Hg-resistant Bacteria Research Articles

Isolation and characterization of mercury and multidrug-resistant Citrobacter freundii strains from tannery effluents in Kolkata, India.

Mercury (Hg) is one of the most potent toxic heavy metals that distresses livestock, humans, and ecological health. Owing to uncontrolled exposure to untreated tannery industrial effluents, metals such as Hg are increasing in nature and are, therefore, becoming a global concern. As a result, understanding the thriving microflora in that severe condition and their characteristics becomes immensely important. During the course of this study, two Hg-resistant bacteria were isolated from tannery wastewater effluents from leather factories in Kolkata, India, which were able to tolerate 2.211 × 10- 3 M (600µg/ml) Hg. 16S rDNA analysis revealed strong sequence homology with Citrobacter freundii, were named as BNC22A and BNC22C for this study. In addition they showed high tolerance to nickel (Ni) and Chromium (Cr) at 6.31 × 10- 3 M (1500µg/ml) and 6.792 × 10- 3 M (2000µg/ml) respectively. However, both the isolates were sensitive to arsenic (As) and cadmium (Cd). Furthermore, their antibiotic sensitivity profiles reveal a concerning trend towards resistance to multiple drugs. Overuse and misuse of antibiotics in healthcare systems and agriculture has been identified as two of the main reasons for the decline in efficacy of antibiotics. Though their ability to produce lipase makes them industrially potent organisms, their competence to resist several antibiotics and metals that are toxic makes this study immensely relevant. In addition, their ability to negate heavy metal toxicity makes them potential candidates for bioremediation. Finally, the green mung bean seed germination test showed a significant favourable effect of BNC22A and BNC22C against Hg-stimulated toxicity.

Isolation and Growth Optimization of Mercury-Resistant Bacteria Isolated from River Ravi

Mercury (Hg) is one of the most toxic heavy metals found on earth and has been reported as a contaminant from industrial and agricultural activities. Its release in the environment has led to extreme toxicity to all life forms. Various strategies like physico-chemical techniques have been proposed for Hg removal; bioremediation has proved to be most promising among all. This study focuses on the isolation of Hg resistant bacteria from Ravi River samples. Several biochemical tests were performed like gram staining, catalase, oxidase, sugar fermentation, antibiotic resistance, and motility test. Minimum inhibitory concentration (MIC) displayed that among all the isolates, Hg4 was able to resist up to 2500 ppm Hg followed by Hg6 and Hg7 which showed resistance till 2000 ppm Hg. These isolates were subjected to mercury reduction assay. Results revealed that Hg4, Hg6, and Hg7 bioremediated 59.82%, 40.82%, and 65.00% of mercury (Hg) respectively after 24 hours. In future, these isolates can be used for the bioremediation of Hg from contaminated sites which will help in creating a healthy environment.

Molecular Identification of Hg-Resistant Bacteria and Their Potential in Reducing Mercury Contamination

The aim of this research was to obtain and determine the identity of Hg-resistant bacteria in soil contaminatedwith gold processing waste and test its ability to reduce mercury contamination. Soil samples as a source of Hgresistant bacterial isolates were obtained from the gold processing location in Ilangata Village, Anggrek District,North Gorontalo Regency. The research was conducted at the Microbiology Laboratory, Department of Biology,Faculty of Mathematics and Natural Sciences. Mercury analysis was carried out at the Laboratory of FisheriesProduct Quality Development and Testing (LPPMHP), Gorontalo Province, and bacterial identification wascarried out at the Hasanuddin University Medical Research Center Research Unit. The parameters observed werethe types of Hg resistant bacteria and the ability of the bacteria to reduce mercury contamination. The resultsshowed that there were four bacterial isolates on the soil contaminated with 4.5 ppm mercury, which were namedILb01, ILB02, ILb03, and ILb04. Molecular identification showed that ILb01 was closely related toStenotrophomonas sp. SB67 and ILb02 close to Enterobacter cloacae strain CM 1, these strains were not resistantto mercury contamination; while ILb03 which is similar to strain BS0591 and ILb04 which is similar to BacteriumBacillus albus strain SQ30 16S could be resistant and was able to reduce mercury contamination by 99% at 10ppm levels. Key words: heavy metals, Hg-resistant bacteria, molecular identification, mercury

Genomic Characterization of a Mercury Resistant Arthrobacter sp. H-02-3 Reveals the Presence of Heavy Metal and Antibiotic Resistance Determinants

Nuclear production and industrial activities led to widespread contamination of the Department of Energy (DOE) managed Savannah River Site (SRS), located in South Carolina, United States. The H-02 wetland system was constructed in 2007 for the treatment of industrial and storm water runoff from the SRS Tritium Facility. Albeit at low levels, mercury (Hg) has been detected in the soils of the H-02 wetland ecosystem. In anoxic sediments, Hg is typically methylated by anaerobic microbiota, forming the highly neurotoxic methylmercury (MeHg), which biomagnifies across food webs. However, in surficial oxic wetland soils, microbially mediated demethylation and/or volatilization processes can transform Hg2+ into the less toxic Hg0 form which is released into the atmosphere, thus circumventing MeHg formation. To obtain a deeper understanding on bacterial Hg volatilization, a robust Hg-resistant (HgR) bacteria, called as strain H-02-3 was isolated from the H-02 soils. A draft genome sequence of this strain was obtained at a coverage of 700×, which assembled in 44 contigs with an N50 of 171,569 bp. The genomic size of the strain H-02-3 was 4,708,612 bp with a total number of 4,240 genes; phylogenomic analysis revealed the strain as an Arthrobacter species. Comparative genomics revealed the presence of 1100 unique genes in strain H-02-3, representing 26.7% of the total genome; many identified previously as metal resistance genes (MRGs). Specific to Hg-cycling, the presence of mercuric ion reductase (merA), the organomercurial lyase (merB), and the mercuric resistance operon regulatory protein, were identified. By inference, it can be proposed that the organomercurial lyase facilitates the demethylation of MeHg into Hg2+ which is then reduced to Hg0 by MerA in strain H-02-3. Furthermore, gene prediction using resistome analysis of strain H-02-3 revealed the presence of several antibiotic resistance genes (ARGs), that statistically correlated with the presence of metal resistant genes (MRGs), suggesting co-occurrence patterns of MRGs and ARGs in the strain. Overall, this study delineates environmentally beneficial traits that likely facilitates survival of Arthrobacter sp. H-02-3 within the H-02 wetland soil. Finally, this study also highlights the largely ignored public health risk associated with the co-development of ARGs and MRGs in bacteria native to historically contaminated soils.

Quantification and characterization of mercury resistant bacteria in sediments contaminated by artisanal small-scale gold mining activities, Kedougou region, Senegal

This study describes Hg-resistant bacterial present in the aquatic sediments artisanal small-scale gold mining ASGM activities along Gambia River Kedougou, Senegal. Mercury (Hg) is used for gold amalgamation in artisanal small-scale gold mining (ASGM) activities. The level of total Hg in sediment samples was determined by automatic mercury analyser. Bacterial (colony-forming units) susceptibility to Hg was evaluated by minimal inhibitory concentrations. The phylogenetic diversity analysis of the Hg-resistant bacteria was performed by PCR amplification of 16S rDNA on isolated bacterial strains, followed by restriction fragment length polymorphism, cloning and sequencing. The results documented high concentrations of Hg in ASGM activity areas ranging from 2.4 to 6.2 mg kg-1. Population densities of heterotrophic bacteria in wet sediment ranging from 3.7 × 106 to 4.6 × 108 CFU g−1. The isolated bacterial strains from highly Hg-contaminated sites can grow to medium containing up to 17 mg L−1 of Hg2+. In this study, bacterial strains resistant to Hg are Stenotrophomonas maltophilia, Dyella ginsengisoli, Arthrobacter defluvi, Arthrobacter pascens, Bacillus firmus and Pseudomonas moraviensis. Our results demonstrate the occurrence the presence of diverse groups of bacterial strains resistant to metal (Hg) under tropical conditions. The isolated strains are particularly interesting for further studies to evaluate their role in bioremediation of Hg in contaminated aquatic ecosystems.

Water-soluble mercury induced by organic amendments affected microbial community assemblage in mercury-polluted paddy soil

Mercury (Hg) pollution or organic amendments (OA) may individually induce changes in the microbial community of paddy soils. However, little is known regarding the interaction of Hg and OA and the effect of different OA applications on the microbial community assemblage in Hg-polluted paddy soil. A soil incubation experiment was performed by applying three organic amendments (OA), namely a food-waste compost (FC), and its HA and FA, into an Hg-polluted paddy soil to examine the changes in the microbial community and merA/merB gene abundance. The results showed that the OA treatments promoted total (SOC) and dissolved organic carbon (DOC) in soils, which may harbor copiotrophic bacteria. The HA and FA treatments decreased microbial diversity and richness along with an increase of water-soluble Hg (WHg) through the complexation of DOC to Hg, which may be mainly attributed to the enhanced Hg biotoxicity to soil microbiome induced by the increased WHg under these two treatments. Additionally, the WHg enhancement also contributed to the increase of Hg-resistant bacteria and merA/merB gene abundance, and consequently, induced changes in the microbial community. These results indicated the interaction of Hg and different OA induced the variation of WHg fraction in paddy soil, which played a fundamental role in the distinct responses of the microbial community assemblage. Collectively, the application of FA and HA to Hg-polluted soil should be limited considering Hg risk to microbiome, and FC can be an alternative.

Multiple Lines of Evidences Reveal Mechanisms Underpinning Mercury Resistance and Volatilization by Stenotrophomonas sp. MA5 Isolated from the Savannah River Site (SRS), USA.

A largely understudied microbially mediated mercury (Hg) bioremediative pathway includes the volatilization of Hg2+ to Hg0. Therefore, studies on Hg resistant bacteria (HgR), isolated from historically long-term contaminated environments, can serve as models to understand mechanisms underpinning Hg cycling. Towards this end, a mercury resistant bacterial strain, identified as Stenotrophomonas sp., strain MA5, was isolated from Mill Branch on the Savannah River Site (SRS); an Hg-impacted ecosystem. Minimum inhibitory concentration (MIC) analysis showed Hg resistance of up to 20 µg/mL by MA5 with 95% of cells retaining viability. Microcosm studies showed that the strain depleted more than 90% of spiked Hg2+ within the first 24 h of growth and the detection of volatilized mercury indicated that the strain was able to reduce Hg2+ to Hg0. To understand molecular mechanisms of Hg volatilization, a draft whole genome sequence was obtained, annotated and analyzed, which revealed the presence of a transposon-derived mer operon (merRTPADE) in MA5, known to transport and reduce Hg2+ into Hg0. Based on the whole genome sequence of strain MA5, qRT-PCR assays were designed on merRTPADE, we found a ~40-fold higher transcription of mer T, P, A, D and E when cells were exposed to 5 µg/mL Hg2+. Interestingly, strain MA5 increased cellular size as a function of increasing Hg concentrations, which is likely an evolutionary response mechanism to cope with Hg stress. Moreover, metal contaminated environments are shown to co-select for antibiotic resistance. When MA5 was screened for antibiotic resistance, broad resistance against penicillin, streptomycin, tetracycline, ampicillin, rifampicin, and erythromycin was found; this correlated with the presence of multiple gene determinants for antibiotic resistance within the whole genome sequence of MA5. Overall, this study provides an in-depth understanding of the underpinnings of Stenotrophomonas-mercury interactions that facilitate cellular survival in a contaminated soil habitat.

The potential of mercury resistant purple nonsulfur bacteria as effective biosorbents to remove mercury from contaminated areas

Abstract This study aimed to investigate the possibility to remove mercury (Hg) using the live or dead biomass of Hg resistant purple nonsulfur bacteria (PNSB) for their potential application to clean up shrimp ponds and during cultivation. The efficiency of Hg2+ removal by three PNSB strains; Rhodovulum sulfidophilum SRW1–5, and Afifella marina strains SSS2-1 and SSW15-1, grown with 2 different growth conditions (microaerobic light and aerobic dark conditions) were tested (2 mg/L HgCl2, 4.5 mg dry cell weight/mL, 30 min). It was found that the efficiency to remove Hg2+ by the dead cells of all strains was significantly higher than for live cells; and the most effective strain was SSS2-1. The highest Hg2+ removal under the optimum conditions using a 4 mg/L initial HgCl2 concentration by live and dead cells grown with both incubation conditions were approximately 87% and 95%, respectively. Under both incubation conditions, Hg2+ biosoption by strain SSS2-1 fitted the Freundlich model for live cells and the Langmuir model for dead cells. The kinetics of Hg2+ biosorption by both cell types of strain SSS2-1 suggests the process as a pseudo-second order kinetic model. This study demonstrates that biomass of the strain SSS2-1 has great potential for its biosorption to remove Hg2+ from contaminated areas like shrimp ponds.

Bacterial diversity associated with poplar trees grown on a Hg-contaminated site: Community characterization and isolation of Hg-resistant plant growth-promoting bacteria

Industrial waste dumps are rarely colonized by vegetation after they have been abandoned, indicating biological infertility. Revegetation of industrial tailings dumps is thus necessary to prevent wind erosion, metal leaching and has been shown to restore soil functions and ecosystem services. However, little is known about the microbial colonization and community structure of vegetated tailings following the application of restoration technologies. In this study, we investigated the rhizosphere and phyllosphere bacterial communities of a poplar tree plantation within a phytomanagement-based restoration program of a Hg-contaminated site. We used Illumina-based sequencing combined with culture-dependent approaches to describe plant-associated bacterial communities and to isolate growth-promoting bacteria (PGPB) and Hg-resistant bacteria. The genus Streptomyces was highly specific to the root community, accounting for 24.4% of the relative abundance but only representing 0.8% of the soil community, whereas OTUs from the Chloroflexi phylum were essentially detected in the soil community. Aboveground habitats were dominated by bacteria from the Deinococcus-Thermus phylum, which were not detected in belowground habitats. Leaf and stem habitats were characterized by several dominant OTUs, such as those from the phylum Firmicutes in the stems or from the genera Methylobacterium, Kineococcus, Sphingomonas and Hymenobacter in the leaves. Belowground habitats hosted more cultivable Hg-resistant bacteria than aboveground habitats and more Hg-resistant bacteria were found on the episphere than in endospheric habitats. Hg-resistant isolates exhibiting plant growth-promoting (PGP) traits, when used as inoculants of Capsicum annuum, were shown to increase its root dry biomass but not Hg concentration. The N2-fixing and Hg-resistant species Pseudomonas graminis, observed in the poplar phyllosphere, may be a key microorganism for the restoration of industrial tailings dumps.

Profiling of heavy metal(loid)-resistant bacterial community structure by metagenomic-DNA fingerprinting using PCR–DGGE for monitoring and bioremediation of contaminated environment

Frequent exposure of microbes to hazardous metalloids/heavy metals in contaminated environment results in the development of heavy metal(loid)-resistance properties. The study attempted to assess the profile of elevated arsenic (As), cadmium (Cd) and mercury (Hg)—resistant bacterial community structures of sludge (S1, India), sludge and sediment (S2 and S3, Japan) and sediment (S4, Vietnam) samples by metagenomic-DNA fingerprinting using polymerase chain reaction-denaturing gradient gel electrophoresis (PCR–DGGE) for monitoring and bioremediation of hazardous metal(loid) contamination in environment. The results revealed that As-resistant bacteria were dominant compared to Cd- and Hg-resistant bacteria with higher species diversity (Lysinibacillus sp., Uncultured soil bacterium clone, Staphylococcus sciuri, Bacillus fastidiosus, Bacillus niacini, Clostridium sp. and Bacillus sp.) in S1 and S4 than that of S2 and S3 samples. The occurrence of dominant As-resistant bacteria may indicate arsenic contamination in the investigated coastal habitats of India, Japan and Vietnam. The As-, Cd- and Hg-resistant bacteria/bacterial consortiums showed appreciable uptake ability of respective metal(loid) (0.042–0.125 mg As/l, 0.696–0.726 mg Cd/l and 0.34–0.412 mg Hg/l). Therefore, it might be concluded that the profiling of metalloids/heavy metal-resistant bacterial community structure by metagenomic-DNA fingerprinting using PCR–DGGE could be used to explore high metal(loid)-resistant bacteria for applying in metal(loid) bioremediation and as an indicator for monitoring hazardous metal(loid) contamination in environment.

Bioremediation of mercury: not properly exploited in contaminated soils!

Contamination of land and water caused by heavy metal mercury (Hg) poses a serious threat to biota worldwide. The seriousness of toxicity of this neurotoxin is characterized by its ability to augment in food chains and bind to thiol groups in living tissue. Therefore, different remediation approaches have been implemented to rehabilitate Hg-contaminated sites. Bioremediation is considered as cheaper and greener technology than the conventional physico-chemical means. Large-scale use of Hg-volatilizing bacteria are used to clean up Hg-contaminated waters, but there is no such approach to remediate Hg-contaminated soils. This review focuses on recent uses of Hg-resistant bacteria in bioremediation of mercury-contaminated sites, limitation and advantages of this approach, and identifies the gaps in existing research.

Assessment of heavy metal contamination and Hg-resistant bacteria in surface water from different regions of Delhi, India.

The present study aims to monitor the surface water quality of different regions in Delhi (India). With many physical and chemical properties, all samples had a high load of pollution in which Najafgarh drain (Nd) exhibited maximum and laboratory tap water (Ltw) minimum contamination. Water samples contained notable amounts of heavy metals including Cr, Cd, As, Cu, Pb and Hg. A total of 88 Hg-resistant bacteria were isolated from all the regions except Ltw. Among all the samples, the density of Hg-resistant bacteria was highest in sample of Nd and their morphotype heterogeneity was highest in sample collected from river Yamuna nearby Kashmiri gate (Kg). Different strains showed different patterns of resistance to different heavy metals and antibiotics. Multiple antibiotic resistance (MAR) indices were high in two samples, the highest reported in a sample taken from river Yamuna nearby Majnu ka tila (Mkt) (0.34). The 12.5% and 24.45% isolates showed β- and α-hemolytic natures, respectively that might be of pathogenic concern. In this account, high concentrations of heavy metals and their resistant bacteria in surface water have severely damaged the quality of water and their resources and produced high risk to the associated life forms.

Anaerobic Mercury Methylation and Demethylation by Geobacter bemidjiensis Bem.

Microbial methylation and demethylation are two competing processes controlling the net production and bioaccumulation of neurotoxic methylmercury (MeHg) in natural ecosystems. Although mercury (Hg) methylation by anaerobic microorganisms and demethylation by aerobic Hg-resistant bacteria have both been extensively studied, little attention has been given to MeHg degradation by anaerobic bacteria, particularly the iron-reducing bacterium Geobacter bemidjiensis Bem. Here we report, for the first time, that the strain G. bemidjiensis Bem can mediate a suite of Hg transformations, including Hg(II) reduction, Hg(0) oxidation, MeHg production and degradation under anoxic conditions. Results suggest that G. bemidjiensis utilizes a reductive demethylation pathway to degrade MeHg, with elemental Hg(0) as the major reaction product, possibly due to the presence of genes encoding homologues of an organomercurial lyase (MerB) and a mercuric reductase (MerA). In addition, the cells can strongly sorb Hg(II) and MeHg, reduce or oxidize Hg, resulting in both time and concentration-dependent Hg species transformations. Moderate concentrations (10-500 μM) of Hg-binding ligands such as cysteine enhance Hg(II) methylation but inhibit MeHg degradation. These findings indicate a cycle of Hg methylation and demethylation among anaerobic bacteria, thereby influencing net MeHg production in anoxic water and sediments.

The effect of aqueous speciation and cellular ligand binding on the biotransformation and bioavailability of methylmercury in mercury-resistant bacteria

Mercury resistant bacteria play a critical role in mercury biogeochemical cycling in that they convert methylmercury (MeHg) and inorganic mercury to elemental mercury, Hg(0). To date there are very few studies on the effects of speciation and bioavailability of MeHg in these organisms, and even fewer studies on the role that binding to cellular ligands plays on MeHg uptake. The objective of this study was to investigate the effects of thiol complexation on the uptake of MeHg by measuring the intracellular demethylation-reduction (transformation) of MeHg to Hg(0) in Hg-resistant bacteria. Short-term intracellular transformation of MeHg was quantified by monitoring the loss of volatile Hg(0) generated during incubations of bacteria containing the complete mer operon (including genes from putative mercury transporters) exposed to MeHg in minimal media compared to negative controls with non-mer or heat-killed cells. The results indicate that the complexes MeHgOH, MeHg-cysteine, and MeHg-glutathione are all bioavailable in these bacteria, and without the mer operon there is very little biological degradation of MeHg. In both Pseudomonas stutzeri and Escherichia coli, there was a pool of MeHg that was not transformed to elemental Hg(0), which was likely rendered unavailable to Mer enzymes by non-specific binding to cellular ligands. Since the rates of MeHg accumulation and transformation varied more between the two species of bacteria examined than among MeHg complexes, microbial bioavailability, and therefore microbial demethylation, of MeHg in aquatic systems likely depends more on the species of microorganism than on the types and relative concentrations of thiols or other MeHg ligands present.

Aerobic Mercury-resistant bacteria alter Mercury speciation and retention in the Tagus Estuary (Portugal)

Aerobic mercury-resistant bacteria were isolated from the sediments of two highly mercury-polluted areas of the Tagus Estuary (Barreiro and Cala do Norte) and one natural reserve area (Alcochete) in order to test their capacity to transform mercury. Bacterial species were identified using 16S rRNA amplification and sequencing techniques and the results indicate the prevalence of Bacillus sp. Resistance patterns to mercurial compounds were established by the determination of minimal inhibitory concentrations. Representative Hg-resistant bacteria were further tested for transformation pathways (reduction, volatilization and methylation) in cultures containing mercury chloride. Bacterial Hg-methylation was carried out by Vibrio fluvialis, Bacillus megaterium and Serratia marcescens that transformed 2–8% of total mercury into methylmercury in 48h. In addition, most of the HgR bacterial isolates showed Hg2+-reduction andHg0-volatilization resulting 6–50% mercury loss from the culture media.In summary, the results obtained under controlled laboratory conditions indicate that aerobic Hg-resistant bacteria from the Tagus Estuary significantly affect both the methylation and reduction of mercury and may have a dual face by providing a pathway for pollution dispersion while forming methylmercury, which is highly toxic for living organisms.

The Use of a Mercury Biosensor to Evaluate the Bioavailability of Mercury-Thiol Complexes and Mechanisms of Mercury Uptake in Bacteria.

As mercury (Hg) biosensors are sensitive to only intracellular Hg, they are useful in the investigation of Hg uptake mechanisms and the effects of speciation on Hg bioavailability to microbes. In this study, bacterial biosensors were used to evaluate the roles that several transporters such as the glutathione, cystine/cysteine, and Mer transporters play in the uptake of Hg from Hg-thiol complexes by comparing uptake rates in strains with functioning transport systems to strains where these transporters had been knocked out by deletion of key genes. The Hg uptake into the biosensors was quantified based on the intracellular conversion of inorganic mercury (Hg(II)) to elemental mercury (Hg(0)) by the enzyme MerA. It was found that uptake of Hg from Hg-cysteine (Hg(CYS)2) and Hg-glutathione (Hg(GSH)2) complexes occurred at the same rate as that of inorganic complexes of Hg(II) into Escherichia coli strains with and without intact Mer transport systems. However, higher rates of Hg uptake were observed in the strain with a functioning Mer transport system. These results demonstrate that thiol-bound Hg is bioavailable to E. coli and that this bioavailability is higher in Hg-resistant bacteria with a complete Mer system than in non-resistant strains. No difference in the uptake rate of Hg from Hg(GSH)2 was observed in E. coli strains with or without functioning glutathione transport systems. There was also no difference in uptake rates between a wildtype Bacillus subtilis strain with a functioning cystine/cysteine transport system, and a mutant strain where this transport system had been knocked out. These results cast doubt on the viability of the hypothesis that the entire Hg-thiol complex is taken up into the cell by a thiol transporter. It is more likely that the Hg in the Hg-thiol complex is transferred to a transport protein on the cell membrane and is subsequently internalized.

Isolation and Characterization of Mercury-Resistant Bacteria From Sediments of Tagus Estuary (Portugal): Implications for Environmental and Human Health Risk Assessment

Mercury (Hg) contamination of aquatic systems has been recognized as a global and serious problem affecting both human and environmental health. In the aquatic ecosystems, mercurial compounds are microbiologically transformed with methylation responsible for generation of methylmercury (MeHg) and subsequent biomagnification in food chain, consequently increasing the risk of poisoning for humans and wildlife. High levels of Hg, especially MeHg, are known to exist in Tagus Estuary as a result of past industrial activities. The aim of this study was to isolate and characterize Hg-resistant bacteria from Tagus Estuary. Mercury-resistant (Hg-R) bacteria were isolated from sediments of two hotspots (Barreiro and North Channel) and one reserve area (Alcochete). Mercury contamination in these areas was examined and bacterial susceptibility to Hg compounds evaluated by determination of minimal inhibitory concentrations (MIC). The isolates characterization was based on morphological observation and biochemical testing. Bacteria characteristics, distribution, and Hg resistance levels were compared with metal levels. Barreiro and North Channel were highly contaminated with Hg, containing 126 and 18 μg/g total Hg, respectively, and in Alcochete, contamination was lower at 0.87 μg/g total Hg. Among the isolates there were aerobic and anaerobic bacteria, namely, sulfate-reducing bacteria, and Hg resistance levels ranged from 0.16 to 140 μg/ml for Hg2+ and from 0.02 to 50.1 μg/ml for MeHg. The distribution of these bacteria and the resistance levels were consistent with Hg contamination along the depth of the sediments. Overall, results show the importance of the characterization of Tagus Estuary bacteria for ecological and human health risk assessment.

Antibiotic Resistance Pattern Among Gram Negative Mercury Resistant Bacteria Isolated From Contaminated Environments

Materials and Methods: The samples were collected from water and sediments of Kor River. We evaluated amounts of mercury in the water and sediment samples and counted the number of bacteria in both Hg containing and non-Hg containing media. Antibiotic resistance pattern was studied using disk diffusion method for Hg resistant and Hg sensitive bacteria. Results: The frequencies of mercury resistant bacteria were between 35% for Pole Khan station and 2.5% for Doroodzan station. These stations were the most and the lowest mercury-contaminated areas of the Kor River respectively. Pseudomonas sp., Entrobacter sp., Escherichia coli, Klebsiella sp. And Serratia marcescens were isolated as mercury resistant bacteria. The highest level of antibiotic resistance was seen for ampicillin, tetracycline and sterptomycin. Antibiotic resistance frequencies among these bacteria were higher than mercury sensitive ones. Conclusions: Our results showed that Hg resistance genes in contaminated areas are exchanged between residing bacteria along with the antibiotic resistance genes and resulted in prevalence of antibiotic resistance among residing bacteria.

Mercury induced community tolerance in microbial biofilms is related to pollution gradients in a long-term polluted river

The net toxicity of different forms of mercury, in the long-term during their transformation processes, leads to the selection of resistant bacterial cells and this result in community tolerance which is pollution induced. Accordingly, based on profiles of a bacterial community structure, analysis of Hg resistant culturable bacteria and quantification of merA genes, we assessed development of pollution induced community tolerance in a mercury-polluted gradient in the Idrijca River. TTGE analysis did not show effects of mercury pollution to bacterial community diversity, while quantification of merA genes showed that merA genes can be correlated precisely (R2=0.83) with the total concentration of mercury in the biofilm microbial communities in the pollution gradient.

Manila clams from Hg polluted sediments of Marano and Grado lagoons (Italy) harbor detoxifying Hg resistant bacteria in soft tissues

A mechanism of mercury detoxification has been suggested by a previous study on Hg bioaccumulation in Manila clams (Ruditapes philippinarum) in the polluted Marano and Grado lagoons and in this study we demonstrate that this event could be partly related to the detoxifying activities of Hg-resistant bacteria (MRB) harbored in clam soft tissues. Therefore, natural clams were collected in six stations during two different periods (winter and spring) from Marano and Grado Lagoons. Siphons, gills and hepatopancreas from acclimatized clams were sterile dissected to isolate MRB. These anatomical parts were glass homogenized or used for whole, and they were lying on a solid medium containing 5mgl−1 HgCl2 and incubated at 30°C. A total of fourteen bacterial strains were isolated and were identified by 16S rDNA sequencing and analysis, revealing that strains were representative of eight bacterial genera, four of which were Gram-positive (Enterococcus, Bacillus, Jeotgalicoccus and Staphylococcus) and other four were Gram-negative (Stenotrophomonas, Vibrio, Raoultella and Enterobacter). Plasmids and merA genes were found and their sequences determined. Fluorescence in situ hybridization (FISH) technique shows the presence of Firmicutes, Actinobacteria and Gammaproteobacteria by using different molecular probes in siphon and gills. Bacterial clumps inside clam flesh were observed and even a Gram-negative endosymbiont was disclosed by transmission electronic microscope inside clam cells. Bacteria harbored in cavities of soft tissue have mercury detoxifying activity. This feature was confirmed by the determination of mercuric reductase in glass-homogenized siphons and gills.