Mitigation effect of alpha-tocopherol and thermo-priming in Brassica napus L. under induced mercuric chloride stress.

Soil pollution with heavy metals has grown to be a big hassle, leading to the loss in farming production particularly in developing countries like Pakistan, where no proper channel is present for irrigation and extraction of these toxic heavy metals. The present study aims to ameliorate the damages caused by heavy metal ions (Hg-Mercury) on rapeseed (Brassica napus L.) via a growth regulator (α-tocopherol 150 mg/L) and thermopriming technique at 4 °C and 50 °C to maintain plant agronomical and physiological characteristics. In pot experiments, we designed total of 11 treatments viz.( T0 (control), T1 (Hg4ppm), T2 (Hg8ppm), T3 (Hg4ppm + 4 °C), T4 (Hg4ppm + 4 °C + tocopherol (150 m/L)), T5 (Hg4ppm + 50 °C), T6 (Hg4ppm + 50 °C + tocopherol (150 mg/L)), T7 (Hg8ppm + 4 °C), T8 (Hg8ppm + 4 °C + tocopherol (150 mg/L)), T9 (Hg8ppm + 50 °C), T10 (Hg8ppm + 50 °C + tocopherol (150 mg/L) the results revealed that chlorophyll content at p < 0.05 with growth regulator and antioxidant enzymes such as catalase, peroxidase, and malondialdehyde enhanced up to the maximum level at T5 = Hg4ppm + 50 °C (50 °C thermopriming under 4 ppm mercuric chloride stress), suggesting that high temperature initiate the antioxidant system to reduce photosystem damage. However, protein, proline, superoxide dismutase at p < 0.05, and carotenoid, soluble sugar, and ascorbate peroxidase were increased non-significantly (p > 0.05) 50 °C thermopriming under 8 ppm high mercuric chloride stress (T9 = Hg8ppm + 50 °C) representing the tolerance of selected specie by synthesizing osmolytes to resist oxidation mechanism. Furthermore, reduction in % MC (moisture content) is easily improved with foliar application of α-tocopherol and 50 °C thermopriming and 4 ppm heavy metal stress at T6 = Hg4ppm + 50 °C + α-tocopherol (150 mg/L), with a remarkable increase in plant vigor and germination energy. It has resulted that the inhibitory effect of only lower concentration (4 ppm) of heavy metal stress was ameliorated by exogenous application of α-tocopherol and thermopriming technique by synthesizing high levels of proline and antioxidant activities in maintaining seedling growth and development on heavy metal contaminated soil.

Effects of naringenin on oxidative damage and apoptosis in liver and kidney in rats subjected to chronic mercury chloride.

Mercury chloride is a type of heavy metal that causes the formation of free radicals, causing hepatotoxicity, nephrotoxicity and apoptosis. In this study, the effects of naringenin on oxidative stress and apoptosis in the liver and kidney of rats exposed to mercury chloride were investigated. In the study, 41 2-month-old male Wistar-Albino rats were divided into five groups. Accordingly, group 1 was set as control group, group 2 as naringenin-100, group 3 as mercury chloride, group 4 as mercury chloride + naringenin-50, and group 5 as mercury chloride + naringenin-100. For the interventions, 1 mL/kg saline was administered to the control, 0.4 mg/kg/day mercury (II) chloride to the mercury chloride groups by i.p., and 50 and 100 mg/kg/day naringenin prepared in corn oil to the naringenin groups by gavage. All the interventions lasted for 20 days. Mercury chloride administration was initiated 1 h following the administration of naringenin. When mercury chloride and the control group were compared, a significant increase in plasma urea, liver and kidney malondialdehyde (MDA) levels, in kidney superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), glutathione-S-transferase (GST) activities (p < .001), and a significant decrease in liver and kidney glutathione (GSH) levels (p < .001), in liver catalase (CAT) activity (p < .01) were observed. In addition, histopathological changes and a significant increase in caspase-3 levels were detected (p < .05). When mercury chloride and treatment groups were compared, the administration of naringenin caused a decrease aspartate transaminase (AST), alanine transaminase (ALT), lactate dehydrogenase (LDH) (p < .01), urea, creatinine levels (p < .001) in plasma, MDA levels in liver and kidney, SOD, GSH-Px, GST activities in kidney (p < .001), and increased GSH levels in liver and kidney. The addition of naringenin-100 increased GSH levels above the control (p < .001). The administration of naringenin was also decreased histopathological changes and caspase-3 levels (p < .05). Accordingly, it was determined that naringenin is protective and therapeutic against mercury chloride-induced oxidative damage and apoptosis in the liver and kidney, and 100 mg/kg naringenin is more effective in preventing histopathological changes and apoptosis.

Selenium alleviates endoplasmic reticulum calcium depletion-induced endoplasmic reticulum stress and apoptosis in chicken myocardium after mercuric chloride exposure.

Mercury is a highly toxic heavy metal with definite cardiotoxic properties and can affect the health of humans and animals through diet. Selenium (Se) is a heart-healthy trace element and dietary Se has the potential to attenuate heavy metal-induced myocardial damage in humans and animals. This study was designed to explore antagonistic effect of Se on the cardiotoxicity of mercuric chloride (HgCl2) in chickens. Hyline brown hens received a normal diet, a diet containing 250 mg/L HgCl2, or a diet containing 250 mg/L HgCl2 and 10 mg/kg Na2SeO3 for 7 weeks, respectively. Histopathological observations demonstrated that Se attenuated HgCl2-induced myocardial injury, which was further confirmed by the results of serum creatine kinase and lactate dehydrogenase levels assay and myocardial tissues oxidative stress indexes assessment. The results showed that Se prevented HgCl2-induced cytoplasmic calcium ion (Ca2+) overload and endoplasmic reticulum (ER) Ca2+ depletion mediated by Ca2+-regulatory dysfunction of ER. Importantly, ER Ca2+ depletion led to unfolded protein response and endoplasmic reticulum stress (ERS), resulting in apoptosis of cardiomyocytes via PERK/ATF4/CHOP pathway. In addition, heat shock protein expression was activated by HgCl2 through these stress responses, which was reversed by Se. Moreover, Se supplementation partially eliminated the effects of HgCl2 on the expression of several ER-settled selenoproteins, including selenoprotein K (SELENOK), SELENOM, SELENON, and SELENOS. In conclusion, these results suggested that Se alleviated ER Ca2+ depletion and oxidative stress-induced ERS-dependent apoptosis in chicken myocardium after HgCl2 exposure.

Mercury Chloride Affects Band 3 Protein-Mediated Anionic Transport in Red Blood Cells: Role of Oxidative Stress and Protective Effect of Olive Oil Polyphenols.

Mercury is a toxic heavy metal widely dispersed in the natural environment. Mercury exposure induces an increase in oxidative stress in red blood cells (RBCs) through the production of reactive species and alteration of the endogenous antioxidant defense system. Recently, among various natural antioxidants, the polyphenols from extra-virgin olive oil (EVOO), an important element of the Mediterranean diet, have generated growing interest. Here, we examined the potential protective effects of hydroxytyrosol (HT) and/or homovanillyl alcohol (HVA) on an oxidative stress model represented by human RBCs treated with HgCl2 (10 µM, 4 h of incubation). Morphological changes as well as markers of oxidative stress, including thiobarbituric acid reactive substance (TBARS) levels, the oxidation of protein sulfhydryl (-SH) groups, methemoglobin formation (% MetHb), apoptotic cells, a reduced glutathione/oxidized glutathione ratio, Band 3 protein (B3p) content, and anion exchange capability through B3p were analyzed in RBCs treated with HgCl2 with or without 10 µM HT and/or HVA pre-treatment for 15 min. Our data show that 10 µM HT and/or HVA pre-incubation impaired both acanthocytes formation, due to 10 µM HgCl2, and mercury-induced oxidative stress injury and, moreover, restored the endogenous antioxidant system. Interestingly, HgCl2 treatment was associated with a decrease in the rate constant for SO42- uptake through B3p as well as MetHb formation. Both alterations were attenuated by pre-treatment with HT and/or HVA. These findings provide mechanistic insights into benefits deriving from the use of naturally occurring polyphenols against oxidative stress induced by HgCl2 on RBCs. Thus, dietary supplementation with polyphenols might be useful in populations exposed to HgCl2 poisoning.

Mercuric chloride induces sequential activation of ferroptosis and necroptosis in chicken embryo kidney cells by triggering ferritinophagy.

Mercuric chloride (HgCl2) is an environmental pollutant with serious nephrotoxic effects, but the underlying mechanism of HgCl2 nephrotoxicity is not well understood. Ferroptosis and necroptosis are two programmed cell death (PCD) modalities that have been reported singly in heavy metal-induced kidney injury. However, the interaction between ferroptosis and necroptosis in HgCl2-induced kidney injury is unclear. Here, we established a model of HgCl2-exposed chicken embryo kidney (CEK) cells to dissect the progresses and mechanisms of these two PCDs. We found that ferroptosis was initially activated in CEK cells after HgCl2 exposure for 12 h, and necroptosis was activated subsequently at 24 h. Importantly, further study indicated that the shift from ferroptosis to necroptosis was driven by ROS, which was produced by iron-dependent Fenton reaction, and the iron chelation by DFO prevented the sequential activation of both ferroptosis and necroptosis. To investigate the source of intracellular iron, the regulation of iron homeostasis was first explored and demonstrated a tendency for intracellular iron overload in CEK cells. Interestingly, the cellular ferritin, a free iron depository, decreased in a time-dependent manner. Further studies revealed that the degradation of ferritin was attributed to the activation of selective cargo receptor nuclear receptor coactivator 4 (NCOA4)-mediated ferritinophagy, and the inhibition of ferritinophagy by CQ prevented the HgCl2-induced cell death. In conclusion, our study demonstrated that HgCl2 released excess free iron via ferritinophagy, led to a sustained accumulation of ROS and ultimately activated ferroptosis and necroptosis sequentially. These findings provide a new understanding for the nephrotoxic mechanism of HgCl2.

Protective mechanism of selenium on mercuric chloride-induced testis injury in chicken via p38 MAPK/ATF2/iNOS signaling pathway.

Mercuric chloride (HgCl2) is a well-known toxic heavy metal contaminant, which causes male reproductive function defects. Selenium (Se) has been recognized as an effective antioxidant against heavy metals-induced male reproductive toxicity. The aim of present study was to explore the potentially protective mechanism of Se on HgCl2-induced testis injury in chicken. Firstly, the results showed that Se mitigated HgCl2-induced testicular injury through increasing the blood-testis barrier (BTB) cell-junction proteins expression of occludin, zonula occludens-1 (ZO-1), connexin 43 (Cx43), and N-cadherin. Secondly, Se alleviated HgCl2-induced oxidative stress through decreasing the malondialdehyde (MDA) content and increasing the superoxidase dismutase (SOD), glutathione peroxidase (GSH-Px) activities as well as the total antioxidant capacity (T-AOC) level. Thirdly, Se inhibited the activation of p38 MAPK signaling through decreasing the proteins expression of phosphorylated-p38 (p-p38) and phosphorylated-ATF2 (p-ATF2), and alleviated inflammation response through decreasing the proteins expression of inducible nitric oxide synthase (iNOS), nuclear factor kappa B (NF-κB), tissue necrosis factor-alpha (TNF-α), and cyclooxygenase 2 (COX2). Collectively, these results demonstrated that Se effectively alleviated HgCl2-induced testes injury via improving antioxidant capacity to reduce inflammation mediated by p38 MAPK/ATF2/iNOS signaling pathway in chicken. Our data shed a new light on potential mechanisms of Se antagonized HgCl2-induced male reproductive toxicity.

Hepatic processing of mercuric ions facilitates delivery to renal proximal tubules.

Mercury (Hg) is a toxic heavy metal to which humans are exposed on a regular basis. Hg has a high affinity for thiol-containing biomolecules with the majority of Hg in blood being bound to albumin. The current study tested the hypothesis that circulating Hg-albumin complexes are taken up into hepatocytes and processed to form Hg-glutathione (GSH) conjugates (GSH-Hg-GSH). Subsequently, GSH-Hg-GSH conjugates are exported from hepatocytes into blood via multidrug resistance transporters (MRP) 3 and 5. To test this hypothesis, the portal vein and hepatic artery in Wistar rats were ligated to prevent delivery of Hg to the liver. Ligated and control rats were injected with HgCl2 or GSH-Hg-GSH (containing radioactive Hg) and the disposition of Hg was assessed in various organs. Renal accumulation of Hg was reduced significantly in ligated rats exposed to HgCl2. In contrast, when rats were exposed to GSH-Hg-GSH, the renal accumulation of Hg was similar in control and ligated rats. Experiments using HepG2 cells indicate that Hg-albumin conjugates are taken up by hepatocytes and additional experiments using inside-out membrane vesicles showed that MRP3 and MRP5 mediate the export of GSH-Hg-GSH from hepatocytes. These data are the first to show that Hg-albumin complexes are processed within hepatocytes to form GSH-Hg-GSH, which is, in part, exported back into blood via MRP3 and MRP5 for eventual excretion in urine.

Ziziphus spina-christi Leaf Extract Mitigates Mercuric Chloride-induced Cortical Damage in Rats.

BACKGROUND: Mercuric chloride (HgCl2) severely impairs the central nervous system when humans are exposed to it. AIMS: We investigated the neuroprotective efficiency of Ziziphus spina-christi leaf extract (ZSCLE) on HgCl2-mediated cortical deficits. METHODS: Twenty-eight rats were distributed equally into four groups: the control, ZSCLE-treated (300 mg/kg), HgCl2-treated (0.4 mg/kg), and ZSCLE+HgCl2-treated groups. Animals received their treatments for 28 days. RESULTS: Supplementation with ZSCLE after HgCl2 exposure prevented the deposition of mercury in the cortical slices. It also lowered malondialdehyde levels and nitrite and nitrate formation, elevated glutathione levels, activated its associated-antioxidant enzymes, glutathione reductase, and glutathione peroxidase, and upregulated the transcription of catalase and superoxide dismutase and their activities were accordingly increased. Moreover, ZSCLE activated the expression of nuclear factor erythroid 2-related factor 2 and heme oxygenase-1 when compared with the HgCl2 group. Notably, post-treatment with ZSCLE increased the activity of acetylcholinesterase and ameliorated the histopathological changes associated with HgCl2 exposure. Furthermore, ZSCLE blocked cortical inflammation, as observed by the lowered mRNA expression and protein levels of interleukin-1 beta and tumor necrosis factor-alpha, as well as decreased mRNA expression of inducible nitric oxide synthase. In addition, ZSCLE decreased neuron loss by preventing apoptosis in the cortical tissue upon HgCl2 intoxication. CONCLUSION: Based on the obtained findings, we suggest that ZSCLE supplementation could be applied as a neuroprotective agent to decrease neuron damage following HgCl2 toxicity.

Ziziphus spina-christi leaf extract attenuates mercuric chloride-induced liver injury in male rats via inhibition of oxidative damage.

Heavy metal contamination including mercury (Hg) has become one of the most serious environmental problems facing humans and other living organisms. Here, the hepatoprotective effects of Z. spina-christi leaf extract (ZCE) against inorganic mercury salt (mercuric chloride; HgCl2)-induced hepatotoxicity model was investigated in rats. Mercury concentration, liver function markers, oxidative stress markers, inflammation, cell death indicators, and histopathology were assessed. ZCE protected against HgCl2-induced hepatotoxicity, decreased Hg concentration, lipid peroxidation, and nitric oxide, increased glutathione, superoxide dismutase, catalase, and glutathione recycling enzymes (peroxidase and reductase), and upregulated nuclear factor-erythroid 2-related factor 2 (Nrf2) gene expression in HgCl2-intoxicated rat hepatic tissue. Nrf2 downstream gene and heme oxygenase-1 were also upregulated, confirming that hepatoprotection by ZCE against HgCl2-induced liver damage involved activation of the Nrf2/antioxidant response element pathway. ZCE also decreased the expression and production of pro-inflammatory cytokines and pro-apoptotic proteins and increased anti-apoptotic protein Bcl-2. Immunohistochemical analysis of liver tissues of HgCl2-treated rats confirmed the alternations of apoptotic-related protein expression. Our data demonstrated that post-administration of ZCE attenuated HgCl2-induced liver damage by activating the Nrf2/HO-1 signaling pathway. Therefore, administering this extract may be a novel therapeutic strategy for inorganic mercury intoxication.

Regulation of Single-Channel Conductance of Voltage-Dependent Anion Channel by Mercuric Chloride in a Planar Lipid Bilayer.

The existence of mercury in various forms, e.g., elemental, organic, and inorganic has been known for decades. In any of these forms, it is poisonous to metabolism. In this, an investigation about the effect of the inorganic form of mercury, i.e., mercuric chloride (HgCl2) to the mitochondrial voltage-dependent anion channel (VDAC), has been done after isolation from the cardiac and brain tissues of Wistar rats. In vitro electrophysiology experiments were performed in Cardiolipin planar lipid bilayer membrane (BLM) to study the change in the conductance, selectivity, and gating charge of VDAC post HgCl2 treatment. A reduction in mean conductance of VDAC from 4.3 ± 0.18 to 1.66 ± 0.11 nS was observed. Further, the Gating charge calculated before (± 3.5) and after HgCl2 treatment (± 2.3) showed significant difference. Later, VDAC's behavior was studied at different concentrations of HgCl2 ranging from 0.1 µM to 1 mM. The Inhibitory concentration (IC50) was calculated from the linear regression plot. The IC50 was found to be 488.1 µM. In the asymmetrical HgCl2 (5:1), a permeability ratio of cation to anion was found to be 4.2. It is interpreted that VDAC functioning is affected due to the application of 4 mM HgCl2 and a reduction in the conductance, gating charge, and permeability of VDAC was detected. The results provide clues to HgCl2-induced toxicity mediated through VDAC in the Cardiolipin BLM.

Interventions of aqueous extract of Solanum melongena fruits (garden eggs) on mercury chloride induced testicular toxicity in adult male Wistar rats.

BACKGROUND: Solanum melongena (SM) is commonly known as the garden egg fruit or eggplant. It can be eaten fresh or cooked and has a large history of consumption in West Africa. This study focused on interventions of aqueous extract of SM (garden eggs) fruits on Mercury chloride (HgCl2) induced testicular toxicity in adult male Wistar rats. METHODS: Thirty-two adult male Wistar rats were randomly divided into four groups (A-D) of eight (n = 8) rats each. Group A Served as control and was given 10 ml/kg/day of distilled water, Group B- 500 mg/kg B.W of SM, Group C received 40 mg/kg B.W HgCl2 and Group D- 500 mg/kg B.W of SM and 40 mg/kg B.W HgCl2). The administration was done by gastric gavage once a day, for twenty-eight consecutive days. Testicular weight, semen analysis revealing the sperm count and sperm motility were assessed, gross parameters of the testis and testicular histology were assessed. Testicular oxidative stress markers viz a viz malondialdehyde (MDA), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx) and reduced glutathione (GSH) were also assessed. RESULTS: assessment of the histological profiles of the testes showed a derangement of the cytoarchitecture and deterioration of sperm quality after HgCl2 administration and a marked improvement was observed after SM administration. Similarly, SM was associated with increased antioxidant parameters (SOD, CAT, GPx, and GSH) and decreased MDA in SM + HgCl2 rats. CONCLUSION: It was concluded that S. melongena offers protection against free radical mediated oxidative stress of rats with mercury chloride induced testicular toxicity.

Plasma Membrane Aquaporin Members PIPs Act in Concert to Regulate Cold Acclimation and Freezing Tolerance Responses in Arabidopsis thaliana.

Aquaporins play a major role in plant water uptake at both optimal and environmentally stressed conditions. However, the functional specificity of aquaporins under cold remains obscure. To get a better insight to the role of aquaporins in cold acclimation and freezing tolerance, we took an integrated approach of physiology, transcript profiling and cell biology in Arabidopsis thaliana. Cold acclimation resulted in specific upregulation of PIP1;4 and PIP2;5 aquaporin (plasma membrane intrinsic proteins) expression, and immunoblotting analysis confirmed the increase in amount of PIP2;5 protein and total amount of PIPs during cold acclimation, suggesting that PIP2;5 plays a major role in tackling the cold milieu. Although single mutants of pip1;4 and pip2;5 or their double mutant showed no phenotypic changes in freezing tolerance, they were more sensitive in root elongation and cell survival response under freezing stress conditions compared with the wild type. Consistently, a single mutation in either PIP1;4 or PIP2;5 altered the expression of a number of aquaporins both at the transcriptional and translational levels. Collectively, our results suggest that aquaporin members including PIP1;4 and PIP2;5 function in concert to regulate cold acclimation and freezing tolerance responses.

Mercury chloride exposure induces DNA damage, reduces fertility, and alters somatic and germline cells in Drosophila melanogaster ovaries.

Mercury exposure has been shown to affect the reproductive system in many organisms, although the molecular mechanisms are still elusive. In the present study, we exposed Drosophila melanogaster Canton-S adult females to concentrations of 0 mM, 0.1 mM, 0.3 mM, 3 mM, and 30 mM of mercury chloride (HgCl2) for 24 h, 48 h, or 72 h to determine how mercury could affect fertility. Alkaline assays performed on dissected ovaries showed that mercury induced DNA damage that is not only dose-dependent but also time-dependent. All ovaries treated for 72 h have incorporated mercury and exhibit size reduction. Females treated with 30 mM HgCl2, the highest dose, had atrophied ovaries and exhibited a drastic 7-fold reduction in egg laying. Confocal microscopy analysis revealed that exposure to HgCl2 disrupts germinal and somatic cell organization in the germarium and leads to the aberrant expression of a germline-specific gene in somatic follicle cells in developing egg chambers. Together, these results highlight the potential long-term impact of mercury on germline and ovarian cells that might involve gene deregulation.

L-α-Phosphatidylcholine attenuates mercury-induced hepato-renal damage through suppressing oxidative stress and inflammation.

The potential ameliorative effects of L-α-phosphatidylcholine (PC) against mercuric chloride (HgCl2)-induced hematological and hepato-renal damage were investigated. Rats were randomly allocated into four groups (n = 12): control, PC (100 mg/kg bwt, intragastrically every other day for 30 consecutive days), HgCl2 (5 mg/kg bwt, intragastrically daily), and PC plus HgCl2. Hematological and hepato-renal dysfunctions were evaluated biochemically and histopathologically. Hepatic and renal oxidative/antioxidative indices were evaluated. The expression of proinflammatory cytokines (tumor necrosis factor-α and interleukin-6) was also detected by ELISA. HgCl2 significantly increased serum aminotransferases (ALT, AST), urea, and creatinine levels that are indicative of hepato-renal damage. HgCl2 also induced a significant accumulation of malondialdehyde (+ 195%) with depletion of glutathione (- 43%) levels in the liver and renal tissues. The apparent hepato-renal oxidative damage was associated with obvious organ dysfunction that was confirmed by impairments in the liver and kidney histoarchitecture. Furthermore, HgCl2 significantly attenuated the expression of proinflammatory cytokines named tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). Conversely, PC treatment attenuated these effects, which improved the hematological and serum biochemical alternations, reduced the oxidative stress and proinflammatory cytokine levels, and ameliorated the intensity of the histopathological alterations in livers and kidneys of HgCl2-treated rats. It could be concluded that PC displayed potential anti-inflammatory and antioxidant activities against HgCl2-induced hepato-renal damage via suppression of proinflammatory cytokines and declining oxidative stress.

Expression and Purification of Transmembrane Protein MerE from Mercury-Resistant Bacillus cereus.

Mercury-resistant (HgR) bacteria were isolated from heavy metal polluted wastewater and soil collected near to tanneries of district Kasur, Pakistan. Bacterial isolates AZ-1, AZ-2 and AZ-3 showed resistance up to 40 µg/ml against mercuric chloride (HgCl2). 16S rDNA ribotyping and phylogenetic analysis were performed for the characterization of selected isolates as Bacillus sp. AZ-1 (KT270477), Bacillus cereus AZ-2 (KT270478) and Bacillus cereus AZ-3 (KT270479). Phylogenetic relationship on the basis of merA nucleotide sequence confirmed 51- 100% homology with the corresponding region of the merA gene of already reported mercuryresistant Gram-positive bacteria. The merE gene involved in the transportation of elemental mercury (Hg0) via cell membrane was cloned for the first time into pHLV vector and transformed in overexpressed C43(DE3) E. coli cells. The recombinant plasmid (pHLMerE) was expressed and the native MerE protein was obtained after thrombin cleavage by size exclusion chromatography (SEC). The purification of fusion/recombinant and native protein MerE by Ni-NTA column, dialysis and fast protein liquid chromatography (FPLC/SEC) involved unfolding/refolding techniques. A small-scale reservoir of wastewater containing 30 µg/ml of HgCl2 was designed to check the detoxification ability of selected strains. It resulted in 83% detoxification of mercury by B. cereus AZ-2 and B. cereus AZ-3, and 76% detoxification by Bacillus sp. AZ-1 respectively (p < 0.05).

A polysaccharide-peptide with mercury clearance activity from dried fruiting bodies of maitake mushroom Grifola frondosa.

Mercury is considered to be "a global pollutant" and raises concern worldwide. Once mercury enters the body, it will be distributed all over the body but will accumulate in the brain, kidney and liver. To date, no substance originating from edible fungi capable of adsorbing mercury has been reported. We found that the mushroom Grifola frondosa exhibited mercury adsorption capacity. A polysaccharide-peptide (GFPP), displaying the unique N-terminal amino acid sequence of APPGMHQKQQ and 7 partial sequences with high reliability obtained by LC-MS/MS, was isolated by hot-water extraction of its fruiting bodies followed by ion exchange chromatography and gel filtration chromatography. Two rat models were employed to determine the dose and the duration of HgCl2 treatment (given by acute administration or continuous treatment) to test if G. frondosa could promote mercury elimination. For rats subjected to acute treatment with HgCl2, both GFPP and G. frondosa fruiting bodies (GFFF) could accelerate the decline of blood mercury level, which fell precipitously by 50% on the second day. GFPP and GFFF also promoted elimination of the burden of mercury in the liver and kidneys. For rats receiving continuous HgCl2 treatment, G. frondosa prevented the progressive increase of blood mercury level, and kept the blood mercury level within a relatively stable range.

Robust Mercury Methylation across Diverse Methanogenic Archaea.

Methylmercury (MeHg) production was compared among nine cultured methanogenic archaea that contain hgcAB, a gene pair that codes for mercury (Hg) methylation. The methanogens tested produced MeHg at inherently different rates, even when normalized to growth rate and Hg availability. Eight of the nine tested were capable of MeHg production greater than that of spent- and uninoculated-medium controls during batch culture growth. Methanococcoides methylutens, an hgcAB+ strain with a fused gene pair, was unable to produce more MeHg than controls. Maximal conversion of Hg to MeHg through a full batch culture growth cycle for each species (except M. methylutens) ranged from 2 to >50% of the added Hg(II) or between 0.2 and 17 pmol of MeHg/mg of protein. Three of the species produced >10% MeHg. The ability to produce MeHg was confirmed in several hgcAB+ methanogens that had not previously been tested (Methanocella paludicola SANAE, Methanocorpusculum bavaricum, Methanofollis liminatans GKZPZ, and Methanosphaerula palustris E1-9c). Maximal methylation was observed at low sulfide concentrations (<100 µM) and in the presence of 0.5 to 5 mM cysteine. For M. hollandica, the addition of up to 5 mM cysteine enhanced MeHg production and cell growth in a concentration-dependent manner. As observed for bacterial Hg methylators, sulfide inhibited MeHg production. An initial evaluation of sulfide and thiol impacts on bioavailability showed methanogens responding to Hg complexation in the same way as do Deltaproteobacteria The mercury methylation rates of several methanogens rival those of the better-studied Hg-methylating sulfate- and iron-reducing DeltaproteobacteriaIMPORTANCEArchaea, specifically methanogenic organisms, play a role in mercury methylation in nature, but their global importance to MeHg production and the subsequent risk to ecosystems are not known. Methanogenesis has been linked to Hg methylation in several natural habitats where methylmercury production incurs risk to people and ecosystems, including rice paddies and permafrost. In this study, we confirm that most methanogens carrying the hgcAB gene pair are capable of Hg methylation. We found that methylation rates vary inherently among hgcAB+ methanogens but that several species are capable of MeHg production at rates that rival those of the better-know Hg-methylating sulfate- and iron-reducing bacteria. Methanogens may need to be considered equally with sulfate and iron reducers in evaluations of MeHg production in nature.

Mercury tolerance and biosorption in bacteria isolated from Ny-Ålesund, Svalbard, Arctic.

Mercury tolerant bacteria Pseudarthrobacter oxydans strain MM20 and Pseudomonas frederiksbergensis strain SS18 were isolated from the tundra ecosystem of Ny-Ålesund, Svalbard, where commercial exploitation of the coal existed till 1960s. Minimum inhibitory concentration (MIC), mercury removal, mercury biosorption, and antibiotic resistance of these strains were analyzed. P. frederiksbergensis strain SS18 showed high tolerance (2.0 ppm) to mercury than P. oxydans strain MM20 (1.5 ppm). Mercury removal and biosorption studies were carried out in liquid media containing 1.0 ppm mercury. More than 90% of mercury was removed from the culture media by the selected strains. The mercury biosorption assay revealed that a part of mercury was accumulated in cell pellets and was 22 and 25% respectively for P. oxydans strain MM20 and P. frederiksbergensis strain SS18. Fourier transform infrared study revealed that alkyl halide, alkynes, alcoholic, aliphatic and aromatic amines, alkanes, nitro compound, primary amines, carboxylic acid, alkenes, and amide groups play a major role in the development of tolerance towards mercury. Out of eleven antibiotics tested, P. oxydans strain MM20 was found to be resistant to lincomycin and novobiocin while P. frederiksbergensis strain SS18 was found to be resistant to seven antibiotics. Our study demonstrates that under experimental conditions, bacterial isolates undergo detailed structural and functional changes to tolerate as well as immobilize toxic elements like mercury.

Mercury (II) impairs nucleotide excision repair (NER) in zebrafish (Danio rerio) embryos by targeting primarily at the stage of DNA incision.

Mercuric ion (Hg2+) is the most prevalent form of inorganic Hg found in polluted aquatic environment. As inhibition of DNA damage repair has been proposed as one of the mechanisms of Hg2+-induced genotoxicity in aquatic animals and mammalian cells, this study explored the susceptibility of different stages of nucleotide excision repair (NER) in zebrafish (Danio rerio) embryos to Hg2+ using UV-damaged DNA as the repair substrate. Exposure of embryos at 1h post fertilization (hpf) to HgCl2 at 0.1-2.5µM for 9h caused a concentration-dependent inhibition of NER capacity monitored by a transcription-based DNA repair assay. The extracts of embryos exposed to 2.5µM Hg2+ almost failed to up-regulate UV-suppressed marker cDNA transcription. No inhibition of ATP production was observed in all Hg2+-exposed embryos. Hg2+ exposure imposed either weak inhibitory or stimulating effects on the gene expression of NER factors, while band shift assay showed the inhibition of photolesion binding activities to about 40% of control in embryos treated with 1-2.5µM HgCl2. The damage incision stage of NER in zebrafish embryos was found to be more sensitive to Hg2+ than photolesion binding capacity due to the complete loss of damage incision activity in the extracts of embryos exposed to 1-2.5µM Hg2+. NER-related DNA incision was induced in UV-irradiated embryos based on the production of short DNA fragments matching the sizes of excision products generated by eukaryotic NER. Pre-exposure of embryos to Hg2+ at 0.1-2.5µM all suppressed DNA incision/excision in UV-irradiated embryos, reflecting a high sensitivity of DNA damage incision/excision to Hg2+. Our results showed the potential of Hg2+ at environmental relevant levels to disturb NER in zebrafish embryos by targeting primarily at the stage of DNA incision/excision.

Zuotai and HgS differ from HgCl2 and methyl mercury in Hg accumulation and toxicity in weanling and aged rats.

Mercury sulfides are used in Ayurvedic medicines, Tibetan medicines, and Chinese medicines for thousands of years and are still used today. Cinnabar (α-HgS) and metacinnabar (ß-HgS) are different from mercury chloride (HgCl2) and methylmercury (MeHg) in their disposition and toxicity. Whether such scenario applies to weanling and aged animals is not known. To address this question, weanling (21d) and aged (450d) rats were orally given Zuotai (54% ß-HgS, 30mg/kg), HgS (α-HgS, 30mg/kg), HgCl2 (34.6mg/kg), or MeHg (MeHgCl, 3.2mg/kg) for 7days. Accumulation of Hg in kidney and liver, and the toxicity-sensitive gene expressions were examined. Animal body weight gain was decreased by HgCl2 and to a lesser extent by MeHg, but unaltered after Zuotai and HgS. HgCl2 and MeHg produced dramatic tissue Hg accumulation, increased kidney (kim-1 and Ngal) and liver (Ho-1) injury-sensitive gene expressions, but such changes are absent or mild after Zuotai and HgS. Aged rats were more susceptible than weanling rats to Hg toxicity. To examine roles of transporters in Hg accumulation, transporter gene expressions were examined. The expression of renal uptake transporters Oat1, Oct2, and Oatp4c1 and hepatic Oatp2 was decreased, while the expression of renal efflux transporter Mrp2, Mrp4 and Mdr1b was increased following HgCl2 and MeHg, but unaffected by Zuotai and HgS. Thus, Zuotai and HgS differ from HgCl2 and MeHg in producing tissue Hg accumulation and toxicity, and aged rats are more susceptible than weanling rats. Transporter expression could be adaptive means to reduce tissue Hg burden.