Anaerobic biotransformations of pollutant chemicals in aquifers

Summary Anaerobic microbial communities sampled from either a methanogenic or sulfate-reducing aquifer site have been tested for their ability to degrade a variety of groundwater pollutants, including halogenated aromatic compounds, simple alkyl phenols and tetrachloroethylene. The haloaromatic chemicals were biodegraded in methanogenic incubations but not under sulfate-reducing conditions. The primary degradative event was typically the reductive removal of the aryl halides. Complete dehalogenation of the aromatic moiety was required before substrate mineralization was observed. The lack of dehalogenation activity in sulfatereducing incubations was due, at least in part, to the high levels of sulfate rather than a lack of metabolic potential. In contrast, the degradation of cresol isomers occurred in both types of incubations but proved faster under sulfate-reducing conditions. The requisite microorganisms were enriched and the degradation pathway forp-cresol under the latter conditions involved the anaerobic oxidation of the aryl methyl group. Tetrachloroethylene was also degraded by reductive dehalogenation but under both incubation conditions. The initial conversion of this substrate to trichloroethylene was generally faster under methanogenic conditions. However, the transformation pathway slowed when dichloroethylene was produced and only trace concentrations of vinyl chloride were detected. These results illustrate that pollutant compounds can be biodegraded under anoxic conditions and a knowledge of the predominant ecological conditions is essential for accurate predictions of the transport and fate of such materials in aquifers.     

Distribution of plasmids in groundwater bacteria

Summary Bacteria isolated from groundwater aquifer core materials of pristine aquifers at Lula and Pickett, OK, and from a site with a history of aromatic hydrocarbon contamination and natural renovation located at Conroe, TX, were screened for the presence of plasmid DNA by alkaline or enzyme lysis and agarose gel techniques. Some of the isolates were also subjected to taxonomic tests in addition to screening for resistance to antibiotics, tolerance to heavy metal salts, and bacteriocin production. There was no significant difference in the distribution of the traits usually associated with plasmid occurrence in isolates from the three sites. These traits, which occurred at low frequencies, were not restricted to plasmid-bearing strains of the communities. Plasmids were found in isolates from all three sites, but on the average there was a significantly higher percentage of isolates containing plasmids in the samples from Conroe (19.4%) than from either Lula (1.8%) or Pickett (7.7%). The sizes of the plasmids found ranged between 3.5 and 202 kilobases but, for the Conroe samples, many more isolates (67%) contained smaller plasmids (<10 kb) rather than larger ones. No plasmids were found in bacteria recovered from naturally renovated aquifer material at the Conroe site.     

Isolation and functional analysis of denitrifiers in an aquifer with high potential for denitrification

Aquifers are among the main freshwater sources. The Raigón aquifer is susceptible to contamination, mainly by nitrate and pesticides, such as atrazine, due to increasing agricultural activities in the area. The capacity of indigenous bacteria to attenuate nitrate contamination in different wells of this aquifer was assessed by measuring denitrification rates with either acetate plus succinate or nitrate amendments. Denitrification activity in nitrate-amended assays was significantly higher than in unamended assays, particularly in groundwater from wells where nitrate concentration was 33.5 mg L⁻¹ or lower. Furthermore, groundwater denitrifiers capable of using acetate or succinate as electron donors were isolated, identified by 16S rRNA gene sequencing and evaluated for functional denitrification genes (nirS, nirK and nosZ). Phylogenetic affiliation of 54 isolates showed that all members belonged to nine different genera within the Proteobacteria (Bosea, Ochrobactrum, Azospira, Zoogloea, Acidovorax, Achromobacter, Vogesella, Stenotrophomonas and Pseudomonas). In addition, isolate AR28 that clustered separately from validly described species could potentially belong to a new genus. The majority of the isolates were related to species belonging to previously reported denitrifying genera. However, the phylogeny of the nirS and nosZ genes revealed new sequences of these functional genes. To our knowledge, this is the first isolation and sequencing of the nirS gene from the genus Vogesella, as well as the nosZ gene from the genera Acidovorax and Zoogloea. The results indicated that indigenous bacteria in the Raigón aquifer had the capacity to overcome high nitrate contamination and exhibited functional gene diversity.     

Geomicrobiology and Redox Geochemistry of the Karstified Miocene Gypsum Aquifer,Western Ukraine: The Study from Zoloushka Cave

The gypsum karst of the western Ukraine developed largely under artesian conditions. The Miocene aquifer is presently entrenched and dewatered over much of the territory, while it remains confined in the zone adjacent to the Carpathian Foredeep. The most prominent geochemical features of the Miocene aquifer system in the confined karst zone are: (1) the almost universal presence of a bioepigenetic calcite bed, enriched in the light carbon isotope, at the top of the gypsum (the "Ratynsky Limestone"), (2) the widespread sulfur mineralization associated with the above calcite bed (the region is one of the world's largest sulfur-bearing basins), and (3) high H 2 S and CO 2 in the groundwater. Intense microbial sulfate-reduction processes occur in the gypsum in this zone. Zoloushka Cave is the third longest (92 km) and the largest by volume (more than 7 2 10 5 m 3 ) gypsum cave in the world. It is a unique example of a young artesian cave that only during the Holocene became partly drained and during the last 50 years progressively dewatered due to a quarry operation. These rapid changes have induced a number of transitional geochemical processes, some of which appear to be bacterially mediated. Six groups of microorganisms have been identified in the cave. Our article discusses the aquifer geochemistry during the transitional stage in the light of the microbiological studies.     

In situ analysis of native microbial communities in complex samples with high particulate loads

In the present study a procedure combining a cell extraction method and Fluorescence In Situ Hybridization (FISH) for molecular monitoring and quantification of bacteria in soil and aquifer samples is presented. FISH was applied to bacterial cells extracted from the matrix by density gradient centrifugation. This separation method was applied to soil and aquifer samples and produced high cell recovery of 76.5%+/-4.4 and 78.0%+/-3.2, respectively. FISH, performed on the harvested cells, permitted a perfect visualization and quantification of bacteria. This approach is therefore promising for in situ detection of indigenous bacterial communities in complex samples.     

Fluorescence in situ hybridization (CARD-FISH) of microorganisms in hydrocarbon contaminated aquifer sediment samples

Groundwater ecosystems are the most important sources of drinking water worldwide but they are threatened by contamination and overexploitation. Petroleum spills account for the most common source of contamination and the high carbon load results in anoxia and steep geochemical gradients. Microbes play a major role in the transformation of petroleum hydrocarbons into less toxic substances. To investigate microbial populations at the single cell level, fluorescence in situ hybridization (FISH) is now a well-established technique. Recently, however, catalyzed reporter deposition (CARD)-FISH has been introduced for the detection of microbes from oligotrophic environments. Nevertheless, petroleum contaminated aquifers present a worst case scenario for FISH techniques due to the combination of high background fluorescence of hydrocarbons and the presence of small microbial cells caused by the low turnover rates characteristic of groundwater ecosystems. It is therefore not surprising that studies of microorganisms from such sites are mostly based on cultivation techniques, fingerprinting, and amplicon sequencing. However, to reveal the population dynamics and interspecies relationships of the key participants of contaminant degradation, FISH is an indispensable tool. In this study, a protocol for FISH was developed in combination with cell quantification using an automated counting microscope. The protocol includes the separation and purification of microbial cells from sediment particles, cell permeabilization and, finally, CARD-FISH in a microwave oven. As a proof of principle, the distribution of Archaea and Bacteria was shown in 60 sediment samples taken across the contaminant plume of an aquifer (Leuna, Germany), which has been heavily contaminated with several ten-thousand tonnes of petroleum hydrocarbons since World War II.     

Decrease of the hydraulic conductivity of sand columns by Methanosarcina barkeri

The extent to which a methanogen can clog sand columns was examined: two permeameters packed with clean quartz sand were sterilized, saturated with water, inoculated with Methanosarcina barkeri and percolated under upward flow conditions. After approx. 5 months, the hydraulic conductivity of the sand had decreased to 3% and 25% of the highest values measured earlier. At that point, gas-filled regions in the sand were clearly visible through the transparent walls of the permeameters, and methane bubbles were continuously released from the columns into the effluent. Scanning electron microscopy observations and biomass assays indicated that cell mass accumulation did not contribute significantly to the observed decrease of the hydraulic conductivity. This decrease was therefore attributed to pore blocking due to the entrapment of methane bubbles.D. Sanchez de Lozada and P. Baveye are with the Department of Soil, Crop and Atmospheric Sciences, Bradfield Hall, Cornell University, Ithaca, NY 13853, USA; P. Vandevivere is with the College of Marine Studies, University of Delaware, Lewes, DE 19958, USA. S. Zinder is with the Department of Microbiology, Rice Hall, Cornell University, Ithaca, NY 14853, USA.     

The isotope hydrology of Quaternary climate change

Understanding the links between climate change and human migration and culture is an important theme in Quaternary archaeology. While oxygen and hydrogen stable isotopes in high-latitude ice cores provide the ultimate detailed record of palaeoclimate extending back to the Middle Pleistocene, groundwater can act as a climate archive for areas at lower latitudes, permitting a degree of calibration for proxy records such as lake sediments, bones, and organic matter. Not only can oxygen and hydrogen stable isotopes be measured on waters, but the temperature of recharge can be calculated from the amount of the atmospheric noble gases neon, argon, krypton, and xenon in solution, while residence time can be estimated from the decay of the radioisotopes carbon-14, chlorine-36, and krypton-81 over timescales comparable to the ice core record. The Pleistocene-Holocene transition is well characterised in aquifers worldwide, and it is apparent that isotope-temperature relationships of the present day are not necessarily transferable to past climatic regimes, with important implications for the interpretation of proxy isotope data. Groundwaters dating back to one million years, i.e., to beyond the Middle Pleistocene, are only found in major aquifer basins and information is relatively sparse and of low resolution. Speleothem fluid inclusions offer a way of considerably increasing this resolution, but both speleothem formation and large-scale groundwater recharge requires humid conditions, which may be relatively infrequent for areas currently experiencing arid climates. Both types of record therefore require caution in their interpretation when considering a particular archaeological context.     

Strontium isotopes in Melanopsis sp. as indicators of variation in hydrology and climate in the Upper Jordan Valley during the Early-Middle Pleistocene, and wider implications

Aquifers dominated by Pleistocene basalts and Jurassic to Cretaceous calcareous rocks feed the Hula basin which is drained by the Jordan River into Lake Kinneret. The sedimentary sequence of Lower-Middle Pleistocene Benot Ya‘akov Formation (BYF) exposed by excavations of the 0.78 Ma lake-side site of Gesher Benot Ya‘aqov (GBY) consists of six cycles representing ca. 100 ka history of the Hula basin. This study characterizes the types of water sources in the catchment, tests the use of the Strontium (Sr) isotopes in the common extant snail Melanopsis sp. as a tracer for water in its habitat, and uses this tracer in the fossil specimens from GBY to investigate the palaeohydrology of the Hula paleolake during the corresponding period.The Sr isotope composition (⁸⁷Sr/⁸⁶Sr) of extant Melanopsis shells in the Hula catchment range widely (0.7046-0.7079). These analyses define distinct groups of water sources and aquifers, while the Jordan River at the GBY site has values around 0.70685. The values for fossil Melanopsis from GBY vary along stratigraphy; they are highest around 0.70710 in Cycles 1 and 2, decrease to around 0.70685 in Cycle 3, and exhibit upward trending fluctuations in the subsequent cycles to 0.70703 in Cycle 6. This trend reveals the dominance of the Hermon Jurassic aquifer during the earlier, colder periods before the Matuyama-Brunhes Boundary (MBB) and enhanced influence of the Golan basaltic aquifers, in subsequent warmer periods, indicating that the MBB coincides with climate warming as supported by other indicators. Hence, this global geochronological indicator of 0.78 Ma is also potentially a global palaeoclimatic marker. The similarity between the Sr isotope composition of the Jordan River waters and Melanopsis and those from Cycle 3 suggests that the current climate corresponds to that of the warmest period within the record of GBY, clarifying the comparative interpretation of this 100 k.yr. climate record.