Groundwater Microbiome Research Articles

A Comparative Metagenomic Analysis of Specified Microorganisms in Groundwater for Non-Sterilized Pharmaceutical Products

In pharmaceutical manufacturing, ensuring product safety involves the detection and identification of microorganisms with human pathogenic potential, including Burkholderia cepacia complex (BCC), Escherichia coli, Pseudomonas aeruginosa, Salmonella enterica, Staphylococcus aureus, Clostridium sporogenes, Candida albicans, and Mycoplasma spp., some of which may be missed or not identified by traditional culture-dependent methods. In this study, we employed a metagenomic approach to detect these taxa, avoiding the limitations of conventional cultivation methods. We assessed the groundwater microbiome’s taxonomic and functional features from samples collected at two locations in the spring and summer. All datasets comprised 436–557 genera with Proteobacteria, Bacteroidota, Firmicutes, Actinobacteria, and Cyanobacteria accounting for > 95% of microbial DNA sequences. The aforementioned species constituted less than 18.3% of relative abundance. Escherichia and Salmonella were mainly detected in Hot Springs, relative to Jefferson, while Clostridium and Pseudomonas were mainly found in Jefferson relative to Hot Springs. Multidrug resistance efflux pumps and BlaR1 family regulatory sensor-transducer disambiguation dominated in Hot Springs and in Jefferson. These initial results provide insight into the detection of specified microorganisms and could constitute a framework for the establishment of comprehensive metagenomic analysis for the microbiological evaluation of pharmaceutical-grade water and other non-sterile pharmaceutical products, ensuring public safety.

Rapid Consumption of Dihydrogen Injected into a Shallow Aquifer by Ecophysiologically Different Microbes.

The envisaged future dihydrogen (H2) economy requires a H2 gas grid as well as large deep underground stores. However, the consequences of an unintended spread of H2 through leaky pipes, wells, or subterranean gas migrations on groundwater resources and their ecosystems are poorly understood. Therefore, we emulated a short-term leakage incident by injecting gaseous H2 into a shallow aquifer at the TestUM test site and monitored the subsequent biogeochemical processes in the groundwater system. At elevated H2 concentrations, an increase in acetate concentrations and a decrease in microbial α-diversity with a concomitant change in microbial β-diversity were observed. Additionally, microbial H2 oxidation was indicated by temporally higher abundances of taxa known for aerobic or anaerobic H2 oxidation. After H2 concentrations diminished below the detection limit, α- and β-diversity approached baseline values. In summary, the emulated H2 leakage resulted in a temporally limited change of the groundwater microbiome and associated geochemical conditions due to the intermediate growth of H2 consumers. The results confirm the general assumption that H2, being an excellent energy and electron source for many microorganisms, is quickly microbiologically consumed in the environment after a leakage.

Metabolic coupling of arsenic, carbon, nitrogen, and sulfur in high arsenic geothermal groundwater: Evidence from molecular mechanisms to community ecology

Groundwater arsenic (As) poses a global environmental problem and is regulated by complex biogeochemical processes. However, the As biogeochemistry and its metabolic coupling with carbon (C), nitrogen (N), and sulfur (S) in high As geothermal groundwater remain unclear. Here, we reported significant shifts in the geothermal groundwater microbiome and its functional ecological clusters along the flow path with increased As levels and dynamic As-C-N-S biogeochemical cycle from the Guide Basin, China. Strong associations among As(III), NH4+, HCO3−, and corresponding functional microbial taxa suggest that microbe-mediated As transformation, ammonification, and organic carbon biodegradation potentially contributed to the As mobilization in the discharge area. And As oxidizers (coupling with denitrification or carbon fixation) and S oxidizers were closely linked to the transformation of As(III) to immobile As(V) in the recharge area. Our study provides a comprehensive insight into the complex microbial As-C-N-S coupling network and its potential role in groundwater As mobilization under hydrological disturbances.

From the Mountain to the Valley - Drivers of Groundwater Prokaryotic Communities along an Alpine River Corridor

Rivers extend into the underlying groundwater which represents the unseen freshwater majority. Understanding microbial community composition and dynamics of shallow groundwater and lotic ecosystems is thus crucial, due to their potential impact on ecosystem processes and functioning. A 300 km section of the Mur river valley, from 2000 m.a.s.l. in the Austrian alps to the flats (200 m.a.s.l.) at the Slovenian border, was followed, analyzing river water from 14 stations and groundwater from 45 wells in early summer and late autumn. The active (RNA derived) and total prokaryotic communities were characterized using high-throughput gene amplicon sequencing. Key physico-chemical parameters and stress indicators were recorded. The groundwater microbiome is analyzed regarding its composition, change with land use, and difference to the river. Community composition and species turnover differed significantly. At high altitudes, dispersal limitation was the main driver of groundwater community assembly, whereas in the lowland, homogeneous selection explained the larger share. Land use was a key determinant of the groundwater microbiome composition. The alpine region was more diverse in prokaryotic taxa, with some early diverging archaeal lineages being highly abundant. This dataset shows a longitudinal change in prokaryotic communities that is dependent on regional differences affected by geomorphology and land use.

Environmental stressors altered the groundwater microbiome and nitrogen cycling: A focus on influencing mechanisms and pathways

Nitrogen cycling, as an important biogeochemical process in groundwater, strongly impacts the energy and matter flow of groundwater ecology. Phthalate esters (PAEs) were screened as key environmental stressors in the groundwater of Beijing, contributing to the alteration of microbial community structure and functions; thus, it could be deduced that these stressors might influence nitrogen cycling that is almost exclusively mediated by microorganisms. Identification of the influences of PAEs on groundwater nitrogen cycling and exploration of the potential influence mechanisms and pathways are vital but still challenging. This study explored the influence mechanisms and pathways of the environmental stressor PAE on nitrogen cycling in groundwater collected from a typical monitoring station in Beijing based on high-throughput sequencing and bioinformatics analysis combined with mediation analysis methods. The results suggested that among the 5 detected PAEs, dimethyl phthalate and diethyl phthalate significantly negatively impacted nitrogen cycling processes, especially nitrogen fixation and denitrification processes (p < 0.05), in groundwater. Their influences were fully or partially mediated by functional microorganisms, particularly assigned keystone genera (such as Dechloromonas, Aeromonas and Noviherbaspirillum), whose abundance was significantly inhibited by these PAEs via dysregulation of carbohydrate metabolism and activation of defense mechanisms. These findings confirmed that the influences of environmental stressors PAEs on nitrogen cycling in groundwater might be mediated by the “PAE stress–groundwater microbiome–nitrogen cycling alteration” pathway. This study may advance the understanding of the consequences of environmental stressors on groundwater ecology and support the ecological hazard assessment of groundwater stressors.

Linking groundwater microbiome and functional ecological clusters to geogenic high hexavalent chromium from deep aquifers in a loess plateau

Geogenic high hexavalent chromium [Cr(Ⅵ)] in groundwater is a global environmental problem. However, the groundwater microbiome and its linkage to geogenic high Cr(Ⅵ) from deep aquifers still need to be elucidated. Here, we evaluated geogenic Cr(Ⅵ), groundwater microbiome with featured functional ecological clusters and their interactive responses in groundwater from a deep aquifer in a loess plateau of Northern Shaanxi, China. We found that the compositions and structures of microbial communities in groundwater from the deep aquifer were significantly different between low Cr(Ⅵ) (LCG, < 50 μg/L) and high Cr(Ⅵ) groundwater (HCG, > 50 μg/L), with higher microbial diversity and richness in HCG (p < 0.05). Functional “specialists” related to Cr biotransformation, including Cr(Ⅵ) reducing bacteria (CRB) Rhodococcus, Nocardioides, Novosphingobium, and Acidovorax and Mn-oxidizing bacteria (MnOB) Sphingobium, and Ralstonia were positively correlated to total Cr and Cr(VI) concentrations in groundwater. Moreover, these CRB and MnOB were dominant in high Cr(VI) groundwater and associated by strong interspecific relation in an ecological cluster (p < 0.05), suggesting their indicator roles for high Cr(Ⅵ) and the contribution of MnOB mediated Cr(III) oxidation to Cr(VI) enrichment. RDA and path analysis further revealed that the geogenic Cr(Ⅵ) directly promoted the key Cr-related functional cluster with the groundwater depth, dissolved oxygen, and total dissolved solids as the cofactors indirectly influencing Cr(Ⅵ) and the functional clusters (p < 0.05). Collectively, our results highlight the significant roles of microbial ecological clusters especially functional “specialists” MnOB and CRB in groundwater Cr(Ⅵ) from deep aquifers in the loess plateau and provide a basis for sustainable management of high Cr(Ⅵ) groundwater.

Ecological differentiation and assembly processes of abundant and rare bacterial subcommunities in karst groundwater.

The ecological health of karst groundwater has been of global concern due to increasing anthropogenic activities. Bacteria comprising a few abundant taxa (AT) and plentiful rare taxa (RT) play essential roles in maintaining ecosystem stability, yet limited information is known about their ecological differentiation and assembly processes in karst groundwater. Based on a metabarcoding analysis of 64 groundwater samples from typical karst regions in southwest China, we revealed the environmental drivers, ecological roles, and assembly mechanisms of abundant and rare bacterial communities. We found a relatively high abundance of potential functional groups associated with parasites and pathogens in karst groundwater, which might be linked to the frequent regional anthropogenic activities. Our study confirmed that AT was dominated by Proteobacteria and Campilobacterota, while Patescibacteria and Chloroflexi flourished more in the RT subcommunity. The node-level topological features of the co-occurrence network indicated that AT might share similar niches and play more important roles in maintaining bacterial community stability. RT in karst groundwater was less environmentally constrained and showed a wider environmental threshold response to various environmental factors than AT. Deterministic processes, especially homogeneous selection, tended to be more important in the community assembly of AT, whereas the community assembly of RT was mainly controlled by stochastic processes. This study expanded our knowledge of the karst groundwater microbiome and was of great significance to the assessment of ecological stability and drinking water safety in karst regions.

Effect of Snowmelt on Groundwater Bacterial Community Composition and Potential Role of Surface Environments as Microbial Seed Bank in Two Distinct Aquifers from the Region of Quebec, Canada.

Events of groundwater recharge are associated with changes in the composition of aquifer microbial communities but also abiotic conditions. Modification in the structure of the community can be the result of different environmental condition favoring or hindering certain taxa, or due to the introduction of surface-derived taxa. Yet, in both cases, the local hydrogeochemical settings of the aquifer is likely to affect the amount of variation observed. Therefore, in our study, we used 16S rRNA gene sequencing to assess how microbial communities change in response to snowmelt and the potential connectivity between subsurface and surface microbiomes in two distinct aquifers located in the region of Vaudreuil-Soulanges (Québec, Canada). At both sites, we observed an increase in groundwater level and decrease in temperature following the onset of snow melt in March 2019. Bacterial community composition of each aquifer was significantly different (p < 0.05) between samples collected prior and after groundwater recharge. Furthermore, microbial source tracking results suggested a low contribution of surface environments to the groundwater microbiome except for in the months associated with recharge (March 2019 and April 2019). Overall, despite differences in soil permeability between both sites, the period of snow melt was followed by important changes in the composition of microbial communities from aquifers.

From the Mountain to the Valley: Drivers of Groundwater Prokaryotic Communities along an Alpine River Corridor.

Rivers are the "tip of the iceberg", with the underlying groundwater being the unseen freshwater majority. Microbial community composition and the dynamics of shallow groundwater ecosystems are thus crucial, due to their potential impact on ecosystem processes and functioning. In early summer and late autumn, samples of river water from 14 stations and groundwater from 45 wells were analyzed along a 300 km transect of the Mur River valley, from the Austrian alps to the flats at the Slovenian border. The active and total prokaryotic communities were characterized using high-throughput gene amplicon sequencing. Key physico-chemical parameters and stress indicators were recorded. The dataset was used to challenge ecological concepts and assembly processes in shallow aquifers. The groundwater microbiome is analyzed regarding its composition, change with land use, and difference to the river. Community composition and species turnover differed significantly. At high altitudes, dispersal limitation was the main driver of groundwater community assembly, whereas in the lowland, homogeneous selection explained the larger share. Land use was a key determinant of the groundwater microbiome composition. The alpine region was more diverse and richer in prokaryotic taxa, with some early diverging archaeal lineages being highly abundant. This dataset shows a longitudinal change in prokaryotic communities that is dependent on regional differences affected by geomorphology and land use.

Formate: A promising electron donor to enhance trichloroethene-to-ethene dechlorination in Dehalococcoides-augmented groundwater ecosystems with minimal bacterial growth

Various substrates have been used to stimulate habitat microbes in chloroethene-contaminated groundwater, however, the specific efficiency and minimum growth of microbes have rarely been studied. This study investigated the effects of seven substrates on trichloroethene (TCE) dechlorination by augmentation of groundwater with Dehalococcoides mccartyi NIT01 and its contribution to the microbial community. Three out of eight test groups completed dechlorination of 1 mM TCE-to-ethene in varying durations; groundwater supplemented with formate (FOR) required 78 days, whereas the microcosms with lactate (LAC) and citrate (CIT) required approximately twice as long (143 days). The calculated efficiency of how much produced H2 was used in dechlorination indicated a higher efficiency in FOR (36%) compared with LAC (1.9%) or CIT (2.9%). FOR showed lower microbial growth (3.4 × 105 copies/mL) than LAC (1.5 × 106) or CIT (4.4 × 106), and maintained a higher Shannon diversity index (5.65) than LAC (4.97) and CIT (4.30). The rapid and higher H2 transfer efficiency with lower bacterial growth by using formate was attributed to the slightly positive Gibbs free energy identified in H2 production requiring a H2-utilizer, lower carbon in the molecule, and adaptation to metabolic potential of the original groundwater microbiome. Formate is, therefore, a promising electron donor for rapid Dehalococcoides-augmented remediation with minimum bacterial growth. Sequential transferring of the FOR culture successfully maintained TCE-to-ethene dechlorination activity and enriched the members of genera Dehalococcoides (33%), Methanosphaerula (23%), Rectinema (13%), and Desulfitobacterium (5.6%). This suggests that formate is transferred to H2 and acetate, and provided to Dehalococcoides.

Bacterial Necromass Is Rapidly Metabolized by Heterotrophic Bacteria and Supports Multiple Trophic Levels of the Groundwater Microbiome.

ABSTRACTPristine groundwater is a highly stable environment with microbes adapted to dark, oligotrophic conditions. Input events like heavy rainfalls can introduce the excess particulate organic matter, including surface-derived microorganisms, thereby disturbing the groundwater microbiome. Some surface-derived bacteria will not survive this translocation, leading to an input of necromass to the groundwater. Here, we investigated the effects of necromass addition to the microbial community in fractured bedrock groundwater, using groundwater mesocosms as model systems. We followed the uptake of 13C-labeled necromass by the bacterial and eukaryotic groundwater community quantitatively and over time using a complementary protein-stable and DNA-stable isotope probing approach. Necromass was rapidly depleted in the mesocosms within 4 days, accompanied by a strong decrease in Shannon diversity and a 10-fold increase in bacterial 16S rRNA gene copy numbers. Species of Flavobacterium, Massilia, Rheinheimera, Rhodoferax, and Undibacterium dominated the microbial community within 2 days and were identified as key players in necromass degradation, based on a 13C incorporation of >90% in their peptides. Their proteomes comprised various proteins for uptake and transport functions and amino acid metabolization. After 4 and 8 days, the autotrophic and mixotrophic taxa Nitrosomonas, Limnohabitans, Paucibacter, and Acidovorax increased in abundance with a 13C incorporation between 0.5% and 23%. Likewise, eukaryotes assimilated necromass-derived carbon either directly or indirectly. Our data point toward a fast and exclusive uptake of labeled necromass by a few specialists followed by a concerted action of groundwater microorganisms, including autotrophs presumably fueled by released, reduced nitrogen and sulfur compounds generated during necromass degradation.IMPORTANCE Subsurface microbiomes provide essential ecosystem services, like the generation of drinking water. These ecosystems are devoid of light-driven primary production, and microbial life is adapted to the resulting oligotrophic conditions. Modern groundwater is most vulnerable to anthropogenic and climatic impacts. Heavy rainfalls, which will increase with climate change, can result in high surface inputs into shallow aquifers by percolation or lateral flow. These inputs include terrestrial organic matter and surface-derived microbes that are not all capable to flourish in aquatic subsurface habitats. Here, we investigated the response of groundwater mesocosms to the addition of bacterial necromass, simulating event-driven surface input. We found that the groundwater microbiome responds with a rapid bloom of only a few primary degraders, followed by the activation of typical groundwater autotrophs and mixotrophs, as well as eukaryotes. Our results suggest that this multiphase strategy is essential to maintain the balance of the groundwater microbiome to provide ecosystem services.

Elevated levels of antibiotic resistance in groundwater during treated wastewater irrigation associated with infiltration and accumulation of antibiotic residues

Treated wastewater irrigation (TWW) releases antibiotics and antibiotic resistance genes (ARGs) into the environment and might thus promote the dissemination of antibiotic resistance in groundwater (GW). We hypothesized that TWW irrigation increases ARG abundance in GW through two potential mechanisms: the contamination of GW with resistant bacteria and the accumulation of antibiotics in GW. To test this, the GW below a real-scale TWW-irrigated field was sampled for six months. Sampling took place before, during and after high-intensity TWW irrigation. Samples were analysed with 16S rRNA amplicon sequencing, qPCR of six ARGs and the class 1 integron-integrase gene intI1, while liquid chromatography tandem mass spectrometry was performed to detect antibiotic and pharmaceutical residues. Absolute abundance of 16S rRNA in GW decreased rather than increased during long-term irrigation. Also, the relative abundance of TWW-related bacteria did not increase in GW during long-term irrigation. In contrast, long-term TWW irrigation increased the relative abundance of sul1 and intI1 in the GW microbiome. Furthermore, GW contained elevated concentrations of sulfonamide antibiotics, especially sulfamethoxazole, to which sul1 confers resistance. Total sulfonamide concentrations in GW correlated with sul1 relative abundance. Consequently, TWW irrigation promoted sul1 and intI1 dissemination in the GW microbiome, most likely due to the accumulation of drug residues.

Energy efficiency and biological interactions define the core microbiome of deep oligotrophic groundwater

While oligotrophic deep groundwaters host active microbes attuned to the low-end of the bioenergetics spectrum, the ecological constraints on microbial niches in these ecosystems and their consequences for microbiome convergence are unknown. Here, we provide a genome-resolved, integrated omics analysis comparing archaeal and bacterial communities in disconnected fracture fluids of the Fennoscandian Shield in Europe. Leveraging a dataset that combines metagenomes, single cell genomes, and metatranscriptomes, we show that groundwaters flowing in similar lithologies offer fixed niches that are occupied by a common core microbiome. Functional expression analysis highlights that these deep groundwater ecosystems foster diverse, yet cooperative communities adapted to this setting. We suggest that these communities stimulate cooperation by expression of functions related to ecological traits, such as aggregate or biofilm formation, while alleviating the burden on microorganisms producing compounds or functions that provide a collective benefit by facilitating reciprocal promiscuous metabolic partnerships with other members of the community. We hypothesize that an episodic lifestyle enabled by reversible bacteriostatic functions ensures the subsistence of the oligotrophic deep groundwater microbiome.

Ecogenomics of Groundwater Phages Suggests Niche Differentiation Linked to Specific Environmental Tolerance.

ABSTRACTViruses are ubiquitous microbiome components, shaping ecosystems via strain-specific predation, horizontal gene transfer and redistribution of nutrients through host lysis. Viral impacts are important in groundwater ecosystems, where microbes drive many nutrient fluxes and metabolic processes; however, little is known about the diversity of viruses in these environments. We analyzed four groundwater plasmidomes (the entire plasmid content of an environment) and identified 200 viral sequences, which clustered into 41 genus-level viral clusters (approximately equivalent to viral genera) including 9 known and 32 putative new genera. We used publicly available bacterial whole-genome sequences (WGS) and WGS from 261 bacterial isolates from this groundwater environment to identify potential viral hosts. We linked 76 of the 200 viral sequences to a range of bacterial phyla, the majority associated with Proteobacteria, followed by Firmicutes, Bacteroidetes, and Actinobacteria. The publicly available WGS enabled mapping bacterial hosts to several viral sequences. The WGS of groundwater isolates increased the depth of host prediction by allowing host identification at the strain level. The latter included 4 viruses that were almost entirely (>99% query coverage, >99% identity) identified as integrated in the genomes of Pseudomonas, Acidovorax, and Castellaniella strains, resulting in high-confidence host assignments. Lastly, 21 of these viruses carried putative auxiliary metabolite genes for metal and antibiotic resistance, which might drive their infection cycles and/or provide selective advantage to infected hosts. Exploring the groundwater virome provides a necessary foundation for integration of viruses into ecosystem models where they are key players in microbial adaption to environmental stress.IMPORTANCE To our knowledge, this is the first study to identify the bacteriophage distribution in a groundwater ecosystem shedding light on their prevalence and distribution across metal-contaminated and background sites. Our study is uniquely based on selective sequencing of solely the extrachromosomal elements of a microbiome followed by analysis for viral signatures, thus establishing a more focused approach for phage identifications. Using this method, we detected several novel phage genera along with those previously established. Our approach of using the whole-genome sequences of hundreds of bacterial isolates from the same site enabled us to make host assignments with high confidence, several at strain levels. Certain phage genes suggest that they provide an environment-specific selective advantage to their bacterial hosts. Our study lays the foundation for future research on directed phage isolations using specific bacterial host strains to further characterize groundwater phages, their life cycles, and their effects on groundwater microbiome and biogeochemistry.

Groundwater bacterial communities evolve over time in response to recharge

Time series analyses are a crucial tool for uncovering the patterns and processes shaping microbial communities and their functions, especially in aquatic ecosystems. Subsurface aquatic environments are perceived to be more stable than surface oceans and lakes, due to the lack of sunlight, the absence of photosysnthetically-driven primary production, low temperature variations, and oligotrophic conditions. However, periodic groundwater recharge should affect the structure and succession of groundwater microbiomes. To disentangle the long-term temporal changes in bacterial communities of shallow fractured bedrock groundwater, and identify the drivers of the observed patterns, we analysed bacterial 16S rRNA gene sequencing data for samples collected monthly from three groundwater wells over a six-year period (n = 230) along a hillslope recharge area. We showed that the bacterial communities in the groundwater of limestone-mudstone alternations were not stable over time and exhibited non-linear dissimilarity patterns which corresponded to periods of groundwater recharge. Further, we observed an increase in dissimilarity over time (generalized additive model P < 0.001) indicating that the successive recharge events result in communities that are increasingly more dissimilar to the initial reference time point. The sampling period was able to explain up to 29.5% of the variability in bacterial community composition and the impact of recharge events on the groundwater microbiome was linked to the strength of the recharge and local environmental selection. Many groundwater bacteria originated from the recharge-related sources (mean = 66.5%, SD = 15.1%) and specific bacterial taxa were identified as being either enriched or repressed during recharge events. Overall, similar to surface aquatic environments, the microbiomes in shallow fractured-rock groundwater vary through time, though we revealed groundwater recharges as unique driving factors for these patterns. The high temporal resolution employed here highlights the dynamics of bacterial communities in groundwater, which is an essential resource for the provision of clean drinking water; understanding the biological complexities of these systems is therefore crucial.

Different Effects of Mineral Versus Vegetal Granular Activated Carbon Filters on the Microbial Community Composition of a Drinking Water Treatment Plant

Drinking water quality and safety is strictly regulated and constantly monitored, but little is known about the microorganisms inhabiting drinking water treatment plants (DWTPs). This lack of knowledge prevents optimization of designs and operational controls. Here we investigated the drinking water microbial community harbored by a groundwater-derived DWTP, involving mineral and vegetal granular activated carbon filters (GACs). We used 16S rRNA gene sequencing to analyze water microbiome variations through the potabilization process, considering (i) different GAC materials and (ii) time from GAC regeneration. Our results revealed the predominance of Cand. Patescibacteria, uncultivable bacteria with limited metabolic capacities and small genomes, from source to downstream water. Microbial communities clustered per sampling date, with the noteworthy exception of groundwater samples. If the groundwater microbiome showed no significant variations over time, the community structure of water downstream GACs (both mineral and vegetal) seemed to be affected by time from GAC regeneration. Looking at a finer scale, different GAC material affected microbiome assembly over time with significant variation in the relative abundances of specific taxa. The significance of our research is in identifying the environmental microorganisms intrinsic of deep groundwater and the community shift after the perturbations induced by potabilization processes. Which microorganisms colonize different GACs and become abundant after GACs regeneration and over time is a first step toward advanced control of microbial communities, improving drinking water safety and management of operational costs.

Monitoring of the effects of a temporally limited heat stress on microbial communities in a shallow aquifer.

Aquifer thermal energy storage (ATES) is a key concept for the use of renewable energy resources. Interest in ATES performed at high temperature (HT-ATES; > 60 °C) is increasing due to higher energetic efficiencies. HT-ATES induces temperature fluctuations that exceed the natural variability in shallow aquifers, which could lead to adverse effects in subsurface ecosystems by altering the groundwater chemistry, biodiversity, and microbial metabolic activity, resulting in changes of the groundwater quality, biogeochemical processes, and ecosystem functions. The aim of this study was to emulate the initial operating phase of a HT-ATES system with a short-term infiltration of warm water into Pleistocene sandur sediment and, consequently, to monitor the thermal effects on the groundwater microbiome inhabiting an imitated affected space of an HT-ATES system. Therefore, local groundwater was withdrawn, heated up to 75 °C, and re-infiltrated into a shallow aquifer located near Wittstock/Dosse (Brandenburg, Germany) for around five days. Groundwater samples taken regularly before and after the infiltration were analyzed by 16S rRNA gene amplicon sequencing for microbial diversity analyses as well as total cell counting. During the infiltration, a thermal plume with groundwater temperatures increasing from 9 ± 2 to up to ~65 °C was recorded. The highest temperature at which groundwater samples were taken was 34.9 °C, a temperature typically arising in the affected space of an HT-ATES system. The microbial communities in the groundwater were mainly composed of Gammaproteobacteria, Alphaproteobacteria, Bacteroidia, and Actinobacteria, and the total cell numbers ranged from 3.2 * 104 to 3.1 * 106 cells ml-1. Neither the compositions of the microbial communities nor the total number of cells in groundwater were significantly changed upon moderate temperature increase, indicating that the diverse groundwater microbiome was resilient to the temporally limited heat stress.

Microbial intrusion and seasonal dynamics in the groundwater microbiome of a porous basaltic rock aquifer used as municipal water reservoir.

Groundwater is a key resource for safe drinking water supply. Yet unconfined aquifers can be vulnerable to microbial contamination during extreme weather events that lead to surface runoff. The current study characterises the groundwater microbiome of a porous basaltic rock aquifer in South-West Iceland used for drinking water extraction and analyses the microbial community dynamics during surface runoff. The groundwater microbial community sampled from 12 wells across the extraction area contained over 745 prokaryotic genera and was phylogenetically similar between wells and most seasons, representing a diverse but homogenous ecosystem. The largest seasonal variation in the microbial community composition was detected during a period of concurrent snow melt and high precipitation leading to surface runoff. This period was characterised by an increased abundance of soil-associated taxa in the groundwater microbiome and specifically of taxa assigned to Aeromonas and Bacillus. A field experiment simulating high surface runoff around a groundwater well confirmed the increased abundance of surface soil microorganisms in the well water, indicating vulnerability of groundwater towards surface microbial intrusion during extreme weather events. As such events are likely to increase due to climate change, novel water management tools such as microbial community analysis could help ensure drinking water safety.

Geochemical, metagenomic, and physiological characterization of the multifaceted interaction between microbiome of an arsenic contaminated groundwater and aquifer sediment

Geomicrobiological details of the interactions between groundwater microbiome (GWM) and arsenic (As)-rich aquifer sediment of Bengal basin was investigated through microcosm incubations. Role of key microorganisms and their specific interactions with As-bearing minerals was demarcated under organic carbon- amended and -unamended conditions. Acinetobacter (50.8 %), Brevundimonas (7.9 %), Sideroxydans (3.4 %), Alkanindiges (3.0 %) dominated the GWM. The microbiome catalysed considerable alterations in As-bearing mineral [Fe-(hydr)oxide and aluminosilicate] phases resulting in substantial changes in overall geochemistry and release of As (65 μg/L) and Fe (118 μg/L). Synergistic roles of autotrophic, NH4+-oxidizing Archaea (Thaumarchaeota) and chemoheterotrophic bacteria (Stenotrophomonas, Pseudomonas, Geobacter) of diverse metabolic abilities (NH4+-oxidizing, NO3−, As/Fe-reducing) were noted for observed changes. Organic carbon supported enhanced microbial growth and As mobilization (upto 403.2 μg As/L) from multiple mineral phases (hematite, magnetite, maghemite, biotite, etc.). In presence of high organic carbon, concerted actions of anaerobic, hydrocarbon-utilizing, As-, Fe-reducing Rhizobium, fermentative Escherichia, anaerobic Bacillales, metal-reducing and organic acid-utilizing Pseudomonas and Achromobacter were implicated in altering sediment mineralogy and biogeochemistry. Increase in abundance of arrA, arsC, bssA genes, and dissolution of Fe, Ca, Mg, Mn confirmed that dissimilatory-, cytosolic-As reduction, and mineral weathering fuelled by anaerobic (hydro)carbon metabolism are the predominant mechanisms of As release in aquifers of Bengal basin.

Organic carbon and nutrients drive prokaryote and metazoan communities in a floodplain aquifer

In an alluvial aquifer in the River Fulda Valley (Germany) the influence of agricultural inputs on the subterranean physical, chemical and biological relationships was examined. A 40-year-old (1977–1981) comprehensive data set on the groundwater microbiome plus metazoa was now analysed for the first time in full (measurements for up to 4 years: hydrological, chemical, physical, prokaryote, and metazoa characteristics). Four hydrogeochemically different groundwater zones were identified across the floodplain. In addition, the prokaryote (Archaea and Bacteria) and metazoan communities differed among the four zones. The hydraulic exchange between the alluvial aquifer and the River Fulda influenced the sites closest to the river, leading to the highest prokaryote and metazoan biomasses at these locations. An organic carbon plume zone of anthropogenic origin exhibited high prokaryote abundances and production, which were higher than in the surrounding mixing zone. This mixing zone represented a transition area to the river-influenced sites as well as to the fourth zone, which was characterized by high nutrient levels from intense agriculture and which exhibited low prokaryote abundance and activity and intermediate metazoan abundance. Despite high prokaryote productivity, metazoa did not favor the organic carbon plume, due probably to low oxygen concentrations. At the sites, where metazoa occurred, their biomass corresponded mostly to about one hundredth of the prokaryote biomass. The main implication from this new analysis of an old data set is that even on a coarse taxonomical resolution, patterns emerge that show – in a geologically homogeneous area – an unprecedented complexity among different groundwater zones resulting from different external influences of natural as well as anthropogenic origin. Future studies need to ascertain an adequate temporal and spatial resolution.