The arsenic biogeochemical cycle is greatly dependent on microbial transformations that affect both the distribution and mobility of arsenic species in the environment. In this study, a microbial biofilm from volcanic rocks was characterized on the basis of its bacterial composition and ability to mobilize arsenic under circumneutral pH. Biofilm microstructure was analyzed by scanning electron microscopy (SEM)-energy-dispersive spectroscopy (EDS). Strains were isolated from biofilms and identified by 16S rDNA sequences analysis. Arsenic oxidation and reduction capacity was assayed with high-performance liquid chromatography coupled to gaseous formation performing the detection by atomic absortion in a quartz bucket (HPLC/HG/QAAS), and polymerase chain reaction was used to detect aox and ars genes. Bacterial communities associated with volcanic rocks were studied by denaturing gradient gel electrophoresis (DGGE). The SEM-EDS studies showed the presence of biofilm after 45 days of incubation. The relative closest GenBank matches of the DNA sequences, of isolated arsenic-resistant strains, showed the existence of four different genus: Burkholderia, Pseudomonas, Erwinia, and Pantoea. Four arsenite-resistant strains were isolates, and only three strains were able to oxidize >97% of the As(III) present (500 uM). All arsenate-resistant isolates were able to reduce between 69 and 86% of total As(V) (1000 uM). Analysis of 16S rDNA sequences obtained by DGGE showed the presence of four bacterial groups (∝-proteobacteria, γ-proteobacteria, Firmicutes, and Actinobacteria). Experiments demonstrate that epilithic bacterial communities play a key role in the mobilization of arsenic and metalloids speciation.
Roxarsone is an organoarsenical compound used as food additive in the poultry industry. Roxarsone has the potential risk to contaminate the environment, mainly by the use of poultry industry manure as fertilizer, releasing inorganic arsenic to the soil and water. The aim of this work was to isolate and characterize a bacterial consortium capable to degrade roxarsone under aerobic conditions. A bacterial consortium was cultured from a soil sample obtained from a field fertilized with poultry litter containing roxarsone. The consortium was cultured in the presence or absence of roxarsone. Roxarsone degradation and growth kinetics were determined by incubation of the bacterial consortium in the presence of roxarsone at room temperature and under aerobiosis. Both consortiums were characterized molecularly by denaturing gradient gel electrophoresis analysis and metabolically using Biolog Ecoplates. Inorganic arsenic was assessed by precipitation with silver nitrate. The consortium was also analyzed by scanning electron microscopy. The results showed that growth rate of the bacterial consortium was 1.4-fold higher in presence of roxarsone and 81.04 % of the roxarsone was transformed after 7 days of incubation. Molecular characterization revealed the presence of different bacterial groups, being alphaproteobacteria and firmicutes the groups that showed the highest count in both consortiums. The metabolic profile of the consortium did not change in the presence of roxarsone, but it showed a greater ability to oxidize amines, suggesting production of functional amines to decrease the stability of the aromatic ring resonance energy, the principal problem associated with aromatic compounds degradation.
Roxarsone is included in chicken food as anticoccidial and mainly excreted unchanged in faeces. Microorganisms biotransform roxarsone into toxic compounds that leach and contaminate underground waters used for human consumption. This study evaluated roxarsone biotransformation by underground water microorganisms and the toxicity of the resulting compounds. Underground water from an agricultural field was used to prepare microcosms, containing 0.05 mM roxarsone, and cultured under aerobic or anaerobic conditions. Bacterial communities of microcosms were characterized by PCR-DGGE. Roxarsone degradation was measured by HPLC/HG/AAS. Toxicity was evaluated using HUVEC cells and the Toxi-ChromoTest kit. Roxarsone degradation analysis, after 15 days, showed that microcosms of underground water with nutrients degraded 90 and 83.3% of roxarsone under anaerobic and aerobic conditions, respectively. Microcosms without nutrients degraded 50 and 33.1% under anaerobic and aerobic conditions, respectively. Microcosms including nutrients showed more roxarsone conversion into toxic inorganic arsenic species. DGGE analyses showed the presence of Proteobacteria, Firmicutes, Actinobacteria, Planctomycetes and Spirochaetes. Toxicity assays showed that roxarsone biotransformation by underground water microorganisms in all microcosms generated degradation products toxic for eukaryotic and prokaryotic cells. Furthermore, toxicity increased when roxarsone leached though a soil column and was further transformed by the bacterial community present in underground water. Therefore, using underground water from areas where roxarsone containing manure is used as fertilizer might be a health risk.
Arsenic (As), a highly toxic metalloid, naturally present in Camarones River (Atacama Desert, Chile) is a great health concern for the local population and authorities. In this study, the taxonomic and functional characterization of bacterial communities associated to metal-rich sediments from three sites of the river (sites M1, M2 and M3), showing different arsenic concentrations, were evaluated using a combination of approaches. Diversity of bacterial communities was evaluated by Illumina sequencing. Strains resistant to arsenic concentrations varying from 0.5 to 100 mM arsenite or arsenate were isolated and the presence of genes coding for enzymes involved in arsenic oxidation (aio) or reduction (arsC) investigated. Bacterial communities showed a moderate diversity which increased as arsenic concentrations decreased along the river. Sequences of the dominant taxonomic groups (abundances ≥1%) present in all three sites were affiliated to Proteobacteria (range 40.3–47.2%), Firmicutes (8.4–24.8%), Acidobacteria (10.4–17.1%), Actinobacteria (5.4–8.1%), Chloroflexi (3.9–7.5%), Planctomycetes (1.2–5.3%), Gemmatimonadetes (1.2–1.5%), and Nitrospirae (1.1–1.2%). Bacterial communities from sites M2 and M3 showed no significant differences in diversity between each other (p = 0.9753) but they were significantly more diverse than M1 (p<0.001 and p<0.001, respectively). Sequences affiliated with Proteobacteria, Firmicutes, Acidobacteria, Chloroflexi and Actinobacteria at M1 accounted for more than 89% of the total classified bacterial sequences present but these phyla were present in lesser proportions in M2 and M3 sites. Strains isolated from the sediment of sample M1, having the greatest arsenic concentration (498 mg kg-1), showed the largest percentages of arsenic oxidation and reduction. Genes aio were more frequently detected in isolates from M1 (54%), whereas arsC genes were present in almost all isolates from all three sediments, suggesting that bacterial communities play an important role in the arsenic biogeochemical cycle and detoxification of arsenical compounds. Overall, results provide further knowledge on the microbial diversity of arsenic contaminated fresh-water sediments.
Exopolysaccharide (EPS) production represents an adaptive strategy developed by extremophiles to cope with environmental stresses. The EPS-producing Bacillus licheniformis B3-15, of shallow marine vent origin (Vulcano Island, Italy), was previously reported as tolerant to arsenate (AsV). In this study, we evaluated: (i) the increasing production of EPS by Bacillus licheniformis B3-15 in the novel SG17 medium; (ii) the arsenic absorption capacity of the EPS by mass spectroscopy; (iii) the functional groups of EPS interacting with As by ATR-FTIR spectroscopy; and (iv) the ability of EPS to prevent arsenic toxicity by the bioluminescent assay. The EPS yield (240 mg L−1) was 45% higher than previously reported. The EPS was mainly constituted of disaccharide repeating units with a manno-pyranosidic configuration and low protein content, attributed to the poly-gamma glutamic acid component as evidenced by NMR analysis. ATR-FTIR spectra indicated that the functional groups of the EPS (O–H, C=O, C–O and C=C and N–O) were involved in the adsorption of the arsenic cations, with greater interactions between EPS and arsenate (AsV) than arsenite (AsIII). Consequently, the EPS at increasing concentration (100 and 300 µg mL−1) adsorbed AsV more efficiently (20.5% and 34.5%) than AsIII (0.7% and 1.8%). The bioluminescence assay showed that the EPS was not toxic, and its addition reduced the toxicity of both As forms by more than twofold. The crude EPS B3-15 could be used in arsenic bioremediation as a possible eco-friendly alternative to other physical or chemical methods.
The Patagonian Lakes have particular environmental conditions with or without intermittent disturbances. The study of the microorganisms present in aquatic ecosystems has increased notably because they can be used as micro-scale bioindicators of, among others, anthropogenic pollution and climatic change. The aim of the work was to compare the composition of the bacterial communities associated with sediments of three Patagonian Lakes with different geomorphologic patterns and disturbances. The lake sediments were characterized by molecular techniques, physiology profiles and physico-chemical analyses. The metabolic and physiological profiles of the microbial community demonstrated that non-impacted Tranquilo Lake is statistically different to impacted Bertrand and Plomo Lakes. Similar results were detected by DGGE profiles. FISH results demonstrated that betaproteobacteria showed the highest count in the Tranquilo Lake while gammaproteobacteria showed the highest counts in the Bertrand and Plomo Lakes, indicating that their sediments are highly dystrophic. The results demonstrate differences in the metabolic activity and structural and functional composition of bacterial communities of the studied lakes, which have different geomorphological patterns due to disturbances such as volcanic activity and the climatic change.
Pseudomonas arsenicoxydans has been recently described as a new arsenite oxidizing bacterial species. Arsenite detoxification activity by this species was determined by HPLC/HG/AAS. P. arsenicoxydans showed a high rate of As(III) conversion, particularly when immobilized (it oxidizes 100 % of 500 μg arsenite present in the medium after 48 of incubation). Arsenite oxidizing activity, mediated by a constitutive periplasmic enzyme, was determined following the transfer of reducing equivalents from arsenite to 2,4-dichlorophenolindophenol (DCIP) showing that approximately 75 % (0.173 µmol DCIP min(-1) mg(-1)) of the total activity (0.231 µmol DCIP min(-1) mg(-1)) was detected in the periplasmic fraction. Using PCR with primers specific for arsenite oxidase gene showed the presence of a gene encoding for arsenite oxidase in P. arsenicoxydans. Results show the potential biotechnological application of P. arsenicoxydans as a candidate for detoxification of As(III).
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