Occurrence of two structural types of mercury reductases among Gram-positive bacteria

Structural variants of mercury reductase containing the N-terminal domain, which is easily cleaved by trypsin, have been found in Gram-positive bacteria with a low genomic G + C content (Bacillus, Staphylococcus and, possibly, some other genera). Mercury reductases without the N-terminal domain and relatively resistant to limited proteolysis are typical for Gram-positive bacteria with a high genomic G + C content (Arthrobacter, Citreobacterium, Micrococcus, Mycobacterium, Rhodococcus). Both types of mercury reductase genes may be located on plasmids.     

Methods and recent advances in speciation analysis of mercury chemical species in environmental samples: a review

Mercury (Hg) and its compounds are much concerned for their high toxicity and wide presence in the environment. Since the toxicity of Hg is species dependent, various methods have been developed for the speciation analysis of Hg. This review focus on the determination and speciation analysis of Hg chemical species in water, sediment, and soil samples. Recent developments on sample pre-treatment and extraction/pre-concentration, separation, and quantification of Hg chemical species, and associated analytical challenges have been reviewed and briefly discussed based on recent reports.     

Diversity of purple nonsulfur bacteria in shrimp ponds with varying mercury levels

This research aimed to study the diversity of purple nonsulfur bacteria (PNSB) and to investigate the effect of Hg concentrations in shrimp ponds on PNSB diversity. Amplification of the pufM gene was detected in 13 and 10 samples of water and sediment collected from 16 shrimp ponds in Southern Thailand. In addition to PNSB, other anoxygenic phototrophic bacteria (APB) were also observed; purple sulfur bacteria (PSB) and aerobic anoxygenic phototrophic bacteria (AAPB) although most of them could not be identified. Among identified groups; AAPB, PSB and PNSB in the samples of water and sediment were 25.71, 11.43 and 8.57%; and 27.78, 11.11 and 22.22%, respectively. In both sample types, Roseobacter denitrificans (AAPB) was the most dominant species followed by Halorhodospira halophila (PSB). In addition two genera, observed most frequently in the sediment samples were a group of PNSB (Rhodovulum kholense, Rhodospirillum centenum and Rhodobium marinum). The UPGMA dendrograms showed 7 and 6 clustered groups in the water and sediment samples, respectively. There was no relationship between the clustered groups and the total Hg (Hg_T) concentrations in the water and sediment samples used (<0.002–0.03 μg/L and 35.40–391.60 μg/kg dry weight) for studying the biodiversity. It can be concluded that there was no effect of the various Hg levels on the diversity of detected APB species; particularly the PNSB in the shrimp ponds.     

Use of a bioassay to evaluate the toxicity of beauvericin to bacteria

An agar diffusion bioassay was used to compare the sensitivities of bacteria to the mycotoxin beauvericin. Bacillus pumilus LACB101 was inhibited by filter-paper disks containing 0.1 g of beauvericin; B. cereus, B. mycoides, B. sphaericus, Paenibacillus alvei, P. azotofixans, P. macquariensis, and P. pulvifaciens by 1 g; and P. validus by 25 g. The anaerobes Eubacterium biforme, Peptostreptococcus anaerobius, P. productus, Bifidobacterium adolescentis, and Clostridium perfringens were also inhibited by beauvericin.     

Mercuric reductase in environmental Gram-positive bacteria sensitive to mercury

According to existing data, mercury resistance operons (mer operons) are in general thought to be rare in bacteria, other than those from mercury-contaminated sites. We have found that a high proportion of strains in environmental isolates of Gram-positive bacteria express mercuric reductase (MerA protein): the majority of these strains are apparently sensitive to mercury. The expression of MerA was also inducible in all cases. These results imply the presence of phenotypically cryptic mer resistance operons, with both the merA (mercuric reductase) and merR (regulatory) genes still present, but the possible absence of the transport function required to complete the resistance mechanism. This indicates that mer operons or parts thereof are more widely spread in nature than is suggested by the frequency of mercury-resistant bacteria.     

Removal of mercury from mercury-contaminated sediments using a combined method of chemical leaching and volatilization of mercury by bacteria

A method for the removal of mercury sulfide frommercury-contaminated sediments was developed, whichconsists of chemical leaching and volatilization ofmercury by bacteria. More than 85% of the mercury insediment containing 0.11–37.4 mg/kg of mercury wasefficiently extracted with 3 M HCl and 74 mMFeCl₃. Subsequent volatilization by bacteriaresulted in the removal of 62.9–75.1% of mercury frommercury-contaminated Minamata Bay sediments. Methylmercury was also eliminated from soil at a highefficiency. Thus, this combined method of chemicaland microbial treatments could be used for efficientremoval of both organic and inorganic mercurials fromnatural sediments.     

Cobalt toxicity in anaerobic granular sludge: influence of chemical speciation

The influence of cobalt speciation on the toxicity of cobalt to methylotrophic methanogenesis in anaerobic granular sludge was investigated. The cobalt speciation was studied with three different media that contained varying concentrations of complexing ligands [carbonates, phosphates and ethylenediaminetetraacetic acid (EDTA)]. Three fractions (nominal added, dissolved and free) of cobalt were determined in the liquid media and were correlated with data from batch toxicity experiments. The average concentration of cobalt that was required for 50% inhibition of methanogenic activity (IC₅₀) for free Co²⁺ in the three sets of measurements was 13 μmol/L with a standard deviation of 22% and a similarity of 72% between the data obtained in the three different media for the range of cobalt concentrations investigated. The standard deviation of the IC₅₀ for the other two fractions was much higher, i.e. 85 and 144% for the added cobalt and dissolved cobalt, respectively, and the similarity was almost 0% for both fractions. Complexation (and precipitation) with EDTA, phosphates and carbonates was shown to decrease the toxicity of cobalt on methylotrophic methanogenesis. The free cobalt concentration is proposed to be the key parameter to correlate with cobalt toxicity. Thus, the toxicity of cobalt to granular sludge can be estimated based on the equilibrium-free cobalt concentration.     

几种杀螺细菌的筛选与功效研究

湖北钉螺是日本血吸虫的唯一中间宿主,生物杀螺具有环保、成本较低以及对钉螺定向特异的优点。从鄱阳湖草洲土壤及病螺体内分离出4株对钉螺具有高毒性作用的细菌。革兰氏染色和显微镜观察表明,一株为芽孢杆菌,其余3株均为革兰氏阴性杆菌。16srDNA扩增及序列比对鉴定其种属类型,结果显示分别为Enterobactersp．、B．cereus、S．putrefaciens和Aeromonassp．。杀螺研究结果表明,4株细菌的发酵液、发酵上清液和菌体对钉螺均表现出不同程度的毒杀作用,其中S．putrefaciens和Aeromonassp．的杀螺效果最好,72h发酵液和发酵上清液对钉螺的毒杀作用分别达100％和95％以上。各细菌发酵上清SDS-PAGE结果发现了丰富蛋白带,推测这些蛋白中存在对钉螺具有毒杀功效的组分。     

Mutants of luminous bacteria selected for bioluminescent toxicity tests

Mutants of the luminescent bacterial strain NRRL B-11177 were isolated with pleiotropic hypersensitivity towards hydrophobic antimicrobial agents. SDS-PAGE analyses of outer membrane proteins and lipopolysaccharides revealed that the outer membrane structure of the ahs-mutants was altered. QSAR analysis showed that the inhibitory effect of chloro-substituted phenols on bioluminescence of the ahs-mutants depended on their hydrophobicity. The effect of chlorinated phenols and detergents on bioluminescence was increased in the ahs-mutants. The potential use of these mutants in bioluminescent toxicity tests was discussed.     

The role of recombination,niche‐specific gene pools and flexible genomes in the ecological speciation of bacteria

Bacteria diversify into genetic clusters analogous to those observed in sexual eukaryotes, but the definition of bacterial species is an ongoing problem. Recent work has focused on adaptation to distinct ecological niches as the main driver of clustering, but there remains debate about the role of recombination in that process. One view is that homologous recombination occurs too rarely for gene flow to constrain divergent selection. Another view is that homologous recombination is frequent enough in many bacterial populations that barriers to gene flow are needed to permit divergence. Niche‐specific gene pools have been proposed as a general mechanism to limit gene flow. We use theoretical models to evaluate additional hypotheses that evolving genetic architecture, specifically the effect sizes of genes and gene gain and loss, can limit gene flow between diverging populations. Our model predicts that (a) in the presence of gene flow and recombination, ecological divergence is concentrated in few loci of large effect and (b) high rates of gene flow plus recombination promote gene loss and favor the evolution of niche‐specific genes. The results show that changing genetic architecture and gene loss can facilitate ecological divergence, even without niche‐specific gene pools. We discuss these results in the context of recent studies of sympatric divergence in microbes.     

Heterogeneity of the populations of marine luminescent bacteria Photobacterium leiognathi under different conditions of cultivation

Manifestation of pleiotropic effects in the isogenic variants of the luminescent bacterium Photobacterium leiognathi 54 was investigated. The decrease or increase of the expression level of bioluminescence was caused by changes in lux operon regulation. The dynamics of the bioluminescence of dark and dim variants did not differ from the dynamics of the initial luminescent variant, but dependence of the level of luminescence intensity on the exogenous autoinducer of the lux operon was revealed. The investigated variants of P. leiognathi 54 inherited fairly stable morphological characteristics, colony architectonics, level of luminescence, and activity of some enzymes; variants with reduced bioluminescence formed colonies of the S type. Stable bright variants with S-and R-type colonies appeared both in the initial strain population and in the dark variant population, but with smaller frequency. Populations of the bright variant with R-type colonies were most heterogeneous; this can be determined by the lack of glucose repression of the bioluminescence in contrast to other investigated inherited variants of P. leiognathi.     

Evaluation and application of aquatic toxicity tests: use of the Microtox test for the prediction of toxicity based upon concentrations of contaminants in soil

The quality control for the reuse of cleaned soil from a contaminated site consisted in the determination of the main contaminants by analytical chemical methods such as GC and HPLC. Since it is not possible to analyze for all contaminants a toxicity test should be used to detect large concentrations of not routinely analyzed chemicals. The aim of the study was to develop a system for toxicity testing, which should be able to predict the toxicity of soil samples based on the concentration of chemicals in the soil and to detect toxic chemicals not analyzed by the routinely conducted soil analysis.Based upon the relative sensitivity to various contaminants as well as practical aspects such as test duration and costs the Microtox® test was favoured over the bioassays with Daphnia magna and Scenedesmus subspicatus. The Microtox® test was used to measure the toxicity of various pesticides and their major metabolites. The toxicity data of the pure compounds were used to predict the toxicity (EC₅₀ and % inhibition of the bioluminescence reaction) of defined mixtures of chemicals in water by applying two different mathematical appriaches which are based on the additivity of the effects of the single chemicals. The predicted values were compared with the experimental data and showed good agreement.In order to be able to predict the toxicity of soil samples using the Microtox® test the soil/water partition coefficient (K _d) was measured for the main contaminants. The toxicity of soil samples was predicted by calculating the concentration of the contaminants in the leachate by using the corresponding concentration in the soil and applying the K _d values determined. From the calculated composition of the leachate the expected toxicity was estimated. This value was compared with the toxicity experimentally determined in the Microtox® test.     

贵州万山汞矿区污染土壤中汞的形态分布特征

利用优化Tessier连续化学浸提法对贵州万山汞矿区污染土壤中汞的形态分布进行研究.结果表明,稻田土壤中汞主要以残渣态形式存在(79.65%),其次为有机结合态(19.97%)、氧化态(0.31%)、特殊吸附态(0.04%)和溶解态与可交换态(0.03%);除特殊吸附态外,其他各形态汞含量均随距污染源距离增加而降低,特殊吸附态汞在分析土壤中含量变化不明显;生物可利用性(溶解态与可交换态和特殊吸附态)汞占总汞比例较低,但在污染土壤中其含量明显高于未受污染地区.     

Detoxification of mercury and organomercurials by nitrogen-fixing soil bacteria

Minimal inhibitory concentration values of HgCl₂ and 5 organomercurials were determined against 24 mercury-resistant N₂-fixing soil bacteria previously isolated from soil and identified in our laboratory. These bacterial strains also displayed multiple antibiotic resistant properties. Typical growth pattern of a highly mercury-resistantBeijerinckia sp (KDr₂) was studied in liquid broth supplemented with toxic levels of mercury compounds. Four bacterial strains were selected for determining their ability to volatilize mercury and their Hg-volatilizing capacity was different. Cell-free extracts prepared from overnight mercury-induced cells catalyzed Hg²⁺-induced NADPH oxidation. Specific activities of Hg²⁺-reductase which is capable of catalyzing conversion of Hg²⁺ →Hg(o) of 10 Hg-resistant bacterial strains are also reported.     

Review of characteristics of mercury speciation and mobility from areas of mercury mining in semi-arid environments

The speciation of mercury—including most phase minerals, secondary phases, gaseous and aqueous species—is very important for evaluating the environmental impact and mobilization of this contaminant. Mining activities produce mercury mine waste, which includes several types of material (mainly mine waste and calcines) with varying mercury content and speciation depending on the ore deposit and processing technology. The main phase minerals are cinnabar, metacinnabar, metallic Hg⁰, corderoite, livingstonite, calomel and schuetteite. The aqueous mobilization of mercury is controlled by complex formation, pH-Eh conditions, the primary phase mineral of mercury, and organic-matter and iron oxyhydroxide content. The possibility of colloidal transport of mercury from mine waste is influenced by the atmospheric emission of metallic Hg⁰ and the leaching of waste by episodic high-intensity precipitations. In these climatic conditions, mercury can be mobilized to pore water, surface water or groundwater by the dissolution of metallic Hg⁰ and cinnabar in acidic conditions, and by the colloidal transport. The presence of Hg-soluble phases (chlorides and oxychlorides) may enhance the mobilization of mercury. In the semi-arid conditions of the Valle del Azogue (SE Spain) the atmospheric emissions of the Hg⁰ present in calcines and mine waste may be significant and the dissolution of Hg⁰ and metal-sulfate salts during episodic runoff events may explain the mobilization of Hg, Fe, Pb, Zn and other heavy metals.     

发光细菌在水环境生物毒性检测中应用的研究进展

发光细菌是一类自身含发光基因且能够发出可见光的细菌,其分布非常广泛。由于具备了日常毒性检测所要求的快速灵敏以及再现性好的特点,发光细菌法愈来愈受到关注,且因其快速和低成本的特点常被用作早期预警系统。基因操作技术的介入更使得发光细菌试验具有了分辨各种毒性的功能。本文综述了发光细菌在环境毒性检测中的进展,重点介绍了基于发光细菌的毒性试验方法,以及在水质检测中的应用。     

Experimental study of the combined effects of pH and salinity on the bioaccumulation of inorganic mercury in the crayfish Astacus leptodactylus

ABSTRACT

The combined effects of pH and salinity were investigated on the bioaccumulation of inorganic mercury in the Turkish crayfish Astacus leptodactylus, at the whole organism and organ levels. Effects on the ionic balance were also analysed during the acclimatization phase and after 3 and 15 days of exposure, via the direct route. The experimental protocol, based on a complete factorial design, took into account nine experimental conditions, resulting from the combination of three levels for pH (6, 7.5 and 9) and for salinity (1, 10 and 100 mM Cl-). The results showed an important Hg accumulation in the organs or tissues in direct contact with the surrounding medium, e.g. gills and carapace. The effects of the nine pH and salinity conditions were complex and variable from one organ to another. Correlations were studied between chemical species concentrations in the water (HgCl₂, HgOHCl, HgCl₃-,HgCl₄², Hg(OH)₂) and amounts of metal accumulated in the different organs. A positive correlation was found between the neutral species HgCl₂ and metal accumulated in the gills and in the carapace. Perturbations of the haemolymph ionic concentrations were no significant, except for Na⁺ after 3 days of exposure.     

Comparative study of the assay of Artemia salina L. and the estimate of the medium lethal dose (LD50 value) in mice, to determine oral acute toxicity of plant extracts

Artemia salina L. (Artemiidae), the brine shrimp larva, is an invertebrate used in the alternative test to determine toxicity of chemical and natural products. In this study the Medium Lethal Concentrations (LC50 value) of 20 plant extracts, Aloe vera (L.) Burm. F. (Aloeaceae), Artemisia absinthium L. (Asteraceae); Citrus aurantium L. (Rutaceae); Cymbopogon citratus (DC. Ex Nees) Stapf (Poaceae); Datura stramonium L. (Solanaceae); Justicia pectoralis Jacq. (Acanthaceae); Musa x paradisiaca L. (Musaceae); Ocimum basilicum L.; O. gratissimum L.; O. tenuiflorum L. (Lamiaceae); Pimenta dioica (L.) Merr. (Myrtaceae); Piper auritum Kunth (Piperaceae); Plantago major L. (Plantaginaceae); Plectranthus amboinicus (Lour.) Spreng. (Lamiaceae); Ruta graveolens L. (Rutaceae); Senna alata (L.) Roxb. (Fabaceae); Stachytarpheta jamaicensis (L.) Vahl (Verbenaceae); and Thuja occidentalis L. (Cupressaceae), were determined using Artemia salina L. (Artemiidae), with the objective of relating the results to the LD50 values reported in mice (tested at three concentrations: 10, 100, and 1000 microg/mL, for each extract). We found good correlation between the in vivo and the in vitro tests (r = 0.85 p < 0.05), and this method is a useful tool for predicting oral acute toxicity in plant extracts.     

Studies on the mercury volatilizing enzymes in nitrogen-fixing Beijerinckia mobilis

Beijerinckia mobilis KDr₂, a broad-spectrum, mercury-resistant nitrogen-fixing organism, possesses multiple metal-resistance properties. Mercuric reductase and organomercurial lyase activities of this bacterial strain were determined using different mercury compounds and heavy metal salts of copper, nickel, cobalt, cadmium, silver, zinc, lead and arsenate as inducers and substrates. Mercuric reductase was partially purified and the effect of some enzyme inhibitors and heavy metal ions on the enzyme activity was studied. The enzyme activity was completely inhibited by CdCl₂, Bi(NO₃)₃ and KCN at 10^-5 M and by AgNO₃, CoCl₂ and CuSO₄ at 10^-4 M.     

The effect of the pH level on the communities of anoxygenic phototrophic bacteria of soda lakes of the southeastern Transbaikal Region

The effect of pH on the structure of the communities of anoxygenic phototrophic bacteria (APB) was studied under laboratory conditions. Samples of natural APB communities were inoculated into media that differed in pH values, which were 7, 9.5, or 10.5. The structure of the APB communities in the obtained enrichment cultures at all pH values depended also on the mineralization levels of the media, which were the same as in the lakes from which samples were taken. The same dependence of the community structure on salinity was observed as in the case of the natural communities that had been described previously. APB were most diverse in the enrichment cultures grown at pH 9.5. The shift of the pH to either neutral or extremely alkaline values restricted the species diversity within the APB community, resulting in marked predominance of the most adapted forms. It was shown that the status of Ectothiorhodospira species within the community could serve not only as an indicator of salinity but also as an indicator of pH in soda lakes with a water mineralization of higher than 5 g/l. The statuses of various APB groups in the community as dependent on pH and salinity are discussed, as well as possible changes in these statuses due to changes in the water level and other environmental parameters in the studied lakes.