Broad Dissemination of Plasmids across Groundwater-Fed Rapid Sand Filter Microbiomes

The supply of clean water for human consumption is being challenged by the appearance of anthropogenic pollutants in groundwater ecosystems. Because many plasmids can transfer horizontally between members of bacterial communities, they comprise promising vectors for the dissemination of pollutant-degrading genetic determinants within water purification plants.

Molecular characterization of bacterial diversity in New Zealand groundwater

<p>Groundwater is a globally important natural resource and an integral part of the water supply in New Zealand. Due to high demand, the quality and availability of groundwater are both extensively monitored in New Zealand and globally, under State-of-the-Environment (SOE) monitoring programmes. SOE groundwater monitoring in New Zealand mainly evaluates hydrochemistry and until this thesis has largely overlooked the biotic component. Microbes including bacteria play a crucial role in ecosystem functioning by mediating biogeochemical processes in subsurface environments. Therefore, analysis of microbiological content will enable better evaluation of the health of groundwater ecosystems that is not fully reflected by chemical data alone. This project characterizes the bacterial diversity in New Zealand groundwater at national and regional scales using molecular methods and explores the underlying factors that shape the bacterial community structure. A simple molecular profiling tool, Terminal Restriction Fragment Length Polymorphism (T-RFLP) was used to determine community structure at local and national scales. The results revealed considerable diversity that was driven by groundwater chemistry. Roche 454-pyrosequencing was then used to obtain a deeper insight into New Zealand groundwater ecosystems, and showed that bacterial communities have many low abundance taxa and relatively few highly abundant species. In addition, microbial diversity is mainly related to the redox potential of the groundwater. But, despite this relationship, Pseudomonas spp. were the dominant genus at many sites even those with diverse chemistries and environmental factors. The final phase of the project set the platform to test whether these Pseudomonas spp. have acquired genetic material from other species via horizontal gene transfer (HGT) enabling them to adapt into a diverse range of habitats. A whole-genome sequencing approach (Illumina MiSeq platform) was used to develop six metagenomic databases as a resource to test this hypothesis. Initial results show some evidence for HGT and further investigations are underway. Overall, the knowledge generated across all phases of this project provides novel insights into New Zealand groundwater ecosystems and creates a scientific basis for the future inclusion of microbial status assessment criteria into regional and national groundwater monitoring programmes and related policies in New Zealand.</p>

Molecular characterization of bacterial diversity in New Zealand groundwater

<p>Groundwater is a globally important natural resource and an integral part of the water supply in New Zealand. Due to high demand, the quality and availability of groundwater are both extensively monitored in New Zealand and globally, under State-of-the-Environment (SOE) monitoring programmes. SOE groundwater monitoring in New Zealand mainly evaluates hydrochemistry and until this thesis has largely overlooked the biotic component. Microbes including bacteria play a crucial role in ecosystem functioning by mediating biogeochemical processes in subsurface environments. Therefore, analysis of microbiological content will enable better evaluation of the health of groundwater ecosystems that is not fully reflected by chemical data alone. This project characterizes the bacterial diversity in New Zealand groundwater at national and regional scales using molecular methods and explores the underlying factors that shape the bacterial community structure. A simple molecular profiling tool, Terminal Restriction Fragment Length Polymorphism (T-RFLP) was used to determine community structure at local and national scales. The results revealed considerable diversity that was driven by groundwater chemistry. Roche 454-pyrosequencing was then used to obtain a deeper insight into New Zealand groundwater ecosystems, and showed that bacterial communities have many low abundance taxa and relatively few highly abundant species. In addition, microbial diversity is mainly related to the redox potential of the groundwater. But, despite this relationship, Pseudomonas spp. were the dominant genus at many sites even those with diverse chemistries and environmental factors. The final phase of the project set the platform to test whether these Pseudomonas spp. have acquired genetic material from other species via horizontal gene transfer (HGT) enabling them to adapt into a diverse range of habitats. A whole-genome sequencing approach (Illumina MiSeq platform) was used to develop six metagenomic databases as a resource to test this hypothesis. Initial results show some evidence for HGT and further investigations are underway. Overall, the knowledge generated across all phases of this project provides novel insights into New Zealand groundwater ecosystems and creates a scientific basis for the future inclusion of microbial status assessment criteria into regional and national groundwater monitoring programmes and related policies in New Zealand.</p>

Application of the D-A-(C) index as a simple tool for microbial-ecological characterization and assessment of groundwater ecosystems—a case study of the Mur River Valley, Austria

The assessment and monitoring of the ecological quality and status of groundwater is a timely issue. At present, various assessment tools have been developed that now await application and validation. One of these, the D‑A‑C index, evaluates the microbiological-ecological quality of groundwater based on of prokaryotic cell counts, microbial activity measurements, and the qualitative characterization of dissolved organic carbon (DOM). The purpose of this paper is to illustrate the different ways of application of the D‑A-(C) index making use of a recently collected data set (n = 61) from the river Mur valley, Austria. First, we present an extension of the D‑A-(C) index by including measurements of dissolved organic matter quality (DOM) derived from fluorescence spectroscopy as additional variables to supplement the analysis of microbial cell density and activity levels. Second, we illustrate how the definition of a reference status for a ‘good’ microbiological-ecological state can improve the analysis and allow for a more sensitive and accurate detection of impacts on groundwater ecosystems. Based on our results, we advocate that the analysis be performed by making use of expert knowledge for the definition of reference sites to which target sites are to be compared.

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.

Novel Protocol for Acute In Situ Ecotoxicity Test Using Native Crustaceans Applied to Groundwater Ecosystems

Current standardized laboratory test protocols use model species that have limitations to accurately assess native species responses to stressors. We developed and tested a novel acute in situ protocol for testing field-collected organisms. We used Asellus aquaticus and NaCl as a reference toxicant to test for the effects of location (laboratory vs. in situ), medium (synthetic vs. field water), substrate (presence vs. absence), and protocol replicability. We further tested the protocol using groundwater-adapted isopods: Proasellus assaforensis for the effect of location, P. cavaticus of medium and P.lusitanicus of substrate. Our results showed that A.aquaticus’ lethality obtained with the novel acute in situ protocol did not significantly differ from those from laboratory testing. However, laboratory tested P.assaforensis showed a higher sensitivity, suggesting that its acclimation to laboratory conditions might have pernicious effects. A. aquaticus and P. cavaticus showed a higher mortality using synthetic medium in situ and under laboratory conditions, which overestimated the stressor’s effect. Besides, substrate use had no significant effect. The novel acute in situ protocol allows the use of native species under realistic scenarios. It is particularly well adapted for assessing the risk of groundwater ecosystems but it can be applied to a wide range of ecosystems.

Using the molecular composition of dissolved organic matter in groundwater to assess the functional stability of subsurface ecosystems

&lt;p&gt;With surface systems changing rapidly on a global scale, it is important to understand how this will affect groundwater resources and ecosystems in the subsurface. The molecular composition of dissolved organic matter (DOM) integrates essential information on metabolic functioning and could therefore reveal changes of groundwater ecosystems in high detail. Here, we evaluate a 6-year time series of ultrahigh-resolution DOM composition analysis of groundwater from a hillslope well transect within the Hainich Critical Zone Exploratory, Germany. We predict ecosystem functionality by assigning molecular sum formulas to metabolic pathways via the KEGG database. Our data support hydrogeological characterizations of a compartmentalized fractured multi-storey aquifer system and reveal distinct metabolic functions that largely depend on the compartment&amp;#8217;s relative surface-connectivity or isolation. We show that seasonal fluctuation of groundwater levels, coinciding with cross-stratal exchange can substantially impact the local inventory of functional metabolites in DOM. Furthermore, we find that extreme conditions of groundwater recharge following pronounced groundwater lowstand cause strong alterations of the functional metabolome in DOM even in aquifer compartments, which usually show minimal variation in DOM composition. Our findings suggest that bedrock groundwater ecosystems might be functionally vulnerable to hydrogeological extremes.&lt;/p&gt;

Rainfall as a trigger of ecological cascade effects in an Australian groundwater ecosystem

Groundwaters host vital resources playing a key role in the near future. Subterranean fauna and microbes are crucial in regulating organic cycles in environments characterized by low energy and scarce carbon availability. However, our knowledge about the functioning of groundwater ecosystems is limited, despite being increasingly exposed to anthropic impacts and climate change-related processes. In this work we apply novel biochemical and genetic techniques to investigate the ecological dynamics of an Australian calcrete under two contrasting rainfall periods (LR—low rainfall and HR—high rainfall). Our results indicate that the microbial gut community of copepods and amphipods experienced a shift in taxonomic diversity and predicted organic functional metabolic pathways during HR. The HR regime triggered a cascade effect driven by microbes (OM processors) and exploited by copepods and amphipods (primary and secondary consumers), which was finally transferred to the aquatic beetles (top predators). Our findings highlight that rainfall triggers ecological shifts towards more deterministic dynamics, revealing a complex web of interactions in seemingly simple environmental settings. Here we show how a combined isotopic-molecular approach can untangle the mechanisms shaping a calcrete community. This design will help manage and preserve one of the most vital but underrated ecosystems worldwide.

Genome-resolved metagenomics reveals site-specific diversity of episymbiotic CPR bacteria and DPANN archaea in groundwater ecosystems

Candidate phyla radiation (CPR) bacteria and DPANN archaea are unisolated, small-celled symbionts that are often detected in groundwater. The effects of groundwater geochemistry on the abundance, distribution, taxonomic diversity and host association of CPR bacteria and DPANN archaea has not been studied. Here, we performed genome-resolved metagenomic analysis of one agricultural and seven pristine groundwater microbial communities and recovered 746 CPR and DPANN genomes in total. The pristine sites, which serve as local sources of drinking water, contained up to 31% CPR bacteria and 4% DPANN archaea. We observed little species-level overlap of metagenome-assembled genomes (MAGs) across the groundwater sites, indicating that CPR and DPANN communities may be differentiated according to physicochemical conditions and host populations. Cryogenic transmission electron microscopy imaging and genomic analyses enabled us to identify CPR and DPANN lineages that reproducibly attach to host cells and showed that the growth of CPR bacteria seems to be stimulated by attachment to host-cell surfaces. Our analysis reveals site-specific diversity of CPR bacteria and DPANN archaea that coexist with diverse hosts in groundwater aquifers. Given that CPR and DPANN organisms have been identified in human microbiomes and their presence is correlated with diseases such as periodontitis, our findings are relevant to considerations of drinking water quality and human health.