Multi-biomarker approach reveals the effects of heavy metal bioaccumulation in the foundation species Prosopis laevigata (Fabaceae).

Mining is a major economic activity in many developing countries. However, it disturbs the environment, producing enormous quantities of waste, known as mine tailings, which can have deleterious environmental impact, due to their high heavy metals (HM) content. Often, foundation species that establish on mine tailings are good candidates to study the effects of HM bioaccumulation at different levels of biological organization. Prosopis laevigata is considered a HM hyperaccumulator which presents attributes of a foundation species (FS) and establishes naturally on mine tailings. We evaluated the bioaccumulation of Cu, Pb, and Zn in P. laevigata foliar tissue, the leaf micro- and macro-morphological characters, DNA damage, and population genetic effects. In total, 80 P. laevigata individuals (20/site) belonging to four populations: The individuals from both sites (exposed and reference) bioaccumulated HMs (Pb > Cu > Zn). However, in the exposed individuals, Pb and Cu bioaccumulation was significantly higher. Also, a significant effect of macro- and micro-morphological characters was registered, showing significantly lower values in individuals from the exposed sites. In addition, we found significant differences in genotoxic damage in P. laevigata individuals, between the exposed and reference sites. In contrast, for the micro-morphological characters, none of the analyzed metals had any influence. P. laevigata did not show significant differences in the genetic structure and diversity between exposed and reference populations. However, four haplotypes and four private alleles were found in the exposed populations. Since P. laevigata is a species that establishes naturally in polluted sites and bioaccumulates HM in its foliar tissues, the resulting genetic, individual and population effects have not been severe enough to show detrimental effects; hence, P. laevigata can be a useful tool in phytoremediation strategies for soils polluted with Pb and Cu, maintaining its important ecological functions.

Glyphosate resistance and biodegradation by Burkholderia cenocepacia CEIB S5-2.

Glyphosate is a broad spectrum and non-selective herbicide employed to control different weeds in agricultural and urban zones and to facilitate the harvest of various crops. Currently, glyphosate-based formulations are the most employed herbicides in agriculture worldwide. Extensive use of glyphosate has been related to environmental pollution events and adverse effects on non-target organisms, including humans. Reducing the presence of glyphosate in the environment and its potential adverse effects requires the development of remediation and treatment alternatives. Bioremediation with microorganisms has been proposed as a feasible alternative for treating glyphosate pollution. The present study reports the glyphosate resistance profile and degradation capacity of the bacterial strain Burkholderia cenocepacia CEIB S5-2, isolated from an agricultural field in Morelos-México. According to the agar plates and the liquid media inhibition assays, the bacterial strain can resist glyphosate exposure at high concentrations, 2000 mg·L-1. In the degradation assays, the bacterial strain was capable of fast degrading glyphosate (50 mg·L-1) and the primary degradation metabolite aminomethylphosphonic acid (AMPA) in just eight hours. The analysis of the genomic data of B. cenocepacia CEIB S5-2 revealed the presence of genes that encode enzymes implicated in glyphosate biodegradation through the two metabolic pathways reported, sarcosine and AMPA. This investigation provides novel information about the potential of species of the genus Burkholderia in the degradation of the herbicide glyphosate and its main degradation metabolite (AMPA). Furthermore, the analysis of genomic information allowed us to propose for the first time a metabolic route related to the degradation of glyphosate in this bacterial group. According to the findings of this study, B. cenocepacia CEIB S5-2 displays a great glyphosate biodegradation capability and has the potential to be implemented in glyphosate bioremediation approaches.

Ecotoxicological effects of heavy metal bioaccumulation in two trophic levels.

The pollution generated by the heavy metals (HM) contained in mining wastes (tailings) is a worldwide recognized environmental concern. Due to the persistence, toxicity, bioaccumulation, and biomagnification capacity through the food chains, the release of HM into the environment causes negative effects on human health and the ecosystems. Wigandia urens Kunth (Boraginaceae) is a plant species that naturally establishes and grows in tailings and is consumed by the grasshopper Sphenarium purpurascens Charpentier (Orthoptera: Pyrgomorphidae). HM accumulation in this plant and their subsequent consumption by defoliating insects allow these contaminants to enter the food webs and favor their biomagnification. This study evaluated the effect of HM bioaccumulation in the leaf tissue of W. urens on the characteristics associated with its physical defense against herbivores and the effect of HM exposure on population parameters of grasshoppers through their ontogeny under controlled conditions. The results showed a significant increase in leaf hardness and in the number of simple and glandular trichomes in the leaves of W. urens growing on mine tailing substrate compared to those grown on the control substrate without HM. W. urens individuals growing on mine tailing substrate presented the following heavy metal foliar bioaccumulation pattern: Fe > Zn > Pb > Cu. These metals were also bioaccumulated in individuals of S. purpurascens fed with leaves of the plants exposed to mine tailings, observing differences in their concentration pattern through ontogeny. Grasshoppers fed on leaf tissue containing HM showed higher mortality in the first two developmental instars and lower body biomass throughout their ontogeny in comparison to the individuals fed on leaf tissue of plants growing on the control treatment without HM. In conclusion, W. urens is a species with phytoremediation potential for soils contaminated with HM, since it is naturally established in contaminated sites, has a wide geographic distribution, and bioaccumulates significant amounts of different HM. Furthermore, as was observed in this report, the W. urens physical and chemical defense against herbivores was enhanced by HM exposure, compromising the fitness and development of the herbivore S. purpurascens through its ontogeny and thus interrupting the entry and transfer of heavy metal through the food chain.

The bioaccumulation potential of heavy metals by Gliricidia sepium (Fabaceae) in mine tailings.

As a result of mining activities, waste of different types is generated. One example is mine tailings that contain potentially toxic elements such as heavy metals that negatively impact the environment and human health. Hence, developing treatments to guarantee its efficient elimination from the environment is necessary. Among these treatments, phytoremediation takes advantage of the potential of different plant species, to remove heavy metals from polluted sites. Gliricidia sepium is a tree that grows up to 15 m high and distributed from southern Mexico to Central America. This study evaluates the heavy metal bioaccumulation capacity in roots and leaves, and the effect of such bioaccumulation on fifteen macro- and one micro-morphological characters of G. sepium growing during 360 days in control, and in mine tailing substrates. G. sepium individuals growing on the exposed substrate registered the following average heavy metal bioaccumulation pattern in the roots: Fe > Pb > Zn > Cu, while in the leaf tissue, the bioaccumulation pattern was Cu > Fe > Pb > Zn. Macro- and micro-morphological characters evaluated in G. sepium decreased in plants exposed to metals. The translocation factor showed that Cu and Pb registered average values greater than 1. In conclusion, G. sepium is a species with potential for the phytoremediation of soils contaminated with Fe, Cu, and Pb, and for phytostabilizing soils polluted with Fe, Pb, Zn, and Cu, along with its ability to establish itself and turn into an abundant plant species in polluted sites, its capacity to bioaccumulate heavy metals in roots and leaves, and its high rate of HM translocation.

Dodonaea viscosa (Sapindaceae) as a phytoremediator for soils contaminated by heavy metals in abandoned mines.

Dodonaea viscosa (L.) Jacq. is a plant with a wide distribution that expands throughout almost all Mexican territory and is used in traditional medicine to treat many ailments. This species has been found associated with polluted areas, including mine tailings. Huautla, Morelos, Mexico, was a metallurgic district where mining activities generated 780,000 tons of waste rich in metals, deposited at 500 m from the town without any treatment; this situation has been related to different environmental threats and human health risks. The study was carried out for 18 months on seedlings developed under greenhouse conditions in two treatments: control substrate and mine tailings substrate. The concentration of six metals (Cd, Cr, Cu, Fe, Pb, and Zn) was measured through atomic absorption spectrophotometry in plant tissues, roots, and leaves. Effects of metal exposure were analyzed by size, micro-morphological character changes, and genetic damage in foliar tissue using the comet assay. The results showed significantly higher metal concentrations in the roots and leaves of individuals growing on the mine tailing substrate in comparison to the same plants tissues growing on control substrate. Positive and significant relationships between exposure time and metal concentration in roots and leaves, and between metal bioaccumulation in leaves and genetic damage were registered. Four out of six micro-morphological and size characters evaluated decreased significantly in exposed plants, except for stomatic index and root biomass. The most important metals in terms of the number of significantly affected micro-morphological and size characters showed the next pattern: Fe > Cd = Cr = Pb > Cu > Zn. D. viscosa is an efficient accumulator of Cu, Cd, Fe, Pb, and Zn in its root and leaf tissues. Overall, metal translocation factors in exposed D. viscosa plants showed the following pattern: Zn > Cu > Cd. We conclude that D. viscosa has the potential to phytoextract (Zn, Cu, and Cd), and phytostabilize (Cu, Cd, Fe, Pb, and Zn) metals from polluted soils, and along with its abundance, natural establishment in mine tailings, high levels of metal translocation, and bioconcentration factors, without affecting plant development, it can be an ideal candidate for phytoremediation of metal polluted soils.

Assisted Phytostabilization of Mine-Tailings with Prosopis laevigata (Fabaceae) and Biochar.

Phytoremediation is a cost-effective technique to remediate heavy metal (HM) polluted sites. However, the toxic effects of HM can limit plant establishment and development, reducing phytoremediation effectiveness. Therefore, the addition of organic amendments to mine wastes, such as biochar, improves the establishment of plants and reduces the bioavailability of toxic HM and its subsequent absorption by plants. Prosopis laevigata can establish naturally in mine tailings and accumulate different HM; however, these individuals show morphological and genetic damage. In this study, the effect of biochar on HM bioaccumulation in roots and aerial tissues, HM translocation, morphological characters and plant growth were evaluated, after three and six months of exposure. Plants grown on mine tailings with biochar presented significantly higher values for most of the evaluated characters, in respect to plants that grew on mine tailing substrate. Biochar addition reduced the bioaccumulation and translocation of Cu, Pb, and Cd, while it favored the translocation of essential metals such as Fe and Mn. The addition of biochar from agro-industrial residues to mine tailings improves the establishment of plants with potential to phytoextract and phytostabilize metals from polluted soils. Using biochar and heavy metal accumulating plants constitutes an assisted phytostabilization strategy with great potential for HM polluted sites such as Cd and Pb.

Proteomic analysis of Burkholderia zhejiangensis CEIB S4-3 during the methyl parathion degradation process.

Methyl parathion is an organophosphorus pesticide widely employed worldwide to control pests in agricultural and domestic environments. However, due to its intensive use, high toxicity, and environmental persistence, methyl parathion is recognized as an important ecosystem and human health threat, causing severe environmental pollution events and numerous human poisoning and deaths each year. Therefore, identifying and characterizing microorganisms capable of fully degrading methyl parathion and its degradation metabolites is a crucial environmental task for the bioremediation of pesticide-polluted sites. Burkholderia zhejiangensis CEIB S4-3 is a bacterial strain isolated from agricultural soils capable of immediately hydrolyzing methyl parathion at a concentration of 50 mg/L and degrading the 100% of the released p-nitrophenol in a 12-hour lapse when cultured in minimal salt medium. In this study, a comparative proteomic analysis was conducted in the presence and absence of methyl parathion to evaluate the biological mechanisms implicated in the methyl parathion biodegradation and resistance by the strain B. zhejiangensis CEIB S4-3. In each treatment, the changes in the protein expression patterns were evaluated at three sampling times, zero, three, and nine hours through the use of two-dimensional polyacrylamide gel electrophoresis (2D-PAGE), and the differentially expressed proteins were identified by mass spectrometry (MALDI-TOF). The proteomic analysis allowed the identification of 72 proteins with differential expression, 35 proteins in the absence of the pesticide, and 37 proteins in the experimental condition in the presence of methyl parathion. The identified proteins are involved in different metabolic processes such as the carbohydrate and amino acids metabolism, carbon metabolism and energy production, fatty acids ß-oxidation, and the aromatic compounds catabolism, including enzymes of the both p-nitrophenol degradation pathways (Hydroquinone dioxygenase and Hydroxyquinol 1,2 dioxygenase), as well as the overexpression of proteins implicated in cellular damage defense mechanisms such as the response and protection of the oxidative stress, reactive oxygen species defense, detoxification of xenobiotics, and DNA repair processes. According to these data, B. zhejiangensis CEIB S4-3 overexpress different proteins related to aromatic compounds catabolism and with the p-nitrophenol  degradation pathways, the higher expression levels observed in the two subunits of the enzyme Hydroquinone dioxygenase, suggest a preferential use of the Hydroquinone metabolic pathway in the p-nitrophenol degradation process. Moreover the overexpression of several proteins implicated in the oxidative stress response, xenobiotics detoxification, and DNA damage repair reveals the mechanisms employed by B. zhejiangensis CEIB S4-3 to counteract the adverse effects caused by the methyl parathion and p-nitrophenol exposure.

Glyphosate Pollution Treatment and Microbial Degradation Alternatives, a Review.

Glyphosate is a broad-spectrum herbicide extensively used worldwide to eliminate weeds in agricultural areas. Since its market introduction in the 70's, the levels of glyphosate agricultural use have increased, mainly due to the introduction of glyphosate-resistant transgenic crops in the 90's. Glyphosate presence in the environment causes pollution, and recent findings have proposed that glyphosate exposure causes adverse effects in different organisms, including humans. In 2015, glyphosate was classified as a probable carcinogen chemical, and several other human health effects have been documented since. Environmental pollution and human health threats derived from glyphosate intensive use require the development of alternatives for its elimination and proper treatment. Bioremediation has been proposed as a suitable alternative for the treatment of glyphosate-related pollution, and several microorganisms have great potential for the biodegradation of this herbicide. The present review highlights the environmental and human health impacts related to glyphosate pollution, the proposed alternatives for its elimination through physicochemical and biological approaches, and recent studies related to glyphosate biodegradation by bacteria and fungi are also reviewed. Microbial remediation strategies have great potential for glyphosate elimination, however, additional studies are needed to characterize the mechanisms employed by the microorganisms to counteract the adverse effects generated by the glyphosate exposure.

Transcriptomic analysis of Burkholderia cenocepacia CEIB S5-2 during methyl parathion degradation.

Methyl parathion (MP) is a highly toxic organophosphorus pesticide associated with water, soil, and air pollution events. The identification and characterization of microorganisms capable of biodegrading pollutants are an important environmental task for bioremediation of pesticide impacted sites. The strain Burkholderia cenocepacia CEIB S5-2 is a bacterium capable of efficiently hydrolyzing MP and biodegrade p-nitrophenol (PNP), the main MP hydrolysis product. Due to the high PNP toxicity over microbial living forms, the reports on bacterial PNP biodegradation are scarce. According to the genomic data, the MP- and PNP-degrading ability observed in B. cenocepacia CEIB S5-2 is related to the presence of the methyl parathion-degrading gene (mpd) and the gene cluster pnpABA'E1E2FDC, which include the genes implicated in the PNP degradation. In this work, the transcriptomic analysis of the strain in the presence of MP revealed the differential expression of 257 genes, including all genes implicated in the PNP degradation, as well as a set of genes related to the sensing of environmental changes, the response to stress, and the degradation of aromatic compounds, such as translational regulators, membrane transporters, efflux pumps, and oxidative stress response genes. These findings suggest that these genes play an important role in the defense against toxic effects derived from the MP and PNP exposure. Therefore, B. cenocepacia CEIB S5-2 has a great potential for application in pesticide bioremediation approaches due to its biodegradation capabilities and the differential expression of genes for resistance to MP and PNP.

Omics Approaches to Pesticide Biodegradation.

Pesticides are xenobiotic molecules necessary to control pests in agriculture, home, and industry. However, water and soil can become contaminated as a consequence of their extensive use. Therefore, because of its eco-friendly characteristics and efficiency, bioremediation of contaminated sites is a powerful tool with advantages over other kinds of treatments. For an efficient pesticides bioremediation, it is necessary to take into account different aspects related to the microbial metabolism and physiology. In this respect, OMICs studies such as genomics, transcriptomics, proteomics, and metabolomics are essential to generate relevant information about the genes and proteins involved in pesticide degradation, the metabolites generated by microbial pesticide degradation, and the cellular strategies to contend against stress caused by pesticide exposition. Pesticides as organochlorines and organophosphorus are the more commonly studied using OMIC approaches. To date, many genomes of microorganisms capable of degrading pesticides have been published, mainly bacterial strains from Burkholderia, Pseudomonas, and Rhodococcus genera. Following the genomic reports, transcriptomic studies, using microarrays and more recently next-generation sequencing technology RNA-Seq, in pesticide microbial degradation are the most numerous. Proteomics, metabolomics, as well as studies that combine different OMIC are gained interest. This review aims to describe a brief overview of pesticide biodegradation mechanisms; new tools to study microorganisms in natural environments; basic concepts of the OMICs approaches; as well as advances in methodologies associated with the analysis of that tools. Additionally, the most recent reports on genomics, transcriptomics, proteomics, and metabolomics during the degradation of pesticides are also analyzed.

Transcriptional analysis reveals the metabolic state of Burkholderia zhejiangensis CEIB S4-3 during methyl parathion degradation.

Burkholderia zhejiangensis CEIB S4-3 has the ability to degrade methyl parathion (MP) and its main hydrolysis byproduct p-nitrophenol (PNP). According to genomic data, several genes related with metabolism of MP and PNP were identified in this strain. However, the metabolic state of the strain during the MP degradation has not been evaluated. In the present study, we analyzed gene expression changes during MP hydrolysis and PNP degradation through a transcriptomic approach. The transcriptional analysis revealed differential changes in the expression of genes involved in important cellular processes, such as energy production and conversion, transcription, amino acid transport and metabolism, translation, ribosomal structure and biogenesis, among others. Transcriptomic data also exhibited the overexpression of both PNP-catabolic gene clusters (pnpABA'E1E2FDC and pnpE1E2FDC) present in the strain. We found and validated by quantitative reverse transcription polymerase chain reaction the expression of the methyl parathion degrading gene, as well as the genes responsible for PNP degradation contained in two clusters. This proves the MP degradation pathway by the strain tested in this work. The exposure to PNP activates, in the first instance, the expression of the transcriptional regulators multiple antibiotic resistance regulator and Isocitrate Lyase Regulator (IclR), which are important in the regulation of genes from aromatic compound catabolism, as well as the expression of genes that encode transporters, permeases, efflux pumps, and porins related to the resistance to multidrugs and other xenobiotics. In the presence of the pesticide, 997 differentially expressed genes grouped in 104 metabolic pathways were observed. This report is the first to describe the transcriptomic analysis of a strain of B. zhejiangensis during the biodegradation of PNP.

Penicillium sp. as an organism that degrades endosulfan and reduces its genotoxic effects.

Endosulfan is an organochloride and persistent pesticide that has caused concern because of its impact in the environment and its toxicity to and bioaccumulation in living organisms. In this study, we isolated an endosulfan-degrading fungus from the activated sludge from an industrial wastewater treatment plant. Through repetitive enrichment and successive subculture in media containing endosulfan as the sole carbon source, a fungus designated CHE 23 was isolated. Based on a phylogenetic analysis, strain CHE 23 was assigned to the genus Penicillium sp. In a mineral salt medium with 50 mg/l endosulfan as the sole source carbon, CHE 23 removed the added endosulfan in a period of six days. To verify the decrease in endosulfan toxicity due to the activity of the fungus, we performed genotoxicity tests trough the single cell gel electrophoresis assay or comet assay, with Eisenia fetida as the bioindicator species. This organism was exposed to the supernatants of the culture of the fungus and endosulfan. Our results indicated that the genotoxicity of endosulfan was completely reduced due the activity of this fungus. These results suggest that the Penicillium sp. CHE 23 strain can be used to degrade endosulfan residues and/or for water and soil bioremediation processes without causing toxicity problems, which are probably due to the generation of no-toxic metabolites during biodegradation.

Greenhouse gas emissions in the state of Morelos, Mexico: a first approximation for establishing mitigation strategies.

In this study, the authors report the first greenhouse gas emission inventory of Morelos, a state in central Mexico, in which the emissions of carbon dioxide (CO2), methane (CH4), and nitrous oxide (N2O) have been identified using the Intergovernmental Panel on Climate Change (IPCC) methodology. Greenhouse gas (GHG) emissions were estimated as CO2 equivalents (CO2 eq) for the years 2005, 2007, and 2009, with 2005 being treated as the base year. The percentage contributions from each category to the CO2 eq emissions in the base year were as follows: 38% from energy, 30% from industrial processes, 23% from waste, 5% from agriculture, and 4% from land use/land use change and forestry (LULUCF). As observed in other state inventories in Mexico, road transportation is the main source of CO2 emissions, wastewater handling and solid waste disposal are the main sources of CH4 emissions, and agricultural soils are the source of the most significant N2O emissions. The information reported in this inventory identifies the main emission sources. Based on these results, the government can propose public policies specifically designed for the state of Morelos to establish GHG mitigation strategies in the near future.

Isolation of the opdE gene that encodes for a new hydrolase of Enterobacter sp. capable of degrading organophosphorus pesticides.

Microbial enzymes that can hydrolyze organophosphorus compounds have been isolated, identified and characterized from different microbial species in order to use them in biodegradation of organophosphorus compounds. We isolated a bacterial strain Cons002 from an agricultural soil bacterial consortium, which can hydrolyze methyl-parathion (MP) and other organophosphate pesticides. HPLC analysis showed that strain Cons002 is capable of degrading pesticides MP, parathion and phorate. Pulsed-field gel electrophoresis and 16S rRNA amplification were performed for strain characterization and identification, respectively, showing that the strain Cons002 is related to the genus Enterobacter sp. which has a single chromosome of 4.6 Mb and has no plasmids. Genomic library was constructed from DNA of Enterobacter sp. Cons002. A gene called opdE (Organophosphate Degradation from Enterobacter) consists of 753 bp and encodes a protein of 25 kDa, which was isolated using activity methods. This gene opdE had no similarity to any genes reported to degrade organophosphates. When kanamycin-resistance cassette was placed in the gene opdE, hydrolase activity was suppressed and Enterobacter sp. Cons002 had no growth with MP as a nutrients source.