A comparative analysis of endophytic bacterial communities associated with hyperaccumulators growing in mine soils

Interactions between endophytic bacterial communities and hyperaccumulators in heavy metal-polluted sites are not fully understood. In this study, the diversity of stem-associated endophytic bacterial communities of two hyperaccumulators (Solanum nigrum L. and Phytolacca acinosa Roxb.) growing in mine soils was investigated using molecular-based methods. The denaturing gradient gel electrophoresis (DGGE) analysis showed that the endophytic bacterial community structures were affected by both the level of heavy metal pollution and the plant species. Heavy metal in contaminated soil determined, to a large extent, the composition of the different endophytic bacterial communities in S. nigrum growing across soil series (five sampling spots, and the concentration of Cd is from 0.2 to 35.5 mg/kg). Detailed analysis of endophytic bacterial populations by cloning of 16S rRNA genes amplified from the stems of the two plants at the same site revealed a different composition. A total of 51 taxa at the genus level that included α-, β-, and γ-Proteobacteria (68.8% of the two libraries clones), Bacteroidetes (9.0% of the two libraries clones), Firmicutes (2.0% of the two libraries clones), Actinobacteria (16.4% of the two libraries clones), and unclassified bacteria (3.8% of the two libraries clones) were found in the two clone libraries. The most abundant genus in S. nigrum was Sphingomonas (23.35%), while Pseudomonas prevailed in P. acinosa (21.40%). These results suggest that both heavy metal pollution and plant species contribute to the shaping of the dynamic endophytic bacterial communities associated with stems of hyperaccumulators.     

Mercury methylation by a microbial community from sediments of the Adour Estuary (Bay of Biscay, France)

In order to study the influence of microorganisms on the mercury biogeochemistry, the metal content and the structure of microbial communities were determined in sediments from stations along the Adour Estuary. The comparison of the bacterial communities and their distribution in function of the environmental parameters by Canonical Correspondence Analysis (CCA) revealed the influence of metals on the bacterial communities structure. Sediments where the bacterial communities are mostly influenced by methylmercury were incubated in slurries with or without mercury, under oxic and anoxic conditions. Methylmercury production was detected in the anoxic biotic slurries with a net methylation yield of 0.3% after 24 h. CCA based on T-RFLP profiles revealed the impact of mercury addition on the bacterial communities structure. In addition, 17 bacterial strains, mainly sulphate-reducing bacteria involved in mercury methylation, were isolated and identified.     

Effect of organic carbon and metal accumulation on the bacterial communities in sulphidogenic sediments

A unique geochemical setting in Lake Cadagno, Switzerland, has led to the accumulation of insoluble metal sulphides in the sedimentary record as the result of past airborne pollution. This offers an exceptional opportunity to study the effect of these metals on the bacterial communities in sediments, and in particular to investigate further the link between metal contamination and an increase in the populations of endospore-forming bacteria observed previously in other metal-contaminated sediments. A decrease in organic carbon and total bacterial counts was correlated with an increase in the numbers of endospores in the oldest sediment samples, showing the first environmental evidence of a decrease in nutrient availability as a trigger of sporulation. Proteobacteria and Firmicutes were the two dominant bacterial phyla throughout the sediment, the former in an area with high sulphidogenic activity, and the latter in the oldest samples. Even though the dominant Firmicutes taxa were stable along the sediment core and did not vary with changes in metal contamination, the prevalence of some molecular species like Clostridium sp. was positively correlated with metal sulphide concentration. However, this cannot be generalized to all endospore-forming species. Overall, the community composition supports the hypothesis of sporulation as the main mechanism explaining the dominance of endospore formers in the deepest part of the sediment core, while metal contamination in the form of insoluble metal sulphide deposits appears not to be linked with sporulation as a mechanism of metal tolerance in this sulphidogenic ecosystem.     

Periphyton as an indicator of water quality in the St Lawrence River (Québec, Canada)

The performance of various algal indices to document improvements in water quality across a low nutrient concentration gradient was assessed during 2 years in the St Lawrence River (Quebec, Canada). Water-quality variables and periphyton samples were collected on navigational buoys near Montreal during the spring, summer and fall of 1994 and 1995. Exposure to urban wastewater varied widely within the sector surrounding the island of Montreal, with some areas upstream receiving no direct effluents and areas further downstream receiving treated and untreated wastewater. Faecal coliform concentrations provided a good tracer of effluents and were significantly correlated to nutrient concentrations (r = 0.33-0.72, p < 0.001) and water transparency (r = 0.70, p < 0.001). Despite a strong gradient in faecal coliform concentration (< 2 to > 20 000 UFC/100 ml), algal biomass and diversity did not reflect differences between sites with varying levels of urban wastewater. Taxonomic composition of periphyton communities, particularly the presence of the cyanophyte Plectonema notatum Schmidle, was related (r = 0.48, p = 0.004) to exposure to urban effluents. Variables describing seasonal changes (temperature, Julian day, river discharge, conductivity, NO2-NO3) explained a large fraction of total variance (38-52% of total variance) and thus exerted the predominant influence on algal biomass and species composition in the St Lawrence River. Variables describing the presence of effluents explained 1-22% of the variance in compositional data. Subtle changes in periphyton species composition were the only response to different levels of exposure to urban wastewater in the Montreal area, which represented relatively small differences in comparison to natural seasonal variability.     

The ecology of acidification and recovery: changes in herbivore-algal food web linkages across a stream pH gradient

We examined the effects of acidification on herbivore-algal food web linkages in headwater streams. We determined the structure and abundance of consumer and benthic algal assemblages, and gauged herbivory, in 10 streams along a pH gradient (mean annual pH 4.6-6.4). Biofilm taxonomic composition changed with pH but total abundance did not vary systematically across the gradient. Mayflies and chironomids dominated under circumneutral conditions but declined with increasing acidity and their consumption of algae was strongly reduced. Contrary to expectations, several putative shredder species consumed algae, maintaining the herbivore-algal linkage where specialist grazers could not persist. These shifts in functioning could render the communities of acidified streams resistant to reinvasion when acidity ameliorates and water chemistry is restored to a pre-acidification condition. This hypothesis is discussed in the light of recent trends in the chemistry and biology of the UK Acid Waters Monitoring Network sites.     

The relationship between trace metal contamination and stream meiofauna

The effect of chemical contaminants on freshwater meiofaunal communities is poorly understood and meiofauna rarely form part of environmental impact assessments in fresh waters. The community composition and diversity of meiofauna in streams of southwest England (Cornwall) representing a gradient in trace metal contamination were investigated. Multivariate and univariate statistical techniques were used to correlate community composition, diversity and environmental variables. Meiofaunal communities were very different at sites with high compared with low metal concentrations. Copper, either singly, or in combination with other environmental variables (aluminium, zinc or dissolved organic carbon), was the most important correlate with community composition. Not all meiofaunal species were adversely affected by metals, however, and some taxa, most notably certain cyclopoid copepods, were abundant at high metal concentrations. Moreover, sites with high metal concentrations were not significantly less diverse than sites of low metal concentrations. These data suggest that metal contamination significantly alters the composition of stream meiofaunal assemblages and highlight the importance of including meiofauna in impact studies of fresh waters.     

Using benthic recruitment to assess the significance of contaminated sediments: the influence of taxonomic resolution

The use of small-scale experimental units as a means of evaluating the ecological effects of contaminated sediments was examined at the species, family, mixed and phylum levels of taxonomic resolution. Sediments were taken from various locations representing a range of contaminant loads. Containers with these sediments were placed in situ at a relatively uncontaminated location for 90 days. The containers were retrieved and the abundance of the macrofauna which recruited to the containers was estimated. The results showed that the composition of the benthic communities in the more highly contaminated sediments differed significantly from those in less contaminated sediments. Analyses at the different taxonomic levels showed that all but the phylum level data showed some differences in community structure among sediment types. The study showed that small-scale experiments are useful for examining the effects of contaminants and that higher levels of taxonomic resolution can be used to describe variations in the structure of benthic communities at this spatial scale.     

Macrobenthic community in the Douro estuary: relations with trace metals and natural sediment characteristics

The relationship between macrobenthic community structure and natural characteristics of sediment and trace metal contamination were studied in the lower Douro estuary (Portugal, NW, Iberian Peninsula), using an innovative threefold approach (SQG, Sediment Quality Guidelines), metal normalization to Fe, and macrobenthic community structure. This study allowed detection of a clear signature of anthropogenic contamination, in terms of Zn, Cu, Pb, and Cr in the north bank of the estuary, which experiences high urban pressure. Using the SQG approach, metal concentrations above ERM (effects range-median) were observed only at one sampling station, but several stations had levels above ERL (effects range-low). The macrobenthic community had a low diversity, with only 19 species found in the entire estuarine area, dominated by opportunistic species. The granulometric distribution of the sediments (estimated from the combination of organic matter, Fe and Al) seemed to be the major structuring factor for the communities, establishing the natural macrobenthic distribution pattern. The metals (Zn, Cu, Pb, and Cr) seemed to act as a disturbing factor over the natural distribution, with deleterious consequences for the macrobenthic communities.     

Oxygen levels versus chemical pollutants: do they have similar influence on macrofaunal assemblages? A case study in a harbour with two opposing entrances

Generally, harbours are polluted zones characterised by low values of hydrodynamism and oxygen in the water column and high concentrations of pollutants in sediments. The harbour of Ceuta, North Africa, has an unusual structure; it is located between two bays connected by a channel, which increases the water movement and exchange in the harbour, maintaining moderate oxygen levels in the water-sediment interface. Nevertheless, high concentration of organic matter, nutrients and heavy metals were measured in sediments from this harbour. Under these unusual conditions (high levels of pollution but total saturation of oxygen in the water column) we studied the responses of soft-bottom macrobenthic communities using uni and multivariate analyses. The number of species was similar inside and outside the harbour but the species composition differed between internal and external stations; oxygen levels seem to control the "quantity" of species whereas pollutants control the "quality" of them.     

Functional recovery of biofilm bacterial communities after copper exposure

Potential of bacterial communities in biofilms to recover after copper exposure was investigated. Biofilms grown outdoor in shallow water on glass dishes were exposed in the laboratory to 0.6, 2.1, 6.8 micromol/l copper amended surface water and a reference and subsequently to un-amended surface water. Transitions of bacterial communities were characterised with denaturing gradient gel electrophoresis (DGGE) and community-level physiological profiles (CLPP). Exposure to 6.8 micromol/l copper provoked distinct changes in DGGE profiles of bacterial consortia, which did not reverse upon copper depuration. Exposure to 2.1 and 6.8 micromol/l copper was found to induce marked changes in CLPP of bacterial communities that proved to be reversible during copper depuration. Furthermore, copper exposure induced the development of copper-tolerance, which was partially lost during depuration. It is concluded that bacterial communities exposed to copper contaminated water for a period of 26 days are capable to restore their metabolic attributes after introduction of unpolluted water in aquaria for 28 days.     

Organic carbon effects on aerobic polychlorinated biphenyl removal and bacterial community composition in soils and sediments

Certain organic compounds, including biphenyl and salicylic acid, stimulate polychlorinated biphenyl (PCB) degradation by microorganisms in some environments. However, the usefulness of these amendments for improving PCB removal by microorganisms from diverse habitats has not been extensively explored. This study evaluated the effects of biphenyl, salicylic acid, and glucose on changes in aerobic PCB removal and bacterial communities from an agricultural soil, a wetland peat soil, a river sediment, and a mixture of these samples. PCB removal patterns were significantly different between soils and sediments amended with carbon compounds: (i) terrestrial soil microorganisms removed more PCBs than river sediment microorganisms, particularly with regard to PCBs with >4 chlorine substituents, (ii) glucose-supplemented, agricultural soil microorganisms removed more hexachlorobiphenyl than unsupplemented samples, (iii) biphenyl-supplemented, river sediment microorganisms removed more di- and tri-chlorobiphenyls than unamended samples. Carbon amendments also caused unique shifts in soil and sediment bacterial communities, as determined by specific changes in bacterial 16S rRNA denaturing gradient gel electrophoresis banding patterns. These results indicate that organic carbon amendments had site-specific effects on bacterial populations and PCB removal. Further work is needed to more accurately characterize PCB degrading communities and functional gene expression in diverse types of environments to better understand how they respond to bioremediation treatments.     

Bacterial community structure in soils contaminated by polycyclic aromatic hydrocarbons

Bacterial community structure was examined in polycyclic aromatic hydrocarbon (PAH) contaminated soil taken from a timber treatment facility in southern Ireland. Profiles of soil bacterial communities were generated using a molecular fingerprinting technique, terminal restriction fragment length polymorphism (TRFLP), and results were interpreted using sophisticated multivariate statistical analysis. Findings suggested that there was a correlation between PAH structure and bacterial community composition. Initial characterisation of soil from the timber treatment facility indicated that PAH contamination was unevenly distributed across the site. Bacterial community composition was correlated with the type of PAH present, with microbial community structure associated with soil contaminated with two-ringed PAHs only being distinctly different to communities in soils contaminated with multi-component PAH mixtures. Typically the number of bacterial ribotypes detected in samples did not appear to be adversely affected by the level of contamination.     

Evidence of population genetic effects in Peromyscus melanophrys chronically exposed to mine tailings in Morelos, Mexico

Effects of environmental chemical pollution can be observed at all levels of biological organization. At the population level, genetic structure and diversity may be affected by exposure to metal contamination. This study was conducted in Huautla, Morelos, Mexico in a mining district where the main contaminants are lead and arsenic. Peromyscus melanophrys is a small mammal species that inhabits Huautla mine tailings and has been considered as a sentinel species. Metal bioaccumulation levels were examined by inductively coupled plasma mass spectrometry and genetic analyses were performed using eight microsatellite loci in 100 P. melanophrys individuals from 3 mine tailings and 2 control sites. The effect of metal bioaccumulation levels on genetic parameters (population and individual genetic diversity, genetic structure) was analyzed. We found a tissue concentration gradient for each metal and for the bioaccumulation index. The highest values of genetic differentiation (Fst and Rst) and the lowest number of migrants per generation (Nm) were registered among the exposed populations. Genetic distance analyses showed that the most polluted population was the most genetically distant among the five populations examined. Moreover, a negative and significant relationship was detected between genetic diversity (expected heterozygosity and internal relatedness) and each metal concentration and for the bioaccumulation index in P. melanophrys. This study highlights that metal stress is a major factor affecting the distribution and genetic diversity levels of P. melanophrys populations living inside mine tailings. We suggest the use of genetic population changes at micro-geographical scales as a population level biomarker.     

Lead pollution in a large, prairie-pothole lake (Rush Lake, WI, USA): effects on abundance and community structure of indigenous sediment bacteria

Rush Lake (WI, USA), the largest prairie-pothole lake east of the Mississippi River, has been contaminated with lead pollution as a result of over 140 years of waterfowl hunting. We examined: (1) the extent of lead pollution in Rush Lake sediments and (2) whether lead pollution in Rush Lake is affecting the abundance and community structure of indigenous sediment bacteria. Sediment lead concentrations did not exceed 59 mg Pb kg(-1) dry sediment. No relationship was observed between sediment lead concentration and the abundance of aerobic (P=0.498) or anaerobic (P=0.416) heterotrophic bacteria. Similarly, lead did not appear to affect bacterial community structure when considering both culturable and nonculturable community members. In contrast, the culturable fraction of sediment bacteria in samples containing 59 mg Pb kg(-1) exhibited a unique community structure. While factors other than lead content likely play roles in determining bacterial community structure in the sediments of Rush Lake, these data suggest that the culturable fraction of sediment bacterial communities is affected by elevated lead levels.     

The roles of biological interactions and pollutant contamination in shaping microbial benthic community structure

Biological interactions between metazoans and the microbial community play a major role in structuring food webs in aquatic sediments. Pollutants can also strongly affect the structure of meiofauna and microbial communities. This study aims investigating, in a non-contaminated sediment, the impact of meiofauna on bacteria facing contamination by a mixture of three PAHs (fluoranthene, phenanthrene and pyrene). Sediment microcosms were incubated in the presence or absence of meiofauna during 30 days. Bioremediation treatments, nutrient amendment and addition of a hydrocarbon-degrading bacterium, were also tested to enhance PAH biodegradation. Results clearly show the important role of meiofauna as structuring factor for bacterial communities with significant changes observed in the molecular fingerprints. However, these structural changes were not concomitant with changes in biomass or function. PAH contamination had a severe impact on total meiofaunal abundance with a strong decrease of nematodes and the complete disappearance of polychaetes and copepods. In contrast, correspondence analysis, based on T-RFLP fingerprints, showed that contamination by PAH resulted in small shifts in microbial composition, with or without meiofauna, suggesting a relative tolerance of bacteria to the PAH cocktail. The PAH bioremediation treatments were highly efficient with more than 95% biodegradation. No significant difference was observed in presence or absence of meiofauna. Nutrient addition strongly enhanced bacterial and meiofaunal abundances as compared to control and contaminated microcosms, as well as inducing important changes in the bacterial community structure. Nutrients thus were the main structural factor in shaping bacterial community composition, while the role of meiofauna was less evident.     

Impact of metal stress on the production of secondary metabolites in <Emphasis Type="Italic">Pteris vittata</Emphasis> L. and associated rhizosphere bacterial communities

Plants adapt to metal stress by modifying their metabolism including the production of secondary metabolites in plant tissues. Such changes may impact the diversity and functions of plant associated microbial communities. Our study aimed to evaluate the influence of metals on the secondary metabolism of plants and the indirect impact on rhizosphere bacterial communities. We then compared the secondary metabolites of the hyperaccumulator Pteris vittata L. collected from a contaminated mining site to a non-contaminated site in Vietnam and identified the discriminant metabolites. Our data showed a significant increase in chlorogenic acid derivatives and A-type procyanidin in plant roots at the contaminated site. We hypothesized that the intensive production of these compounds could be part of the antioxidant defense mechanism in response to metals. In parallel, the structure and diversity of bulk soil and rhizosphere communities was studied using high-throughput sequencing. The results showed strong differences in bacterial composition, characterized by the dominance of Proteobacteria and Nitrospira in the contaminated bulk soil, and the enrichment of some potential human pathogens, i.e., Acinetobacter, Mycobacterium, and Cupriavidus in P. vittata’s rhizosphere at the mining site. Overall, metal pollution modified the production of P. vittata secondary metabolites and altered the diversity and structure of bacterial communities. Further investigations are needed to understand whether the plant recruits specific bacteria to adapt to metal stress.     

Bacterial community structure analyses to assess pollution of water and sediments in the Lake Shkodra/Skadar, Balkan Peninsula

- Goal, Scope, Background. Lake Skadar is the largest lake in Balkan Peninsula, located on the Montenegro-Albanian border. The unique features of the lake and wide range of endemic and rare or endangered plant and animal species resulted in the classification of the Skadar as a wetland site of international significance. In spite of its importance the Lake is influenced by inflowing waters from river Morača and other regional rivers contaminated by the industry, municipal and agricultural activities in the area. Therefore, the Lake has been subject of various physical, chemical, biological and toxicological examinations. However, community-level analyses are most relevant to assess the effect of stressors on aquatic ecosystems. In the present study bacterial community structure among differently polluted sites of the lake was compared by genetic fingerprinting technique. Methods Water and sediment samples were collected from five differently polluted sampling sites on the Lake Skadar in spring and autumn of the same year. The bacterial community structure in the samples was characterized and compared by temporal temperature gel electrophoresis (TTGE) analysis of polymerase chain reaction-amplified bacterial 16S rRNA genes. Results and Discussion The TTGE analysis resulted in many distinguishable and reproducible band patterns, allowing reliable comparison of bacterial communities among sampling sites. Results on the bacterial community structure revealed that three of the selected locations can be considered as sites that have not shown any pollution degradation determined by our method, due to similar structure of bacterial community in the sediment samples. On the other hand, significant shifts in bacterial community structure in the mouth of the river Morača and Plavnica were shown. Since the results coincide with some of the bioassays and chemical analysis performed previously, the changes in bacterial community structure are explained as an effect of antropogenic pollution on the lake ecosystem by waters of river Morača and stream Plavnica. Conclusion The TTGE has proven to be an efficient and reliable method to monitor bacterial dynamics and community shifts in aquatic environment, especially in the sediments. Within the variety of environmental quality assessments the use of TTGE analyses of bacterial community is strongly recommended, particularly as an initial investigation. However, in any conclusion on the state of the environment, the TTGE results should be combined to some other biological, chemical and hydrological data. Recommendation and Outlook Since prokaryotes are a crucial group of organisms in the biosphere, the ecosystem function studies are largely based on bacterial communities. Therefore, bacterial community structure analysis should be a part of an integrated weight of evidence approach in pollution assessment. In case of Triad approach, consisting of chemical analyses, bioassays, and community studies in the field, the TTGE bacterial community structure analyses should be placed in the later Triad leg. In comparison to other community studies, based on various biotic indices, the TTGE bacterial community analysis has proven to be very sensitive, reliable and less time consuming.     

Oribatid mite communities and metal bioaccumulation in oribatid species (Acari, Oribatida) along the heavy metal gradient in forest ecosystems

The responses of oribatid communities to heavy metal contamination were studied. Concentration of cadmium, copper and zinc in nine oribatid species along a gradient of heavy metal pollution was measured. Oribatid mites were sampled seasonally during two years in five forests located at different distances from the zinc smelter in the Olkusz District, southern Poland. The most numerous and diverse oribatid communities were found in the forest with moderate concentrations of heavy metals. Analysis by atomic absorption spectrophotometry revealed large differences in metal body burdens among species. All studied oribatid species appeared to be accumulators of copper with Oppiella nova, Nothrus silvestris and Adoristes ovatus characterized by the highest bioaccumulation factors. Most species poorly accumulate cadmium and zinc. The accumulation of heavy metals in the body of oribatids was not strictly determined by their body size or the trophic level at which they operate.     

Application of 16S rDNA-PCR amplification and DGGE fingerprinting for detection of shift in microbial community diversity in Cu-, Zn-, and Cd-contaminated paddy soils

Seven soils were sampled from farmland at different distances (0.01-5 km) from a copper and zinc smelter. The total contents of heavy metals in these soils ranged from 46 to 4895 mg Cu kg-1, 96 to 1133 mg Zn kg-1, and 6.9 to 28.8 mg Cd kg-1, respectively. The available fractions were highly correlated with total contents of the metals. In order to assess the impact of combined contamination of heavy metals on soil bacterial communities, denaturing gradient gel electrophoresis (DGGE) of polymerase chain reaction (PCR) amplicons of 16S rDNA sequence of bacteria in soil was used. Bacterial community structure was affected to some extent by heavy metals. The number of DGGE bands in soils increased with increasing distance from the copper and zinc smelter. Clustering analysis of the DGGE profiles showed that bacteria in the seven soils belonged to three clusters. Bacterial communities in three soils sampled at 0.01-0.60 km from the smelter belonged to one cluster, and those in three soils sampled at 0.8-1.2 km from the smelter belong to another cluster. Bacterial community in soil farthest from the smelter belonged to a single cluster. This study demonstrated that heavy metal contamination decreased both biomass and diversity of bacterial community in the soil.