Lucélia Cabral

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.

Mercury (II) removal by resistant bacterial isolates and mercuric (II) reductase activity in a new strain of Pseudomonas sp. B50A

This study aimed to isolate mercury resistant bacteria, determine the minimum inhibitory concentration for Hg, estimate mercury removal by selected isolates, explore the mer genes, and detect and characterize the activity of the enzyme mercuric (II) reductase produced by a new strain of Pseudomonas sp. B50A. The Hg removal capacity of the isolates was determined by incubating the isolates in Luria Bertani broth and the remaining mercury quantified by atomic absorption spectrophotometry. A PCR reaction was carried out to detect the merA gene and the mercury (II) reductase activity was determined in a spectrophotometer at 340 nm. Eight Gram-negative bacterial isolates were resistant to high mercury concentrations and capable of removing mercury, and of these, five were positive for the gene merA. The isolate Pseudomonas sp. B50A removed 86% of the mercury present in the culture medium and was chosen for further analysis of its enzyme activity. Mercuric (II) reductase activity was detected in the crude extract of this strain. This enzyme showed optimal activity at pH 8 and at temperatures between 37 °C and 45 °C. The ions NH4(+), Ba(2+), Sn(2+), Ni(2+) and Cd(2+) neither inhibited nor stimulated the enzyme activity but it decreased in the presence of the ions Ca(2+), Cu(+) and K(+). The isolate and the enzyme detected were effective in reducing Hg(II) to Hg(0), showing the potential to develop bioremediation technologies and processes to clean-up the environment and waste contaminated with mercury.

Anthropogenic impact on mangrove sediments triggers differential responses in the heavy metals and antibiotic resistomes of microbial communities.

Mangroves are complex and dynamic ecosystems highly dependent on diverse microbial activities. In the last decades, these ecosystems have been exposed to and affected by diverse human activities, such as waste disposal and accidental oil spills. Complex microbial communities inhabiting the soil and sediment of mangroves comprise microorganisms that have developed mechanisms to adapt to organic and inorganic contaminants. The resistance of these microbes to contaminants is an attractive property and also the reason why soil and sediment living microorganisms and their enzymes have been considered promising for environmental detoxification. The aim of the present study was to identify active microbial genes in heavy metals, i.e., Cu, Zn, Cd, Pb and Hg, and antibiotic resistomes of polluted and pristine mangrove sediments through the comparative analysis of metatranscriptome data. The concentration of the heavy metals Zn, Cr, Pb, Cu, Ni, Cd, and Hg and abundance of genes and transcripts involved in resistance to toxic compounds (the cobalt-zinc-cadmium resistance protein complex; the cobalt-zinc-cadmium resistance protein CzcA and the cation efflux system protein CusA) have been closely associated with sites impacted with petroleum, sludge and other urban waste. The taxonomic profiling of metatranscriptome sequences suggests that members of Gammaproteobacteria and Deltaproteobacteria classes contribute to the detoxification of the polluted soil. Desulfobacterium autotrophicum was the most abundant microorganism in the oil-impacted site and displayed specific functions related to heavy metal resistance, potentially playing a key role in the successful persistence of the microbial community of this site.

Metal resistance mechanisms in Gram-negative bacteria and their potential to remove Hg in the presence of other metals

Contamination of the environment by heavy metals has been increasing in recent years due to industrial activities. Thus research involving microorganisms capable of surviving in multi-contaminated environments is extremely important. The objectives of the present study were to evaluate the removal of mercury alone and in the presence of cadmium, nickel and lead by four mercury-resistant microorganisms; estimate the removal of Cd, Ni and Pb; understand the mechanisms involved (reduction, siderophores, biofilms, biosorption and bioaccumulation) in the metal resistance of the isolate Pseudomonas sp. B50D; and determine the capacity of Pseudomonas sp. B50D in removing Hg, Cd, Ni and Pb from an industrial effluent. It was shown that the four isolates evaluated were capable of removing from 62% to 95% of mercury from a culture medium with no addition of other metals. The isolate Pseudomonas sp. B50D showed the best performance in the removal of mercury when evaluated concomitantly with other metals. This isolate was capable of removing 75% of Hg in the presence of Cd and 91% in the presence of Ni and Pb. With respect to the other metals it removed 60%, 15% and 85% of Cd, Ni and Pb, respectively. In tests with effluent, the isolate Pseudomonas sp. B50D removed 85% of Hg but did not remove the other metals. This isolate presented reduction, biosorption, biofilm production and siderophore production as its metal resistance mechanisms. Pseudomonas sp. B50D was thus a candidate with potential for application in the bioremediation of effluents with complex metal contaminations.

Diversity of aromatic hydroxylating dioxygenase genes in mangrove microbiome and their biogeographic patterns across global sites

Aromatic hydrocarbons (AH), such as polycyclic aromatic hydrocarbons, are compounds largely found in nature. Aromatic‐ring‐hydroxylating dioxygenases (ARHD) are proteins involved in AH degradation pathways. We used ARHD functional genes from an oil‐impacted mangrove area and compared their diversity with other sites around the world to understand the ARHD biogeographic distribution patterns. For this, a comprehensive database was established with 166 operational protein families (OPFs) from 1,758 gene sequences obtained from 15 different sites worldwide, of which twelve are already published studies and three are unpublished. Based on a deduced ARHD peptide sequences consensus phylogeny, we examined trends and divergences in the sequence phylogenetic clustering from the different sites. The taxonomic affiliation of the OPF revealed that Pseudomonas, Streptomyces, Variovorax, Bordetella and Rhodococcus were the five most abundant genera, considering all sites. The functional diversity analysis showed the enzymatic prevalence of benzene 1,2‐dioxygenase, 3‐phenylpropionate dioxygenase and naphthalene 1,2‐dioxygenase, in addition to 10.98% of undefined category ARHDs. The ARHD gene correlation analysis among different sites was essentially important to gain insights on spatial distribution patterns, genetic congruence and ecological coherence of the bacterial groups found. This work revealed the genetic potential from the mangrove sediment for AH biodegradation and a considerable evolutionary proximity among the dioxygenase OPFs found in Antarctica and South America sites, in addition to high level of endemism in each continental region.

Potential of semiarid soil from Caatinga biome as a novel source for mining lignocellulose-degrading enzymes

&NA; The litterfall is the major organic material deposited in soil of Brazilian Caatinga biome, thus providing the ideal conditions for plant biomass‐degrading microorganisms to thrive. Herein, the phylogenetic composition and lignocellulose‐degrading capacity have been explored for the first time from a fosmid library dataset of Caatinga soil by sequence‐based screening. A complex bacterial community dominated by Proteobacteria and Actinobacteria was unraveled. SEED subsystems‐based annotations revealed a broad range of genes assigned to carbohydrate and aromatic compounds metabolism, indicating microbial ability to utilize plant‐derived material. CAZy‐based annotation identified 7275 genes encoding 37 glycoside hydrolases (GHs) families related to hydrolysis of cellulose, hemicellulose, oligosaccharides and other lignin‐modifying enzymes. Taxonomic affiliation of genes showed high genetic potential of the phylum Acidobacteria for hemicellulose degradation, whereas Actinobacteria members appear to play an important role in celullose hydrolysis. Additionally, comparative analyses revealed greater GHs profile similarity among soils as compared to the digestive tract of animals capable of digesting plant biomass, particularly in the hemicellulases content. Combined results suggest a complex synergistic interaction of community members required for biomass degradation into fermentable sugars. This large repertoire of lignocellulolytic enzymes opens perspectives for mining potential candidates of biochemical catalysts for biofuels production from renewable resources and other environmental applications.

Methylmercury degradation by Pseudomonas putida V1

Environmental contamination of mercury (Hg) has caused public health concerns with focuses on the neurotoxic substance methylmercury, due to its bioaccumulation and biomagnification in food chains. The goals of the present study were to examine: (i) the transformation of methylmercury, thimerosal, phenylmercuric acetate and mercuric chloride by cultures of Pseudomonas putida V1, (ii) the presence of the genes merA and merB in P. putida V1, and (iii) the degradation pathways of methylmercury by P. putida V1. Strain V1 cultures readily degraded methylmercury, thimerosal, phenylmercury acetate, and reduced mercuric chloride into gaseous Hg(0). However, the Hg transformation in LB broth by P. putida V1 was influenced by the type of Hg compounds. The merA gene was detected in P. putida V1, on the other hand, the merB gene was not detected. The sequencing of this gene, showed high similarity (100%) to the mercuric reductase gene of other Pseudomonas spp. Furthermore, tests using radioactive (14)C-methylmercury indicated an uncommon release of (14)CO2 concomitant with the production of Hg(0). The results of the present work suggest that P. putida V1 has the potential to remove methylmercury from contaminated sites. More studies are warranted to determine the mechanism of removal of methylmercury by P. putida V1.

Functional metagenomics of oil-impacted mangrove sediments reveals high abundance of hydrolases of biotechnological interest

Mangroves are located in coastal wetlands and are susceptible to the consequences of oil spills, what may threaten the diversity of microorganisms responsible for the nutrient cycling and the consequent ecosystem functioning. Previous reports show that high concentration of oil favors the incidence of epoxide hydrolases and haloalkane dehalogenases in mangroves. This finding has guided the goals of this study in an attempt to broaden the analysis to other hydrolases and thereby verify whether oil contamination interferes with the prevalence of particular hydrolases and their assigned microorganisms. For this, an in-depth survey of the taxonomic and functional microbial diversity recovered in a fosmid library (Library_Oil Mgv) constructed from oil-impacted Brazilian mangrove sediment was carried out. Fosmid DNA of the whole library was extracted and submitted to Illumina HiSeq sequencing. The resulting Library Oil_Mgv dataset was further compared with those obtained by direct sequencing of environmental DNA from Brazilian mangroves (from distinct regions and affected by distinct sources of contamination), focusing on hydrolases with potential use in biotechnological processes. The most abundant hydrolases found were proteases, esterases and amylases, with similar occurrence profile in all datasets. The main microbial groups harboring such hydrolase-encoding genes were distinct in each mangrove, and in the fosmid library these enzymes were mainly assigned to Chloroflexaceae (for amylases), Planctomycetaceae (for esterases) and Bradyrhizobiaceae (for proteases). Assembly and analysis of Library_Oil Mgv reads revealed three potentially novel enzymes, one epoxide hydrolase, one xylanase and one amylase, to be further investigated via heterologous expression assays.

Microbial functional responses to long-term anthropogenic impact in mangrove soils

Mangroves are coastal ecosystems of transition between terrestrial and marine environments, that have been particularly contaminated in the last decades. Organic compounds are part of these contaminants, which have increased in the environment due to industrial activities and accidental oil spills. These contaminants are toxic to higher organisms, but microorganisms can metabolize most of these compounds and thus offer a tool for bioremediation purposes. The aim of the present study was to characterize the microbial potential and activity for degradation of aromatic compounds in sediment samples from mangroves using metagenomic and metatranscriptomic approaches. Sediment samples were collected for DNA and RNA extraction from each of the mangrove sites: highly oil-impacted (Oil Mgv), anthropogenically impacted (Ant Mgv) and pristine (Prs Mgv) mangrove. Hydrocarbon concentrations in Oil Mgv sediments were higher than those observed in Ant Mgv and Prs Mgv. Genes and transcripts associated with aromatic compound degradation, particularly the meta and ortho-pathways, were more abundant in Oil Mgv and Ant Mgv suggesting that many of the aromatic compounds are being aerobically degraded by the microbiome in these sites. Functions involved in the degradation of aromatic compounds were also found in pristine site, although in lower abundance. Members of the genera Aromatoleum, Desulfococcus, Desulfatibacillum, Desulfitobacterium and Vibrio were actively involved in the detoxification of sediments affected by the oil spill. Results obtained from this study provided strong evidence that microbial degradation of aromatic compounds plays an active role in the biological response to mangrove sediment pollution and subsequent ecosystem recovery.