoremediation. AMF have broad occurrence in contaminated soils, and evidences suggest they improve plant tolerance to excess of certain trace elements. In this review, the use of AMF in phytoremediation and mechanisms involved in their trace element tolerance are discussed. Additionally, we present some techniques used to study the retention of trace elements by AMF, as well as a summary of studies showing major benefits of AMF for phytoremediation.

Solubilisation of inorganic phosphates by inoculant strains from tropical legumes

Microbial solubilisation of low soluble inorganic phosphates is an important process contributing for the phosphorus available to plants in tropical soils. This study evaluates the ability of inoculant strains for tropical legumes to solubilise inorganic phosphates of low solubility that are found in tropical soils. Seven strains of Leguminosae nodulating bacteria (LNB) were compared with one another and with a non-nodulating positive control, Burkholderia cepacia (LMG 1222T). Four of the strains are used as inoculants for cowpeas (Vigna unguiculata) (Bradyrhizobium sp. UFLA 03-84; Bradyrhizobium elkani INPA 03-11B and Bradyrhizobium japonicum BR3267) or for common beans (Phaseolus vulgaris) (Rhizobium tropici CIAT 899T). Rhizobium etli UFLA 02-100 and Rhizobium leguminosarum 316C10a are also efficient nodulators of beans and Cupriavidus taiwanensis LMG 19424T nodulates on Mimosa pudica. Two experiments, with solid and liquid media, were performed to determine whether the strains were able to solubilise CaHPO4, Al(H2PO4)3 or FePO4.2H2O. On solid GELP medium none of the strains dissolved FePO4.2H2O, but LMG 1222, UFLA 03-84 and CIAT 899 solubilised CaHPO4 particularly well. These strains, along with LMG 19424 and BR 3267, were also able to increase the solubility of Al(H2PO4)3. In liquid GELP medium, LMG 1222 solubilised all phosphate sources, but no legume nodulating strain could increase the solubility of Al(H2PO4)3. The strains CIAT 899 and UFLA 02-100 were the only legume nodulating bacteria able to solubilise the other phosphate sources in liquid media, dissolving both CaHPO4 and FePO4.2H2O. There was a negative correlation between the pH of the culture medium and the concentration of soluble phosphate when the phosphorus source was CaHPO4 or FePO4.2H2O. The contribution of these strains to increasing the phosphorus nutrition of legumes and non-legume plant species should be investigated further by in vivo experiments.

Arbuscular mycorrhizal fungi in arsenic-contaminated areas in Brazil

Arbuscular mycorrhizal fungi (AMF) are ubiquitous and establish important symbiotic relationships with the majority of the plants, even in soils contaminated with arsenic (As). In order to better understand the ecological relationships of these fungi with excess As in soils and their effects on plants in tropical conditions, occurrence and diversity of AMF were evaluated in areas affected by gold mining activity in Minas Gerais State, Brazil. Soils of four areas with different As concentrations (mg dm(-3)) were sampled: reference Area (10); B1 (subsuperficial layer) (396); barren material (573), and mine waste (1046). Soil sampling was carried out in rainy and dry seasons, including six composite samples per area (n = 24). AMF occurred widespread in all areas, being influenced by As concentrations and sampling periods. A total of 23 species were identified, belonging to the following genus: Acaulospora (10 species), Scutellospora (4 species), Racocetra (3 species), Glomus (4 species), Gigaspora (1 species) and Paraglomus (1 species). The most frequent species occurring in all areas were Paraglomus occultum, Acaulospora morrowiae and Glomus clarum. The predominance of these species indicates their high tolerance to excess As. Although arsenic contamination reduced AMF species richness, presence of host plants tended to counterbalance this reduction.

Phytoprotective Effect of Arbuscular Mycorrhizal Fungi Species Against Arsenic Toxicity in Tropical Leguminous Species

Arbuscular mycorrhizal fungi (AMF) improve the tolerance of hosting plants to arsenic (As) in contaminated soils. This work assessed the phytoprotective effect of Glomus etunicatum, Acaulospora morrowiae, Gigaspora gigantea, and Acaulospora sp. on four leguminous species (Acacia mangium, Crotalaria juncea, Enterolobium contortisiliquum, and Stizolobium aterrimum) in an As-contaminated soil from a gold mining area. AMF root colonization, biomass production, As and P accumulation, as well as arsenic translocation index (TI) from roots to shoots were measured. The AMF phytoprotective effect was assessed by the P/As ratio and the activity of plant antioxidant enzymes. The AMF colonization ranged from 24 to 28%. In general, all leguminous species had low As TI when inoculated with AMF species. Inoculation of C. juncea with Acaulospora sp. improved significantly As accumulation in roots, and decreased the activity of ascorbate peroxidase (APX) and superoxide dismutase (SOD), highlighting its phytoprotective effect and the potential use of this symbiosis for phytoremediation of As-contaminated soils. However, S. aterrimum has also shown a potential for phytoremediation irrespectively of AMF inoculation. APX was a good indicator of the phytoprotective effect against As contamination in C. juncea and A. mangium. In general P/As ratio in shoots was the best indicator of the phytoprotective effect of all AMF species in all plant species.

Eficiência simbiótica de estirpes de Cupriavidus necator tolerantes a zinco, cádmio, cobre e chumbo

The objective of this work was to evaluate the tolerance of Cupriavidus necator strains to zinc, cadmium, copper and lead, as well as to determine the symbiotic efficiency of the most tolerant ones in legume species suited for use in revegetation. Tolerance was evaluated in LB medium supplemented with 2.5, 5.0, 7.5, 10, 12.5 and 15 mmol L-1 of ZnSO4.7H2O, CdSO4.8H2O, CuSO4.5H2O and PbCl2, respectively, in comparison to a control without metal. The symbiotic efficiency of the four C. necator most metal-tolerant strains (UFLA02-71, UFLA02-73, UFLA01-659 and UFLA01-663) was determined, and these strains were inoculated in the species: Leucaena leucocephala, Enterolobium contortisiliquum, Acacia mangium, Mimosa caesalpiniifolia, M. pudica, M. pigra and M. acutistipula. Leucaena leucocephala, M. pudica and M. caesalpiniifolia and the strains UFLA02-71 and UFLA01-659, which presented the high symbiotic efficiency, were evaluated in pots with soil. UFLA02-71 provided increments of 870% in shoot dry matter of M. caesalpiniifolia, and UFLA01-659 provided 885% in M. pudica and 924% in L. leucocephala. These strains should be assessed for potential use in programs to restore degraded areas, since they showed high efficiency in nitrogen fixation and were competitive with indigenous rhizobia populations, besides being highly tolerant to heavy metals.

Biological attributes of rehabilitated soils contaminated with heavy metals.

This study aimed to evaluate the effects of two rehabilitation systems in sites contaminated by Zn, Cu, Pb, and Cd on biological soil attributes [microbial biomass carbon (Cmic), basal and induced respiration, enzymatic activities, microorganism plate count, and bacterial and fungal community diversity and structure by denaturing gradient gel electrophoresis (DGGE)]. These systems (S1 and S2) consisted of excavation (trenching) and replacement of contaminated soil by uncontaminated soil in rows with Eucalyptus camaldulensis planting (S1-R and S2-R), free of understory vegetation (S1-BR), or completely covered by Brachiaria decumbens (S2-BR) in between rows. A contaminated, non-rehabilitated (NR) site and two contamination-free sites [Cerrado (C) and pasture (P)] were used as controls. Cmic, densities of bacteria and actinobacteria, and enzymatic activities (β-glucosidase, acid phosphatase, and urease) were significantly higher in the rehabilitated sites of system 2 (S2-R and S2-BR). However, even under high heavy metal contents (S1-R), the rehabilitation with eucalyptus was also effective. DGGE analysis revealed similarity in the diversity and structure of bacteria and fungi communities between rehabilitated sites and C site (uncontaminated). Principal component analysis showed clustering of rehabilitated sites (S2-R and S2-BR) with contamination-free sites, and S1-R was intermediate between the most and least contaminated sites, demonstrating that the soil replacement and revegetation improved the biological condition of the soil. The attributes that most explained these clustering were bacterial density, acid phosphatase, β-glucosidase, fungal and actinobacterial densities, Cmic, and induced respiration.

Espécies tropicais de pteridófitas em associação com fungos micorrízicos arbusculares em solo contaminado com arsênio

The symbiosis of plants with mycorrhizal fungi represents an alternative to be considered during the processes of revegetation and rehabilitation of arsenic-contaminated soil. The aim of this study was to evaluate under greenhouse conditions the effect of arsenic on the mycorrhizal association of two species of tropical fern (Thelypteris salzmannii and Dicranopteris flexuosa). T. salzmannii had higher rates of colonization and higher density of spores while D. flexuosa showed greater sensitivity to smaller concentrations of arsenic and association with mycorrhizal fungi. Our results indicate that screening and selection of mycorrhizal fungal isolates/species is possible and effective for phytoremediation of arsenic-contaminated soils.