Antimicrobial resistance (AMR) is among the top 10 public health threats, with nearly 5 million deaths in 2019 linked to AMR-related bacterial infections. (1) A One Health approach is needed to combat AMR.
Healthcare-based surveillance (HBS) of AMR provides incomplete information about the scope of the AMR threat. HBS screens only patients seeking medical attention, lacking community-level representativeness, and suffers from underreporting. (2) Consequently, researchers are turning to wastewater-based surveillance (WBS) to complement HBS. (3) WBS can provide information about AMR circulating within communities and hospitals, offering a comprehensive understanding of AMR prevalence. However, the surveillance targets and data obtained from WBS are distinct from those derived from HBS, creating uncertainty regarding their utility to the public health sector and ability to yield policy relevant information. In May 2024, participants in a workshop during the 7^th Environmental Dimension of Antimicrobial Resistance (EDAR7) conference (Montréal, Canada) sought to answer four questions aimed at advancing the policy relevance of AMR data generated by WBS.

The effect of metals on freshwater microbiomes is poorly understood compared to other factors, such as nutrients or climate. While deleterious effects of metals on plant and animal biodiversity are well documented, the role of metals in shaping the biodiversity, composition and functional potential of sediment microbial communities remains unknown. Therefore, we explored if metal concentrations can be linked to alterations in biodiversity and composition of freshwater sediment microbial communities. We collected sediments from 34 streams and lakes in Switzerland and grouped them based on their metal content. Microbial diversity and community composition were determined using 16S rRNA gene amplicon sequencing. Most of the sediments were not contaminated with metals according to Sediment Environmental Quality Criteria, although some stations exceeded the limits for Cu, Zn, and Pb. Nevertheless, correlational analysis indicated links of metal concentrations to various aspects of sediment microbial biodiversity. Al concentrations were significantly (p < 0.05) correlated with microbial richness. We further observed a predominantly negative correlation between some metals and abundances of dominant taxa. Predicted microbial functional potential analysis indicated that different types of metals have different effects on microbial functional potential. For example, Mn exhibited a significant positive correlation with nitrogen fixation potential, whereas Cu, Pb, and Zn displayed a significant negative correlation. Overall, our findings indicate that metal concentrations may alter microbial community diversity and functional potential in freshwater sediments even at ambient concentrations. Further research into the role of metals as drivers of microbial biodiversity and factors in biodiversity loss is warranted.

Spatial environmental heterogeneity is an important driver of aquatic biodiversity. Ecological and evolutionary theory often consider spatial heterogeneity as being driven by exogenous factors, yet heterogeneity can also be generated and modified by organisms. Here we used a mesocosm experiment to investigate if consumers influence the build-up of spatial heterogeneity. We expected that consumer effects on heterogeneity would depend on consumer composition and differ among response variables. We constructed metacommunities consisting of three mesocosms and manipulated the presence and composition of consumers, using four treatment levels: (1) no consumers, (2) two genotypes of Daphnia galeata, (3) D. galeata and Daphnia longispina, and (4) D. galeata and a hybrid of D. galeata × D. longispina. We then continuously increased heterogeneity among the three patches of each metacommunity by adding nutrients and dissolved organic carbon (DOC), respectively, to two of the three mesocosms. We found that consumers affected the build-up of heterogeneity, but the direction and magnitude of this effect differed among consumer compositions. Metacommunities with only D. galeata had increased heterogeneity in phytoplankton biomass, whereas metacommunities with D. longispina or the hybrid had low phytoplankton heterogeneity. The differential effects of Daphnia taxa on phytoplankton heterogeneity cascaded down to the abiotic environment and resulted in taxon-specific effects on heterogeneity in light extinction, dissolved inorganic nitrogen, and total inorganic carbon. Our results imply that changes in consumer species (e.g., due to environmental change or invasion) might affect not only the local environment but could also impact heterogeneity among environments, with important consequences for aquatic biodiversity.

Nearly half (46%) the world’s population is now served by non-sewered sanitation. In urban areas of low- and middle-income countries, this translates to onsite storage of wastewater in tanks and pits until it can be collected and transported by road to treatment, which is commonly referred to as fecal sludge management. The microbial communities that develop during storage of this wastewater remain understudied, leaving practitioners and scientists to speculate on best management practices such as downstream treatment and climate mitigation measures. In this study, we collected samples from 135 randomly selected containments across the city of Lusaka, Zambia, and evaluated statistical relations of 16S rRNA gene sequence data to types and volume of wastewater going into containments, disturbances (i.e., emptying events), characteristics of accumulated wastewater during storage, and metrics of downstream treatment processes. At the phyla level, 80% of the identified microorganisms belonged to Firmicutes, Proteobacteria and Bacteroidota. Focusing in at the genera level, microbial diversity and composition were statistically related to volumes of water usage, properties of wastewater in containments (total organic carbon, total kjeldahl nitrogen, ammonium nitrogen, pH), and metrics of stabilization and dewatering performance. In Lusaka, a core community was identified with 104 of the 1,247 identified genera being present in >90% of the containments. In contrast, 936 genera were present in <60% of the containments, indicating that niche or transient organisms may also be important in unravelling metabolic processes such as sulfur reduction, methanogenesis, and ammonia tolerance. Community similarity was independent of time since last emptied, indicating stability of microbial communities over time. Identified metabolic differences between pit latrines (i.e., less water usage) and septic tanks (i.e., more water usage) indicate that methanogens more actively convert organic matter to methane in the more dilute wastewater in septic tanks, which could be globally relevant for greenhouse gas mitigation from non-sewered sanitation.

This report summarizes a pilot study conducted by the Eionet Working Group (WG) on antimicrobial resistance (AMR) monitoring in European surface waters. With AMR recognized as a major public health threat, the study aimed to develop a harmonized approach for collecting and reporting AMR data in surface waters, supporting revisions of EU water directives.
Led by the European Environment Agency (EEA) and the European Topic Centre Biodiversity and Ecosystems (ETC-BE), the WG developed methodologies and practical guidelines covering sampling, analysis, data reporting, and quality control. Sampling focused on rivers downstream of urban wastewater treatment plants (WWTPs). Target indicators included six gene markers (e.g., 16S rRNA, intl1, aadA1) via qPCR and culturing of E. coli and ESBL-producing E. coli (ESBL-Ec).
Ten of fourteen nominated countries participated, using qPCR as the main method for its cost-effectiveness. WHO’s Tricycle protocol was applied for culturing due to the absence of standardized EU methods for water. A custom data reporting template aligned with EEA’s WISE-6 model was also developed.
Results showed AMR gene abundance followed a clear gradient: WWTP inlet > outlet > downstream > upstream, consistent with previous studies. Gene removal by WWTPs ranged between 94.5%-99.9%, and ESBL-Ec levels varied between countries (0–14.5%). Wastewater treatment effectively reduced both E. coli and ESBL-Ec by over 99%.
The pilot faced challenges including funding limits, varied national capacities, and differing technical expertise. Despite this, it was viewed as a successful step toward EU-wide AMR monitoring. Priority areas for future EU-wide monitoring were identified and include defining objectives, harmonizing methods, improving QA/QC, and establishing a centralized reporting system.

Background Environmental reservoirs of antibiotic resistance pose a threat to human and animal health. Aquatic biofilms impacted by wastewater effluent (WW) are known environmental reservoirs for antibiotic resistance; however, the relative importance of biotic factors and abiotic factors from WW on the abundance of antibiotic resistance genes (ARGs) within aquatic biofilms remains unclear. Additionally, experimental evidence is limited within complex aquatic microbial communities as to whether genes bearing low sequence similarity to validated reference ARGs are functional as ARGs.

Results To disentangle the effects of abiotic and biotic factors on ARG abundances, natural biofilms were previously grown in flume systems with different proportions of stream water and either ultrafiltered or non-ultrafiltered WW. In this study, we conducted deep shotgun metagenomic sequencing of 75 biofilm, stream, and WW samples from these flume systems and compared the taxonomic and functional microbiome and resistome composition. Statistical analysis revealed an alignment of the resistome and microbiome composition and a significant association with experimental treatment. Several ARG classes exhibited an increase in normalized metagenomic abundances in biofilms grown with increasing percentages of non-ultrafiltered WW. In contrast, sulfonamide and extended-spectrum beta-lactamase ARGs showed greater abundances in biofilms grown in ultrafiltered WW compared to non-ultrafiltered WW. Overall, our results pointed toward the dominance of biotic factors over abiotic factors in determining ARG abundances in WW-impacted stream biofilms and suggested gene family-specific mechanisms for ARGs that exhibited divergent abundance patterns. To investigate one of these specific ARG families experimentally, we biochemically characterized a new beta-lactamase from the Planctomycetota (Phycisphaeraceae). This beta-lactamase displayed activity in the cleavage of cephalosporin analog despite sharing a low sequence identity with known ARGs.

Conclusions This discovery of a functional planctomycete beta-lactamase ARG is noteworthy, not only because it was the first beta-lactamase to be biochemically characterized from this phylum, but also because it was not detected by standard homology-based ARG tools. In summary, this study conducted a metagenomic analysis of the relative importance of biotic and abiotic factors in the context of WW discharge and their impact on both known and new ARGs in aquatic biofilms.

The global nitrogen (N) cycle has been strongly altered by anthropogenic activities, including increased input of bioavailable N into aquatic ecosystems. Freshwater sediments are hotspots with regards to the turnover and elimination of fixed N, yet the environmental controls on the microbial pathways involved in benthic N removal are not fully understood. Here, we analyze the abundance and expression of microbial genes involved in N transformations using metagenomics and -transcriptomics across sediments of 12 Swiss lakes that differ in sedimentation rates and trophic regimes. Our results indicate that microbial N loss in these sediments is primarily driven by nitrification coupled to denitrification. N-transformation gene compositions indicated three groups of lakes: agriculture-influenced lakes characterized by rapid depletion of oxidants in the sediment porewater, pristine-alpine lakes with relatively deep sedimentary penetration of oxygen and nitrate, and large, deep lakes with intermediate porewater hydrochemical properties. Sedimentary organic matter (OM) characteristics showed the strongest correlations with the community structure of microbial N-cycling communities. Most transformation pathways were expressed, but expression deviated from gene abundance and did not correlate with benthic geochemistry. Cryptic N-cycling may maintain transcriptional activity even when substrate levels are below detection. Sediments of large, deep lakes generally showed lower in-situ N gene expression than agriculture-influenced lakes, and half of the pristine-alpine lakes. This implies that prolonged OM mineralization in the water column can lead to the suppression of benthic N gene expression.

In this study, we investigated the temporal and spatial quantitative changes in the concentration of antibiotic resistance gene (ARG) markers in a municipal wastewater treatment plant (WWTP). Four ARGs conferring resistance to different classes of antibiotics (ermB, sul1, tet[W], and bla_CTXM) and a gene used as a proxy for ARG pollution (intl1) were quantified in two separate sampling campaigns covering two and half years of operation of the WWTP. First, a systematic monthly monitoring of multiple points in the inlet and the outlet revealed an absolute decrease in the concentration of all analyzed ARGs. However, the relative abundance of sul1 and intl1 genes relative to the total bacterial load (estimated using the universal marker 16S rDNA) increased in the outlet samples as compared to the inlet. To pinpoint the exact stage of removal and/or enrichment within the WWTP, a second sampling including the stages of the biological treatment was performed bimonthly. This revealed a distinct enrichment of sul1 and intl1 genes during the biological treatment phase. Moreover, the temporal and spatial variations in ARG abundance patterns within the WWTP underscored the complexity of the dynamics associated with the removal of ARGs during wastewater treatment. Understanding these dynamics is pivotal for developing efficient strategies to mitigate the dissemination of ARGs in aquatic environments. Practitioner points: Regular monitoring of ARG markers in WWTPs is essential to assess temporal and spatial changes, aiding in the development of effective mitigation strategies. Understanding the dynamics of ARG abundance during biological treatment is crucial for optimizing processes and minimizing dissemination in aquatic environments. Increased relative abundance of certain ARGs highlights potential enrichment during wastewater treatment, necessitating targeted interventions. Systematic monitoring of multiple points within WWTPs can provide valuable insights into the efficacy of treatment processes in reducing ARG levels over time. The complexity of ARG abundance patterns underscores the need to develop holistic approaches to tackle antibiotic resistance in wastewater systems.

It has been debated whether wastewater treatment plants (WWTPs) primarily act to attenuate or amplify antibiotic resistance genes (ARGs). However, ARGs are highly diverse with respect to their resistance mechanisms, mobilities, and taxonomic hosts and therefore their behavior in WWTPs should not be expected to be universally conserved. We applied metagenomic sequencing to wastewater influent and effluent samples from 12 international WWTPs to classify the behavior of specific ARGs entering and exiting WWTPs. In total, 1079 different ARGs originating from a variety of bacteria were detected. This included ARGs that could be mapped to assembled scaffolds corresponding to nine human pathogens. While the relative abundance (per 16S rRNA gene) of ARGs decreased during treatment at 11 of the 12 WWTPs sampled and absolute abundance (per mL) decreased at all 12 WWTPs, increases in relative abundance were observed for 40% of the ARGs detected at the 12th WWTP. Also, the relative abundance of mobile genetic elements (MGE) increased during treatment, but the fraction of ARGs known to be transmissible between species decreased, thus demonstrating that increased MGE prevalence may not be generally indicative of an increase in ARGs. A distinct conserved resistome was documented in both influent and effluent across samples, suggesting that well-functioning WWTPs generally attenuate influent antibiotic resistance loads. This work helps inform strategies for wastewater surveillance of antibiotic resistance, highlighting the utility of tracking ARGs as indicators of treatment performance and relative risk reduction.

Fresh produce is suggested to contribute highly to shaping the gut resistome. We investigated the impact of pig manure and irrigation water quality on microbiome and resistome of field-grown lettuce over an entire growth period. Lettuce was grown under four regimes, combining soil amendment with manure (with/without) with sprinkler irrigation using river water with an upstream wastewater input, disinfected by UV (with/without). Lettuce leaves, soil, and water samples were collected weekly and analyzed by bacterial cultivation, 16S rRNA gene amplicon sequencing, and shotgun metagenomics from total community DNA. Cultivation yielded only few clinically relevant antibiotic-resistant bacteria (ARB), but numbers of ARB on lettuce increased over time, while no treatment-dependent changes were observed. Microbiome analysis confirmed a temporal trend. Antibiotic resistance genes (ARGs) unique to lettuce and water included multidrug and β-lactam ARGs, whereas lettuce and soil uniquely shared mainly glycopeptide and tetracycline ARGs. Surface water carried clinically relevant ARB (e.g. ESBL-producing Escherichia coli or Serratia fonticola) without affecting the overall lettuce resistome significantly. Resistance markers including biocide and metal resistance were increased in lettuce grown with manure, especially young lettuce (increased soil contact). Overall, while all investigated environments had their share as sources of the lettuce resistome, manure was the main source especially on young plants. We therefore suggest minimizing soil-vegetable contact to minimize resistance markers on fresh produce.

Microbial biodiversity is fundamental to maintain ecosystem functioning in seasonally variable ecosystems. However, it remains unclear how alterations in water availability caused by episodic drying compromise the ability of stream microbes to maintain multiple functions simultaneously (e.g., primary production and carbon cycling). Using data from 32 streams, we investigated how the phenology of annual drying influences stream sediment microbial biodiversity and their capacity to sustain multifunctionality. Our results showed that stream multifunctionality and most bacteria did not respond to changes in drying phenology. Only two bacterial groups, the drying-resistant Sphingobacteriia and the drying-sensitive Acidobacteria_Gp7, exhibited positive associations with multifunctionality; whereas, bacterial diversity showed a negative correlation with functions. Among these biodiversity aspects, Sphingobacteriia showed the strongest capacity to maintain multifunctionality at low and moderate performance levels. Our findings will help to better understand the mechanisms through which biodiversity sustains the functioning of seasonally variable streams and their responses to global change.

Nitrit ist eine ökotoxikologisch problematische Substanz und führt auf Kläranlagen zur Bildung von klimaschädlichem Lachgas. Auf nitrifizierenden Belebtschlammanlagen kann Nitrit saisonal akkumulieren und erhöhte Ablaufwerte treten auf. Hier diskutieren wir die Häufigkeit von Überschreitungen des Nitrit-Richtwerts auf Schweizer ARA und schlagen Gegenmassnahmen vor.

When antimicrobial resistant bacteria (ARB) and genes (ARGs) reach novel habitats, they can become part of the habitat’s microbiome in the long term if they are able to overcome the habitat's biotic resilience towards immigration. This process should become more difficult with increasing biodiversity, as exploitable niches in a given habitat are reduced for immigrants when more diverse competitors are present. Consequently, microbial diversity could provide a natural barrier towards antimicrobial resistance by reducing the persistence time of immigrating ARB and ARG. To test this hypothesis, a pan-European sampling campaign was performed for structured forest soil and dynamic riverbed environments of low anthropogenic impact. In soils, higher diversity, evenness and richness were significantly negatively correlated with relative abundance of >85% of ARGs. Furthermore, the number of detected ARGs per sample were inversely correlated with diversity. However, no such effects were present in the more dynamic riverbeds. Hence, microbiome diversity can serve as a barrier towards antimicrobial resistance dissemination in stationary, structured environments, where long-term, diversity-based resilience against immigration can evolve.

The factors that govern the geographical distribution of nitrogen fixation are fundamental to providing accurate nitrogen budgets in aquatic environments. Model-based insights have demonstrated that regional hydrodynamics strongly impact nitrogen fixation. However, the mechanisms establishing this physical-biological coupling have yet to be constrained in field surveys. Here, we examine the distribution of nitrogen fixation in Lake Tanganyika – a model system with well-defined hydrodynamic regimes. We report that nitrogen fixation is five times higher under stratified than under upwelling conditions. Under stratified conditions, the limited resupply of inorganic nitrogen to surface waters, combined with greater light penetration, promotes the activity of bloom-forming photoautotrophic diazotrophs. In contrast, upwelling conditions support predominantly heterotrophic diazotrophs, which are uniquely suited to chemotactic foraging in a more dynamic nutrient landscape. We suggest that these hydrodynamic regimes (stratification versus mixing) play an important role in governing both the rates and the mode of nitrogen fixation.

As motivation to address environmental dissemination of antimicrobial resistance (AMR) is mounting, there is a need to characterize mechanisms by which AMR can propagate under environmental conditions. Here we investigated the effect of temperature and stagnation on the persistence of wastewater-associated antibiotic resistance markers in riverine biofilms and the invasion success of genetically-tagged Escherichia coli. Biofilms grown on glass slides incubated in-situ downstream of a wastewater treatment plant effluent discharge point were transferred to laboratory-scale flumes fed with filtered river water under potentially stressful temperature and flow conditions: recirculation flow at 20 °C, stagnation at 20 °C, and stagnation at 30 °C. After 14 days, quantitative PCR and amplicon sequencing were used to quantify bacteria, biofilms diversity, resistance markers (sul1, sul2, ermB, tetW, tetM, tetB, blaCTX-M-1, intI1) and E. coli. Resistance markers significantly decreased over time regardless of the treatment applied. Although invading E. coli were initially able to colonize the biofilms, its abundance subsequently declined. Stagnation was associated with a shift in biofilm taxonomic composition, but there was no apparent effect of flow conditions or the simulated river-pool warming (30 °C) on AMR persistence or invasion success of E. coli. Results however indicated that antibiotic resistance markers in the riverine biofilms decreased under the experimental conditions in the absence of exposure to external inputs of antibiotics and AMR.

Real-time quantitative PCR (qPCR) has been widely used to quantify gene copy numbers in microbial ecology. Despite its simplicity and straightforwardness, establishing qPCR assays is often impeded by the tedious process of producing qPCR standards by cloning the target DNA into plasmids. Here, we designed double-stranded synthetic DNA fragments from consensus sequences as qPCR standards by aligning microbial gene sequences (10–20 sequences per gene). Efficiency of standards from synthetic DNA was compared with plasmid standards by qPCR assays for different phylogenetic marker and functional genes involved in carbon (C) and nitrogen (N) cycling, tested with DNA extracted from a broad range of soils. Results showed that qPCR standard curves using synthetic DNA performed equally well to those from plasmids for all the genes tested. Furthermore, gene copy numbers from DNA extracted from soils obtained by using synthetic standards or plasmid standards were comparable. Our approach therefore demonstrates that a synthetic DNA fragment as qPCR standard provides comparable sensitivity and reliability to a traditional plasmid standard, while being more time- and cost-efficient.

Wastewater treatment plants (WWTPs) are key sources of antimicrobial resistance genes (ARGs) that could influence the resistomes of microbial communities in various habitats of the receiving river ecosystem. However, it is currently unknown which habitats are most impacted and whether ARGs, like certain chemical contaminants, could be accumulated or enriched in the river ecosystem. We conducted a systematic metagenomic survey on the antibiotic resistomes of WWTP effluent, four riverine habitats (water, suspended particles, sediment, epilithic biofilm), and freshwater amphipod gut microbiomes. The impact of WWTP effluent on the downstream habitats was assessed in nine Swiss rivers. While there were significant differences in resistomes across habitats, the wastewater resistome was more similar to the resistome of receiving river water than to the resistomes of other habitats, and river water was the habitat most strongly impacted by the WWTPs effluent. The sulfonamide, beta-lactam, and aminoglycoside resistance genes were among the most abundant ARGs in the WWTP effluents, and especially aadA, sul1, and class A beta-lactamase genes showed significantly increased abundance in the river water of downstream compared to upstream locations (p < 0.05). However, this was not the case for the sediment, biofilm, and amphipod gut habitats. Accordingly, evidence for accumulation or enrichment of ARGs through the riverine food web was not identified. Our study suggests that monitoring riverine antimicrobial resistance determinants could be conducted using "co-occurrence" of aadA, sul1, and class A beta-lactamase genes as an indicator of wastewater-related pollution and should focus on the water as the most affected habitat.

Ein zeitnahes Biomonitoring mit Nanopore-Sequenzierung gewährt neue Einblicke in die biologischen Prozesse einer Abwasserreinigungsanlage (ARA). Im Rahmen des Projekts «MicROcensus» wurde an verschiedenen ARA die Einsatzfähigkeit solcher Mikrobiom-Analysen als Diagnoseinstrument evaluiert. So unterstützen Momentaufnahmemessungen die Ursachenfindung von Prozessinstabilitäten oder Begleitphänomenen. Während wöchentliche Analysen des Mikrobioms – bei Bekanntsein der jahreszeitlichen Dynamiken - als Frühwarnsystem dienen und praxisrelevante Daten für den ARA-Betrieb liefern können.

Surveillance of antibiotic resistance genes (ARGs) has been increasingly conducted in environmental sectors to complement the surveys in human and animal sectors under the "One-Health" framework. However, there are substantial challenges in comparing and synthesizing the results of multiple studies that employ different test methods and approaches in bioinformatic analysis. In this article, we consider the commonly used quantification units (ARG copy per cell, ARG copy per genome, ARG density, ARG copy per 16S rRNA gene, RPKM, coverage, PPM, etc.) for profiling ARGs and suggest a universal unit (ARG copy per cell) for reporting such biological measurements of samples and improving the comparability of different surveillance efforts.

The nitrogen (N) cycle is of global importance, as N is an essential element and a limiting nutrient in terrestrial and aquatic ecosystems. Excessive anthropogenic N fertilizer usage threatens sensitive downstream aquatic ecosystems. Although freshwater lake sediments remove N through various microbially mediated processes, few studies have investigated the microbial communities involved. In an integrated biogeochemical and microbiological study on a eutrophic and oligotrophic lake, we estimated N removal rates from pore water concentration gradients in sediments. Simultaneously, the abundance of different microbial N transformation genes was investigated using metagenomics on a seasonal and spatial scale. We observed that contrasting nutrient concentrations in sediments were associated with distinct microbial community compositions and significant differences in abundances of various N transformation genes. For both characteristics, we observed a more pronounced spatial than seasonal variability within each lake. The eutrophic Lake Baldegg showed a higher denitrification potential with higher nosZ gene (N₂O reductase) abundances and higher nirS:nirK (nitrite reductase) ratios, indicating a greater capacity for complete denitrification. Correspondingly, this lake had a higher N removal efficiency. The oligotrophic Lake Sarnen, in contrast, had a higher potential for nitrification. Specifically, it harbored a high abundance of Nitrospira, including some with the potential for comammox. Our results demonstrate that knowledge of the genomic N transformation potential is important for interpreting N process rates and understanding how the lacustrine sedimentary N cycle responds to variations in trophic conditions.

Zu den kommenden Herausforderungen für die Überwachung der mikrobiologischen Qualität des Trinkwassers gehört die Bewertung möglicher Risiken durch antibiotikaresistente Bakterien. Analysemethoden sind aufwendig und teuer. Lässt sich das Vorkommen von Antibiotikaresistenzen mit bereits etablierten Methoden der Trinkwasserüberwachung, wie der Durchflusszytometrie, verknüpfen? Dieser Frage gingen Forscher der Eawag nach.

New findings enable concrete measures at individual interfaces of AMR spread

The aim of the synthesis process on this topic was to derive recommendations from NRP 72 research that promote the implementation of new findings in practice. The focus of many projects was on the interfaces where antimicrobial resistance (AMR) can spread between humans, animals and the environment. In this One Health context, many research findings of NRP 72 provide the basis for concrete measures to interrupt or restrict transmission chains.
In addition to these concrete findings, it has also become apparent that the methods used in NRP 72 research are of great importance: It is a common feature of the projects presented in this thematic synthesis that they have applied new gene sequencing methods, such as whole genome sequencing (WGS), plasmid sequencing and metagenomics. These methods have developed very quickly in the last few years and are a prerequisite for the new insights presented here. [...]

Background: There is concern that the microbially rich activated sludge environment of wastewater treatment plants (WWTPs) may contribute to the dissemination of antibiotic resistance genes (ARGs). We applied long-read (nanopore) sequencing to profile ARGs and their neighboring genes to illuminate their fate in the activated sludge treatment by comparing their abundance, genetic locations, mobility potential, and bacterial hosts within activated sludge relative to those in influent sewage across five WWTPs from three continents.
Results: The abundances (gene copies per Gb of reads, aka gc/Gb) of all ARGs and those carried by putative pathogens decreased 75-90% from influent sewage (192-605 gc/Gb) to activated sludge (31-62 gc/Gb) at all five WWTPs. Long reads enabled quantification of the percent abundance of ARGs with mobility potential (i.e., located on plasmids or co-located with other mobile genetic elements (MGEs)). The abundance of plasmid-associated ARGs decreased at four of five WWTPs (from 40-73 to 31–68%), and ARGs co-located with transposable, integrative, and conjugative element hallmark genes showed similar trends. Most ARG-associated elements decreased 0.35-13.52% while integrative and transposable elements displayed slight increases at two WWTPs (1.4-2.4%). While resistome and taxonomic compositions both shifted significantly, host phyla for chromosomal ARG classes remained relatively consistent, indicating vertical gene transfer via active biomass growth in activated sludge as the key pathway of chromosomal ARG dissemination.
Conclusions: Overall, our results suggest that the activated sludge process acted as a barrier against the proliferation of most ARGs, while those that persisted or increased warrant further attention.

Background: During wastewater treatment, the wastewater microbiome facilitates the degradation of organic matter, reduction of nutrients, and removal of gut parasites. While the latter function is essential to minimize public health risks, the range of parasites involved and how they are removed is still poorly understood.
Results: Using shotgun metagenomic (DNA) and metatranscriptomic (RNA) sequencing data from ten wastewater treatment plants in Switzerland, we were able to assess the entire wastewater microbiome, including the often neglected microeukaryotes (protists). In the latter group, we found a surprising richness and relative abundance of active parasites, particularly in the inflow. Using network analysis, we tracked these taxa across the various treatment compartments and linked their removal to trophic interactions.
Conclusions: Our results indicate that the combination of DNA and RNA data is essential for assessing the full spectrum of taxa present in wastewater. In particular, we shed light on an important but poorly understood function of wastewater treatment - parasite removal.

Lachgas (N₂O) ist ein starkes Treibhausgas und die wichtigste ozonzerstörende Substanz in der Stratosphäre. Auf Abwasserreinigungsanlagen wird N₂O während des biologischen Stickstoffabbaus gebildet, der somit die grösste Treibhausgasquelle während des Reinigungsprozesses darstellt. Reduktionsmassnahmen sind deshalb von grosser Bedeutung und gemäss neuen Erkenntnissen durch eine Optimierung der Nitrifikation und Denitrifikation auch erreichbar.

The development of new wastewater treatment processes can assist in reducing the impact of wastewater treatment on the environment. The recently developed partial nitritation anammox (PNA) process, for example, consumes less energy for aeration and reduces nitrate in the effluent without requiring additional organic carbon. However, achieving stable nitritation (ammonium oxidation; NH₄⁺ → NO₂⁻) at mainstream conditions (T = 10–25 °C, C:N > 10, influent ammonium < 50 mgNH₄-N/L and effluent < 1 mgNH₄-N/L) remains challenging. This study explores the potential and mechanism of nitrite-oxidizing bacteria (NOB) suppression in a bottom-fed sequencing batch reactor (SBR). Two bench-scale (11 L) reactors and a pilot-scale reactor (8 m³) were operated for over a year and were fed with organic substrate depleted municipal wastewater. Initially, nitratation (nitrite oxidation; NO₂⁻ → NO₃⁻) occurred occasionally until an anaerobic phase was integrated into the operating cycle. The introduction of the anaerobic phase effectively suppressed the regrowth of NOB while nitritation was stable over 300 days, down to 8 °C and at ammonium influent concentrations < 25 mgNH₄-N/L. Batch experiments and process data revealed that parameters typically affecting NOB growth (e.g., dissolved oxygen, alkalinity, trace elements, lag-phase after anoxia, free nitrous acid (FNA), free ammonia (FA), pH, sulfide, or solids retention time (SRT)) could not fully explain the suppression of nitratation. Experiments in which fresh nitrifying microbial biomass was added to the nitritation system indicated that NOB inactivation explained NOB suppression better than NOB washout at high SRT. This study concludes that bottom-fed SBRs with anaerobic phases allow for stable nitritation over a broad range of operational parameters. Coupling this type of SBR to an anammox reactor can enable efficient mainstream anammox-based wastewater treatment.

For two decades now, partial nitritation anammox (PNA) systems were suggested to more efficiently remove nitrogen (N) from mainstream municipal wastewater. Yet to date, only a few pilot-scale systems and even fewer full-scale implementations of this technology have been described. Process instability continues to restrict the broad application of PNA. Especially problematic are insufficient anammox biomass retention, the growth of undesired aerobic nitrite-oxidizers, and nitrous oxide (N₂O) emissions. In this study, a two-stage mainstream pilot-scale PNA system, consisting of three reactors (carbon pre-treatment, nitritation, anammox - 8 m³ each), was operated over a year, treating municipal wastewater. The aim was to test whether both, robust autotrophic N removal and high effluent quality, can be achieved throughout the year. A second aim was to better understand rate limiting processes, potentially affecting the overall performance of PNA systems. In this pilot study, excellent effluent quality, in terms of inorganic nitrogen, was accomplished (average effluent concentrations: 0.4 mgNH₄-N/L, 0.1 mgNO₂-N/L, 0.9 mgNO₃-N/L) even at wastewater temperatures previously considered problematic (as low as 8 °C). N removal was limited by nitritation rates (84 ± 43 mgNH₄-N/L/d), while surplus anammox activity was observed at all times (178 ± 43 mgN/L/d). Throughout the study, nitrite-oxidation was maintained at a low level (<2.5% of ammonium consumption rate). Unfortunately, high N₂O emissions from the nitritation stage (1.2% of total nitrogen in the influent) were observed, and, based on natural isotope abundance measurements, could be attributed to heterotrophic denitrification. In situ batch experiments were conducted to identify the role of dissolved oxygen (DO) and organic substrate availability in N₂O emission-mitigation. The addition of organic substrate, to promote complete denitrification, was not successful in decreasing N₂O emission, but increasing the DO from 0.3 to 2.9 mgO₂/L decreased N₂O emissions by a factor of 3.4.

Untreated combined sewage (bypass) is often discharged by wastewater treatment plants to receiving rivers during stormwater events, where it may contribute to increased levels of antibiotic resistance genes (ARGs) and multi-resistance risk factors (multi-resistant bacteria and multi-resistance genomic determinants (MGDs)) in the receiving water. Other contamination sources, such as soil runoff and resuspended river sediment could also play a role during stormwater events. Here we report on stormwater event-based sampling campaigns to determine temporal dynamics of ARGs and multi-resistance risk factors in bypass, treated effluent, and the receiving river, as well as complimentary data on catchment soils and surface sediments. Both indicator ARGs (qPCR) and resistome (ARG profiles revealed by metagenomics) indicated bypass as the main contributor to the increased levels of ARGs in the river during stormwater events. Furthermore, we showed for the first time that the risk of exposure to bypass-borne multi-resistance risk factors increase under stormwater events and that many of these MGDs were plasmid associated and thus potentially mobile. In addition, elevated resistance risk factors persisted for some time (up to 22 h) in the receiving water after stormwater events, likely due to inputs from distributed overflows in the catchment. This indicates temporal dynamics should be considered when interpreting the risks of exposure to resistance from event-based contamination. We propose that reducing bypass from wastewater treatment plants may be an important intervention option for reducing dissemination of antibiotic resistance.

Wastewater-based surveillance (WBS) for disease monitoring is highly promising but requires consistent methodologies that incorporate predetermined objectives, targets, and metrics. Herein, we describe a comprehensive metagenomics-based approach for global surveillance of antibiotic resistance in sewage that enables assessment of 1) which antibiotic resistance genes (ARGs) are shared across regions/communities; 2) which ARGs are discriminatory; and 3) factors associated with overall trends in ARGs, such as antibiotic concentrations. Across an internationally sourced transect of sewage samples collected using a centralized, standardized protocol, ARG relative abundances (16S rRNA gene-normalized) were highest in Hong Kong and India and lowest in Sweden and Switzerland, reflecting national policy, measured antibiotic concentrations, and metal resistance genes. Asian versus European/US resistomes were distinct, with macrolide-lincosamide-streptogramin, phenicol, quinolone, and tetracycline versus multidrug resistance ARGs being discriminatory, respectively. Regional trends in measured antibiotic concentrations differed from trends expected from public sales data. This could reflect unaccounted uses, captured only by the WBS approach. If properly benchmarked, antibiotic WBS might complement public sales and consumption statistics in the future. The WBS approach defined herein demonstrates multisite comparability and sensitivity to local/regional factors.

The nitrogen (N) cycle is of global importance as N is an essential element and limiting nutrient in terrestrial and aquatic ecosystems. On a global scale, N is transported from terrestrial ecosystems through runoff into surface waters and further into the ocean. Excessive anthropogenic N fertilizer usage threatens sensitive downstream aquatic ecosystems and can lead to eutrophication in surface waters, especially in N-limited coastal ecosystems. Lakes effectively reduce the reactive N load by transforming it into inert N2 gas through microbial denitrification or anammox, thus acting as N sinks. All N transforming processes are mediated by microbes, and regulated by various environmental parameters. Among those, oxygen (O2) is one of the main controls. Thus, the classification of aerobic oxidation processes, such as nitrification, and anaerobic reduction processes, such as denitrification, anammox, or dissimilatory nitrate (NO_3^-) reduction to ammonia (DNRA) are commonly applied. Further environmental controls on N removal rates in lakes are organic matter (OM) composition, external N load, NO_3^- concentrations, and the productivity (trophic state) of a lake. Furthermore, the N cycle is tightly linked with the carbon and phosphorus cycles, can be coupled to the sulfur cycle, and shows a complex internal connection of the various N transforming processes. Even though the research on N removal and cycle in lakes has been emphasized, the environmental controls on the microbial communities involved in the turnover and elimination of reactive N in lakes are not fully understood. [...]

In stratified lakes, methane oxidizing bacteria are critical methane converters that significantly reduce emissions of this greenhouse gas to the atmosphere. Efforts to better understand their ecology uncovered a surprising diversity, vertical structure, and seasonal succession. It is an open question how this diversity has to be considered in models of microbial methane oxidation. Likewise, it is unclear to what extent simple microbial traits related to the kinetics of the oxidation process and temperature optimum, suggested by previous studies, suffice to understand the observed ecology of methane oxidizing bacteria. Here we incorporate niche partitioning in a mechanistic model of seasonal lake mixing and microbial methane oxidation in a stratified lake. Can we model MOB diversity and niche partitioning based on differences in methane oxidation kinetics and temperature adaptation? We found that our model approach can closely reproduce diversity and niche preference patterns of methanotrophs that were observed in seasonally stratified lakes. We show that the combination of trait values resulting in coexisting methanotroph communities is limited to very confined regions within the parameter space of potential trait combinations. However, our model also indicates that the sequence of community assembly, and variations in the stratification and mixing behavior of the lake result in different stable combinations. A scenario analysis introducing variable mixing conditions showed that annual weather conditions and the pre-existing species also affect the developing stable methanotrophic species composition of the lake. Both, effect of pre-existing species and the environmental impact suggest that the MOB community in lakes may differ from year to year, and a stable community may never truly occur. The model further shows that there are always better-adapted species in the trait parameter space that would destabilize and replace an existing stable community. Thus, natural selection may drive trait values into the specific configurations observed in nature based on physiological limits and tradeoffs between traits.

Nitrous oxide (N₂O) is a strong greenhouse gas and causal for stratospheric ozone depletion. During biological nitrogen removal in wastewater treatment plants (WWTP), high N₂O fluxes to the atmosphere can occur, typically exhibiting a seasonal emission pattern. Attempts to explain the peak emission phases in winter and spring using physico-chemical process data from WWTP were so far unsuccessful and new approaches are required. The complex and diverse microbial community of activated sludge used in biological treatment systems also exhibit substantial seasonal patterns. However, a potentially causal link between the seasonal patterns of microbial diversity and N₂O emissions has not yet been investigated. Here we show that in a full-scale WWTP nitrification failure and N₂O peak emissions, bad settleability of the activated sludge and a turbid effluent strongly correlate with a significant reduction in the microbial community diversity and shifts in community composition. During episodes of impaired performance, we observed a significant reduction in abundance for filamentous and nitrite oxidizing bacteria in all affected reactors. In some reactors that did not exhibit nitrification and settling failures, we observed a stable microbial community and no drastic loss of species. Standard engineering approaches to stabilize nitrification, such as increasing the aerobic sludge age and oxygen availability failed to improve the plant performance on this particular WWTP and replacing the activated sludge was the only measure applied by the operators to recover treatment performance in affected reactors. Our results demonstrate that disturbances of the sludge microbiome affect key structural and functional microbial groups, which lead to seasonal N₂O emission patterns. To reduce N₂O emissions from WWTP, it is therefore crucial to understand the drivers that lead to the microbial population dynamics in the activated sludge.

Antimicrobial resistance (AMR) is a serious threat to public health as recognized by many international and governmental entities. Rivers are important routes through which anthropogenic AMR is transmitted to other environments. Various factors could affect the fate of riverine AMR, e.g., hydrogeologic processes, various sources/sinks, and weather. In this study, the interplay among those factors and their impact on riverine resistome were studied in high wastewater-impacted rivers in Switzerland. [...]

Antibiotikaresistenzen (ABR) sind ernsthafte Bedrohungen für die öffentliche Gesundheit, welche von vielen internationalen und staatlichen Stellen anerkannt sind. Flüsse sind wichtige Wege, über die anthropogene ABR in andere Ökosysteme übertragen werden können. Der Verbleib von ABR in Flüssen kann von verschiedenen Faktoren beeinflusst werden, z.B. hydrogeologische Prozesse, verschiedene Quellen/Senken und das Wetter. In dieser Studie wurde das Zusammenspiel dieser Faktoren und ihr Einfluss auf flussgebundene ABR in stark abwasserbelasteten Flüssen in der Schweiz untersucht. [...]

River networks are one of the main routes by which the public could be exposed to environmental sources of antibiotic resistance, that may be introduced e.g. via treated wastewater. In this study, we applied a comprehensive integrated analysis encompassing mass-flow concepts, chemistry, bacterial plate counts, resistance gene quantification and shotgun metagenomics to track the fate of the resistome (collective antibiotic resistance genes (ARGs) in a microbial community) of treated wastewater in two Swiss rivers at the kilometer scale. The levels of certain ARGs and the class 1 integron integrase gene (intI1) commonly associated with anthropogenic sources of ARGs decreased quickly over short distances (2-2.5 km) downstream of wastewater discharge points. Mass-flow analysis based on conservative tracers suggested this decrease was attributable mainly to dilution but ARG loadings frequently also decreased (e.g., 55.0-98.5 % for ermB and tetW) over the longest studied distances (6.8 and 13.7 km downstream). Metagenomic analysis confirmed that ARG of wastewater-origin did not persist in rivers after 5 ∼ 6.8 km downstream distance. sul1 and intI1 levels and loadings were more variable and even increased sharply at 5 ∼ 6.8 km downstream distance on one occasion. While input from agriculture and in-situ positive selection pressure for organisms carrying ARGs cannot be excluded, in-system growth of biomass is a more probable explanation. The potential for direct human exposure to the resistome of wastewater-origin thus appeared to typically abate rapidly in the studied rivers. However, the riverine aquatic resistome was also dynamic, as evidenced by the increase of certain gene markers downstream, without obvious sources of anthropogenic contamination. This study provides new insight into drivers of riverine resistomes and pinpoints key monitoring targets indicative of where human sources and exposures are likely to be most acute.

Anaerobic ammonium oxidation (anammox) plays an important role in aquatic systems as a sink of bioavailable nitrogen (N), and in engineered processes by removing ammonium from wastewater. The isotope effects anammox imparts in the N isotope signatures (¹⁵N/¹⁴N) of ammonium, nitrite, and nitrate can be used to estimate its role in environmental settings, to describe physiological and ecological variations in the anammox process, and possibly to optimize anammox-based wastewater treatment. We measured the stable N-isotope composition of ammonium, nitrite, and nitrate in wastewater cultivations of anammox bacteria. We find that the N isotope enrichment factor ¹⁵ε for the reduction of nitrite to N₂ is consistent across all experimental conditions (13.5‰ ± 3.7‰), suggesting it reflects the composition of the anammox bacteria community. Values of ¹⁵ε for the oxidation of nitrite to nitrate (inverse isotope effect, - 16 to - 43‰) and for the reduction of ammonium to N₂ (normal isotope effect, 19-32‰) are more variable, and likely controlled by experimental conditions. We argue that the variations in the isotope effects can be tied to the metabolism and physiology of anammox bacteria, and that the broad range of isotope effects observed for anammox introduces complications for analyzing N-isotope mass balances in natural systems.

Mainstream anaerobic ammonium oxidation (anammox) represents one of the most promising energy-efficient mechanisms of fixed nitrogen elimination from wastewaters. However, little is known about the exact processes and drivers of microbial community assembly within the complex microbial biofilms that support anammox in engineered ecosystems. Here, we followed anammox biofilm development on fresh carriers in an established 8m³ mainstream anammox reactor that is exposed to seasonal temperature changes (∼25-12°C) and varying NH₄⁺ concentrations (5-25 mg/L). We use fluorescence in situ hybridization and 16S rRNA gene sequencing to show that three distinct stages of biofilm development emerge naturally from microbial community composition and biofilm structure. Neutral modelling and network analysis are employed to elucidate the relative importance of stochastic versus deterministic processes and synergistic and antagonistic interactions in the biofilms during their development. We find that the different phases are characterized by a dynamic succession and an interplay of both stochastic and deterministic processes. The observed growth stages (Colonization, Succession and Maturation) appear to be the prerequisite for the anticipated growth of anammox bacteria and for reaching a biofilm community structure that supports the desired metabolic and functional capacities observed for biofilm carriers already present in the system (∼100g_NH4-N m³ d^-1). We discuss the relevance of this improved understanding of anammox-community ecology and biofilm development in the context of its practical application in the start-up, configuration, and optimization of anammox biofilm reactors.

Autotrophic nitrogen removal by anaerobic ammonium oxidizing (anammox) bacteria is an energy-efficient nitrogen removal process in wastewater treatment. However, full-scale deployment under mainstream conditions remains challenging for practitioners due to the high stress susceptibility of anammox bacteria towards fluctuations in dissolved oxygen (DO) and temperature. Here, we investigated the response of microbial biofilms with verified anammox activity to DO shocks under 20 °C and 14 °C. While pulse disturbances of 0.3 mg L⁻¹ DO prompted only moderate declines in the NH₄⁺ removal rates, 1.0 mg L⁻¹ DO led to complete but reversible inhibition of the NH₄⁺ removal activity in all reactors. Genome-centric metagenomics and metatranscriptomics were used to investigate the stress response on various biological levels. We show that temperature regime and strength of DO perturbations induced divergent responses from the process level down to the transcriptional profile of individual taxa. Community-wide gene expression differed significantly depending on the temperature regime in all reactors, and we found a noticeable impact of DO disturbances on genes involved in transcription, translation, replication and posttranslational modification at 20 °C but not 14 °C. Genome-centric analysis revealed that different anammox species and other key biofilm taxa differed in their transcriptional responses to distinct temperature regimes and DO disturbances.

The global rise and spread of antibiotic resistance greatly challenge the treatment of bacterial infections. Wastewater treatment plants (WWTPs) harbor and discharge antibiotic resistance genes (ARGs) as environmental contaminants. However, the knowledge gap on the host identity, activity, and functionality of ARGs limits transmission and health risk assessment of the WWTP resistome. Hereby, a genome-centric quantitative metatranscriptomic approach was exploited to realize high-resolution qualitative and quantitative analyses of bacterial hosts of ARGs (i.e., multiresistance, pathogenicity, activity, and niches) in the 12 urban WWTPs. We found that ∼45% of 248 recovered genomes expressed ARGs against multiple classes of antibiotics, among which bacitracin and aminoglycoside resistance genes in Proteobacteria were the most prevalent scenario. Both potential pathogens and indigenous denitrifying bacteria were transcriptionally active hosts of ARGs. The almost unchanged relative expression levels of ARGs in the most resistant populations (66.9%) and the surviving ARG hosts including globally emerging pathogens (e.g., Aliarcobacter cryaerophilus) in treated WWTP effluent prioritize future examination on the health risks related to resistance propagation and human exposure in the receiving environment.

Many seasonally stratified lakes accumulate substantial amounts of the greenhouse gas methane in the anoxic zone. Methane oxidizing bacteria in the water column act as a converter, oxidizing methane into carbon dioxide and biomass before it reaches the atmosphere. Current observations and estimates of this methane oxidation efficiency are diverging, especially for the lake overturn period. Here, we combine a model of turbulent mixing, gas exchange, and microbial growth with a comprehensive data set for autumn mixing to quantify the relevant physical and microbial processes for a 16 m deep, wind-sheltered Swiss lake. Scenario analysis suggests that the methane converter is efficient and robust under a large range of mixing velocities and only rare events of pronounced surface cooling can trigger substantial outgassing. This case study combines in situ observation and a deterministic physically based model and suggests that the frequency of storms may strongly impact methane emissions for similar temperate lakes.

Have you ever sought to use metagenomic DNA sequences reported in scientific publications? Were you successful? Here, we reveal that metagenomes from no fewer than 20% of the papers found in our literature search, published between 2016 and 2019, were not deposited in a repository or were simply inaccessible. The proportion of inaccessible data within the literature has been increasing year-on-year. Noncompliance with Open Data is best predicted by the scientific discipline of the journal. The number of citations, journal type (e.g., Open Access or subscription journals), and publisher are not good predictors of data accessibility. However, many publications in high-impact factor journals do display a higher likelihood of accessible metagenomic data sets. Twenty-first century science demands compliance with the ethical standard of data sharing of metagenomes and DNA sequence data more broadly. Data accessibility must become one of the routine and mandatory components of manuscript submissions - a requirement that should be applicable across the increasing number of disciplines using metagenomics. Compliance must be ensured and reinforced by funders, publishers, editors, reviewers, and, ultimately, the authors.

In stratified lakes, methane-oxidizing bacteria (MOB) are strongly mitigating methane fluxes to the atmosphere by consuming methane entering the water column from the sediments. MOB communities in lakes are diverse and vertically structured, but their spatio-temporal dynamics along the water column as well as physico-chemical parameters and interactions with other bacterial species that drive the community assembly have so far not been explored in depth. Here, we present a detailed investigation of the MOB and bacterial community composition and a large set of physico-chemical parameters in a shallow, seasonally stratified, and sub-alpine lake. Four highly resolved vertical profiles were sampled in three different years and during various stages of development of the stratified water column. Non-randomly assembled MOB communities were detected in all compartments. We could identify methane and oxygen gradients and physico-chemical parameters like pH, light, available copper and iron, and total dissolved nitrogen as important drivers of the MOB community structure. In addition, MOB were well-integrated into a bacterial-environmental network. Partial redundancy analysis of the relevance network of physico-chemical variables and bacteria explained up to 84% of the MOB abundances. Spatio-temporal MOB community changes were 51% congruent with shifts in the total bacterial community and 22% of variance in MOB abundances could be explained exclusively by the bacterial community composition. Our results show that microbial interactions may play an important role in structuring the MOB community along the depth gradient of stratified lakes.

The World Health Organization Global Action Plan recommends integrated surveillance programs as crucial strategies for monitoring antibiotic resistance. Although several national surveillance programs are in place for clinical and veterinary settings, no such schemes exist for monitoring antibiotic-resistant bacteria in the environment. In this transnational study, we developed, validated, and tested a low-cost surveillance and easy to implement approach to evaluate antibiotic resistance in wastewater treatment plants (WWTPs) by targeting cefotaxime-resistant (CTX-R) coliforms as indicators. The rationale for this approach was: i) coliform quantification methods are internationally accepted as indicators of fecal contamination in recreational waters and are therefore routinely applied in analytical labs; ii) CTX-R coliforms are clinically relevant, associated with extended-spectrum β-lactamases (ESBLs), and are rare in pristine environments. We analyzed 57 WWTPs in 22 countries across Europe, Asia, Africa, Australia, and North America. CTX-R coliforms were ubiquitous in raw sewage and their relative abundance varied significantly (<0.1% to 38.3%), being positively correlated (p < 0.001) with regional atmospheric temperatures. Although most WWTPs removed large proportions of CTX-R coliforms, loads over 10³ colony-forming units per mL were occasionally observed in final effluents. We demonstrate that CTX-R coliform monitoring is a feasible and affordable approach to assess wastewater antibiotic resistance status.

Lakes and reservoirs contribute substantially to atmospheric concentrations of the potent greenhouse gas methane. Lake sediments produce large amounts of methane, which accumulate in the oxygen-depleted bottom waters of stratified lakes. Climate change and eutrophication may increase the number of lakes with methane storage in the future. Whether stored methane escapes to the atmosphere during annual lake overturn is a matter of controversy and depends critically on the response of the methanotroph assemblage. Here we show, by combining 16S rRNA gene and pmoA mRNA amplicon sequencing, qPCR, CARD-FISH and potential methane-oxidation rate measurements, that the methanotroph assemblage in a mixing lake underwent both a substantial bloom and ecological succession. As a result, methane oxidation kept pace with the methane supplied from methane-rich bottom water and most methane was oxidized. This aspect of freshwater methanotroph ecology represents an effective mechanism limiting methane transfer from lakes to the atmosphere.

In lakes, large amounts of methane are produced in anoxic sediments. Methane-oxidizing bacteria effectively convert this potent greenhouse gas into biomass and carbon dioxide. These bacteria are present throughout the water column, where methane concentrations can range from nanomolar to millimolar. In this study, we tested the hypothesis that methanotroph assemblages in a seasonally stratified freshwater lake are adapted to the contrasting methane concentrations in the epi- and hypolimnion. We further hypothesized that lake overturn would change the apparent methane oxidation kinetics as more methane becomes available in the epilimnion. In addition to the change in the methane oxidation kinetics, we investigated changes in the transcription of genes encoding methane monooxygenase, the enzyme responsible for the first step of methane oxidation, with metatranscriptomics. Using laboratory incubations of the natural microbial communities, we show that the half-saturation constant (K_m) for methane - the methane concentration at which half the maximum methane oxidation rate is reached - was 20 times higher in the hypolimnion than in the epilimnion during stable stratification. During lake overturn, however, the kinetic constants in the epi- and hypolimnion converged along with a change in the transcriptionally active methanotroph assemblage. Conventional particulate methane monooxygenase appeared to be responsible for methane oxidation under different methane concentrations. Our results suggest that methane availability is one important factor for creating niches for methanotroph assemblages with well-adapted methane oxidation kinetics. This rapid selection and succession of adapted lacustrine methanotroph assemblages allowed the previously reported high removal efficiency of methane transported to the epilimnion to be maintained – even under rapidly changing conditions during lake overturn. Consequently, only a small fraction of methane stored in the anoxic hypolimnion is emitted to the atmosphere.

Lakes are a significant source of atmospheric methane, although methane-oxidizing bacteria consume most methane diffusing upward from anoxic sediments. Diverse methane-oxidizing bacteria form an effective methane filter in the water column of stratified lakes, yet, niche partitioning of different methane-oxidizing bacteria along the oxygen–methane counter gradient remains poorly understood. In our study, we reveal vertical distribution patterns of active methane-oxidizing bacteria along the oxygen–methane counter gradient of four lakes, based on amplicon sequencing analysis of 16S rRNA and pmoA genes, and 16S rRNA and pmoA transcripts, and potential methane oxidation rates. Differential distribution patterns indicated that ecologically different methane-oxidizing bacteria occupied the methane-deficient and oxygen-deficient part above and below the oxygen–methane interface. The interface sometimes harbored additional taxa. Within the dominant Methylococcales, an uncultivated taxon (CABC2E06) occurred mainly under methane-deficient conditions, whereas Crenothrix-related taxa preferred oxygen-deficient conditions. Candidatus Methylomirabilis limnetica (NC10 phylum) abundantly populated the oxygen-deficient part in two of four lakes. We reason that the methane filter in lakes is structured and that methane-oxidizing bacteria may rely on niche-specific adaptations for methane oxidation along the oxygen–methane counter gradient. Niche partitioning of methane-oxidizing bacteria might support greater overall resource consumption, contributing to the high effectivity of the lacustrine methane filter.

Aerobic methane-oxidizing bacteria (MOB) substantially reduce methane fluxes from freshwater sediments to the atmosphere. Their metalloenzyme methane monooxygenase (MMO) catalyses the first oxidation step converting methane to methanol. Its most prevalent form is the copper-dependent particulate pMMO, however, some MOB are also able to express the iron-containing, soluble sMMO under conditions of copper scarcity. So far, the link between copper availability in different forms and biological methane consumption in freshwater systems is poorly understood. Here, we present high-resolution profiles of MOB abundance and pMMO and sMMO functional genes in relation to copper, methane and oxygen profiles across the oxic-anoxic boundary of a stratified lake. We show that even at low nanomolar copper concentrations, MOB species containing the gene for pMMO expression are present at high abundance. The findings highlight the importance of copper as a micronutrient for MOB species and the potential usage of copper acquisition strategies, even under conditions of abundant iron, and shed light on the spatial distribution of these microorganisms.

Wastewater treatment plants (WWTPs) are implicated as hotspots for the dissemination of antibacterial resistance into the environment. However, the in situ processes governing removal, persistence, and evolution of resistance genes during wastewater treatment remain poorly understood. Here, we used quantitative metagenomic and metatranscriptomic approaches to achieve a broad-spectrum view of the flow and expression of genes related to antibacterial resistance to over 20 classes of antibiotics, 65 biocides, and 22 metals. All compartments of 12 WWTPs share persistent resistance genes with detectable transcriptional activities that were comparatively higher in the secondary effluent, where mobility genes also show higher relative abundance and expression ratios. The richness and abundance of resistance genes vary greatly across metagenomes from different treatment compartments, and their relative and absolute abundances correlate with bacterial community composition and biomass concentration. No strong drivers of resistome composition could be identified among the chemical stressors analyzed, although the sub-inhibitory concentration (hundreds of ng/L) of macrolide antibiotics in wastewater correlates with macrolide and vancomycin resistance genes. Contig-based analysis shows considerable co-localization between resistance and mobility genes and implies a history of substantial horizontal resistance transfer involving human bacterial pathogens. Based on these findings, we propose future inclusion of mobility incidence (M%) and host pathogenicity of antibiotic resistance genes in their quantitative health risk ranking models with an ultimate goal to assess the biological significance of wastewater resistomes with regard to disease control in humans or domestic livestock.

The fate of dissolved organic carbon (DOC) is partly determined by its availability to microbial degradation. Organisms at upper trophic levels could influence the bioavailability of DOC via cascading effects on primary producers and bacteria. Here we experimentally tested whether the presence of fish in aquatic food webs can indirectly affect the composition of the DOC pool. We found that fish had strong positive effects on phytoplankton biomass that affected the dynamics of DOC composition. Specifically, fish increased protein-like, algae-derived DOC mid-experiment, concurrent with the strongest fish-induced increase in phytoplankton biomass. Fish also increased bacterial abundance, altered the community composition and diversity of bacteria, and temporarily increased DOC compounds with fluorescence properties indicative of microbially-reprocessed organic matter. Overall, our experiment revealed that fish can positively influence the substrate (algae-produced DOC) and the key players (bacteria) of the microbial carbon pump. Consequently, fish could contribute to carbon sequestration by stimulating both the production of bioavailable DOC and the microbial degradation of bioavailable to persistent DOC. We propose this as a novel mechanism whereby the loss of predators from global ecosystems could alter carbon cycling.

To compare the performance and antibiotic-resistance character in different process configurations under different levels of antibiotics, anoxic/oxic-membrane bioreactors (MBR) 1#, MBR2# and a sequencing batch reactor (SBR) were operated with identical operating parameters. MBR1# and SBR were operated under high and increasing levels of antibiotics, MBR2# received constant and low concentration of antibiotics. Microbiological community and antibiotic resistance genes (ARGs) were investigated using 16S rDNA gene high-throughput sequencing and qPCR. More than 90% of penicillin and chlortetracycline were removed due to strong hydrolysis, followed by sulfamethoxazole (69.27%–86.25%) through biodegradation and norfloxacin (28.66%–53.86%) through adsorption. Process configuration affected total nitrogen removal more, while antibiotics concentration affected total phosphorus removal more. MBR1# outperformed SBR in reducing sulfamethoxazole, norfloxacin and ARGs due to the retention effect of the membrane module. Retention efficiency of ARGs in MBRs increased along the operation. Compared to the operational taxonomic unit (OTU) number before antibiotics addition, the OTU number in MBR1# and SBR decreased by 23.7% and 28.7%, while that in MBR2# kept relatively stable. Process configuration contributed to higher dissimilarity of microbial community than antibiotics concentration. The research provides an insight into the influence factors of antibiotics-containing wastewater treatment.

Freshwater lakes and impoundments are an important source of methane into the atmosphere. Methane is the second most important greenhouse gas after carbon dioxide and accounts for about 75% of the climate impact of lakes. The only process that reduces methane emissions from lakes is the oxidation by methanotrophic bacteria. This diverse group of bacteria has the unique ability to use methane and oxygen to gain energy and produce biomass.
Aim of this dissertation was to improve our understanding of the ecology of these bacteria and their importance to prevent methane emissions from freshwater lakes. From the outset, the interdisciplinary project was designed to complement field studies with modelling. The focus was in particular on the methane dynamics during the autumnal overturn of stratified lakes. During this critical phase, large amounts of methane that accumulate in the anoxic hypolimnion during stratification are transported to the surface. [...]

Water and sanitation represents a key battlefront in combating the spread of antimicrobial resistance (AMR). Basic water sanitation infrastructure is an essential first step to protecting public health, thereby limiting the spread of pathogens and the need for antibiotics. AMR presents unique human health risks, meriting new risk assessment frameworks specifically adapted to water and sanitation-borne AMR. There are numerous exposure routes to AMR originating from human waste, each of which must be quantified for its relative risk to human health. Wastewater treatment plants (WWTPs) play a vital role in centralized collection and treatment of human sewage, but there are numerous unresolved questions in terms of the microbial ecological processes occurring within and the extent to which they attenuate or amplify AMR. Research is needed to advance understanding of the fate of resistant bacteria and antibiotic resistance genes (ARGs) in various waste management systems, depending on the local constraints and intended re-use applications. WHO and national AMR action plans would benefit from a more holistic 'One Water' understanding. Here we provide a framework for research, policy, practice, and public engagement aimed at limiting the spread of AMR from water and sanitation in both low-, medium- and high-income countries, alike.

Aerobic methane-oxidizing bacteria (MOB) have the ability to thrive on methane, a potent greenhouse gas with critical climatic implications. They use methane as their sole source of carbon and energy. They phylogenetically mainly group as Alpha- and Gammaproteobacteria, as well as Verrucomicrobia. The metalloenzyme methane-monooxygenase (MMO) catalyses the initial oxidation step, the conversion of methane to methanol. The membrane-bound, particulate MMO (pMMO) is widespread among MOB, however, its atomic structure and active site composition remains a matter of debate. Nevertheless, it is generally agreed that pMMO is a copper-dependent enzyme.
Freshwater lakes are significant contributors to global natural methane emissions. However, MOB inhabiting their water column substantially reduce methane fluxes from the sediments to the atmosphere. MOB are commonly highly enriched at oxic-anoxic transition zones, but increasing evidence suggests that methane oxidation by MOB is also occurring at depths depleted in oxygen. [...]

Methan ist ein starkes Treibhausgas mit kritischen klimatischen Auswirkungen. Aerobe Methan oxidierende Bakterien (MOB) haben die Fähigkeit, Methan als einzige Kohlenstoff- und Energiequelle zu nutzen. Phylogenetisch werden sie hauptsächlich in Alpha- und Gammaproteobakterien sowie Verrucomicrobia eingeteilt. Das Enzym Methan-Monooxygenase (MMO) katalysiert den ersten Oxidationsschritt, die Umsetzung von Methan zu Methanol. Die membrangebundene, partikuläre MMO (pMMO) ist unter MOB weit verbreitet, jedoch sind seine chemische Struktur und die Zusammensetzung seines aktiven Zentrums weiterhin umstritten. Nichtsdestotrotz besteht ein allgemeiner Konsens, dass pMMO ein kupferabhängiges Enzym ist.
Süsswasserseen tragen wesentlich zu den globalen natürlichen Methanemissionen bei. Die in ihrer Wassersäule lebenden MOB konsumieren eine erhebliche Menge an Methan, welches aus den Sedimenten austritt, und andernfalls in die Atmosphäre gelangen würde. MOB findet man normalerweise stark angereichert in oxisch-anoxischen Übergangszonen, es gibt jedoch zunehmend Hinweise, dass Methanoxidation durch MOB auch in sauerstoffarmen Tiefen stattfindet. [...]

Resistance in bacteria of human clinical isolates

Since 2008, different trends have been observed in Gram-positive and Gram-negative bacteria. Methicillin-resistant Staphylococcus aureus MRSA) rates have continued to decrease significantly in invasive isolates, mainly in the western part of Switzerland. This trend was also observed in several other European countries, including the neighboring countries Germany, France and Austria. In contrast, MRSA rates are increasing in wound and abscess samples from outpatients. Penicillin resistance in Streptococcus pneumoniae has also decreased over time. This effect is mainly due to a reduction in the prevalence of more resistant serotypes, due to the introduction of pneumococcal vaccines. Vancomycin resistance in enterococci is still very low, but increasing rates observed during the last months are worrisome.

In contrast, we have observed a steady increase in quinolone resistance and 3rd/4th generation cephalosporin resistance in Escherichia coli and Klebsiella pneumoniae. This increase is observed in most European countries and is consistent with the wide distribution of extended-spectrum-beta-lactamase-(ESBL-)producing isolates. During the last two years, this trend seems to have stabilized in Switzerland, as well as in some other European countries. Fortunately, carbapenem resistance still is rare in E. coli and K. pneumoniae. While carbapenem resistance in E. coli is rare in most European countries as well, increasing carbapenem resistance is observed in Europe in K. pneumoniae; in 2016 resistance rates above 25 % have even been described in Italy, Greece and Romania. To allow a closer monitoring of the distribution of carbapenemase-producing Enterobacteriaceae, an obligation to report these microorganisms was introduced in Switzerland on 1.1.2016.

In Pseudomonas aeruginosa, the increasing resistance rates for piperacillin-tazobactam and ceftazidime peaked in 2015 and have slightly decreased since then, while resistance rates for aminoglycosides are steadily increasing. No significant trends were observed in Acinetobacter spp. and in contrast to Europe, carbapenemase rates were stable. […]

The introduction of antibiotics for both medical and non-medical purposes has had a positive effect on human welfare and agricultural output in the past century. However, there is also an important ecological legacy regarding the use of antibiotics and the consequences of increased levels of these compounds in the environment as a consequence of their use and disposal. This legacy was investigated by quantifying two antibiotic resistance genes (ARG) conferring resistance to tetracycline (tet(W)) and sulfonamide (sul1) in bacterial seed bank DNA in sediments. The industrial introduction of antibiotics caused an abrupt increase in the total abundance of tet(W) and a steady increase in sul1. The abrupt change in tet(W) corresponded to an increase in relative abundance from ca. 1960 that peaked around 1976. This pattern of accumulation was highly correlated with the abundance of specific members of the seed bank community belonging to the phylum Firmicutes. In contrast, the relative abundance of sul1 increased after 1976. This correlated with a taxonomically broad spectrum of bacteria, reflecting sul1 dissemination through horizontal gene transfer. The accumulation patterns of both ARGs correspond broadly to the temporal scale of medical antibiotic use. Our results show that the bacterial seed bank can be used to look back at the historical usage of antibiotics and resistance prevalence.

Aquatic ecosystems serve as a dissemination pathway and a reservoir of both antibiotic resistant bacteria (ARB) and antibiotic resistance genes (ARG). In this study, we investigate the role of the bacterial sporobiota to act as a vector for ARG dispersal in aquatic ecosystems. The sporobiota was operationally defined as the resilient fraction of the bacterial community withstanding a harsh extraction treatment eliminating the easily lysed fraction of the total bacterial community. The sporobiota has been identified as a critical component of the human microbiome, and therefore potentially a key element in the dissemination of ARG in human-impacted environments. A region of Lake Geneva in which the accumulation of ARG in the sediments has been previously linked to the deposition of treated wastewater was selected to investigate the dissemination of tet(W) and sul1, two genes conferring resistance to tetracycline and sulfonamide, respectively. Analysis of the abundance of these ARG within the sporobiome (collection of genes of the sporobiota) and correlation with community composition and environmental parameters demonstrated that ARG can spread across the environment with the sporobiota being the dispersal vector. A highly abundant OTU affiliated with the genus Clostridium was identified as a potential specific vector for the dissemination of tet(W), due to a strong correlation with tet(W) frequency (ARG copy numbers/ng DNA). The high dispersal rate, long-term survival, and potential reactivation of the sporobiota constitute a serious concern in terms of dissemination and persistence of ARG in the environment

Here we used flow cytometry (FCM) and filtration paired with amplicon sequencing to determine the abundance and composition of small low nucleic acid (LNA)-content bacteria in a variety of freshwater ecosystems. We found that FCM clusters associated with LNA-content bacteria were ubiquitous across several ecosystems, varying from 50 to 90% of aquatic bacteria. Using filter-size separation, we separated small LNA-content bacteria (passing 0.4 µm filter) from large bacteria (captured on 0.4 µm filter) and characterized communities with 16S amplicon sequencing. Small and large bacteria each represented different sub-communities within the ecosystems’ community. Moreover, we were able to identify individual operational taxonomical units (OTUs) that appeared exclusively with small bacteria (434 OTUs) or exclusively with large bacteria (441 OTUs). Surprisingly, these exclusive OTUs clustered at the phylum level, with many OTUs appearing exclusively with small bacteria identified as candidate phyla (i.e. lacking cultured representatives) and symbionts. We propose that LNA-content bacteria observed with FCM encompass several previously characterized categories of bacteria (ultramicrobacteria, ultra-small bacteria, candidate phyla radiation) that share many traits including small size and metabolic dependencies on other microorganisms.

Über Fäkalien und Gülle können Antibiotikarückstände und antibiotikaresistente Bakterien ins Abwasser oder in den Boden und schliesslich auch in Gewässer und das Grundwasser gelangen. Ist dies eine Gefahr für das Trinkwasser? Eine Untersuchung von SVGW und Eawag ging dieser Frage nach. Um zu klären, ob sich resistente Bakterien und Resistenzgene auch im Trinkwasser nachweisen lassen und welche Auswirkung dabei die Trinkwasseraufbereitung und die Netzpassage haben, wurde Roh-, Rein- und Netzwasser auf Antibiotikaresistenzen bei heterotrophen Keimen und molekularbiologisch auf das Vorkommen verschiedener Resistenzgene untersucht.

Ammonia oxidation decreases the pH in wastewaters where alkalinity is limited relative to total ammonia. The activity of ammonia oxidizing bacteria (AOB), however, typically decreases with pH and often ceases completely in slightly acidic wastewaters. Nevertheless, nitrification at low pH has been reported in reactors treating human urine, but it has been unclear which organisms are involved. In this study, we followed the population dynamics of ammonia oxidizing organisms and reactor performance in synthetic fully hydrolyzed urine as the pH decreased over time in response to a decrease in the loading rate. Populations of the β-proteobacterial Nitrosomonas europaea lineage were abundant at the initial pH close to 6, but the growth of a possibly novel Nitrosococcus-related AOB genus decreased the pH to the new level of 2.2, challenging the perception that nitrification is inhibited entirely at low pH values, or governed exclusively by β-proteobacterial AOB or archaea. With the pH shift, nitrite oxidizing bacteria were not further detected, but nitrous acid (HNO₂) was still removed through chemical decomposition to nitric oxide (NO) and nitrate. The growth of acid-tolerant γ-proteobacterial AOB should be prevented, by keeping the pH above 5.4, which is a typical pH limit for the N. europaea lineage. Otherwise, the microbial community responsible for high-rate nitrification can be lost, and strong emissions of hazardous volatile nitrogen compounds such as NO are likely.

Although the health of rivers is threatened by multiple anthropogenic stressors with increasing frequency, it remains an open question how riverine microbial communities respond to emerging micropollutants. Here, by using 16S rDNA amplicon sequencing of 60 water samples collected during different hydrological seasons, we investigated the spatio-temporal variation and the co-occurrence patterns of microbial communities in the anthropogenically impacted Jiulong River in China. The results indicated that the riverine microbial co-occurrence network had a non-random, modular structure, which was mainly shaped by the taxonomic relatedness of co-occurring species. Fecal indicator bacteria may survive for prolonged periods of time in river water, but they formed an independent module which had fewer interactions with typical freshwater bacteria. Multivariate analysis demonstrated that nutrients and micropollutants (i.e. pharmaceuticals and personal care products, PPCPs) exerted combined effects in shaping α- and β-diversity of riverine microbial communities. Remarkably, we showed that a hitherto unrecognized disruptive effect of PPCPs on the abundance variations of central species and module communities was stronger than the influence of physico-chemical factors, suggesting the key role played by micropollutants for the microbial co-occurrence relationships in lotic ecosystems. Overall, our findings provide novel insights into community assembly in aquatic environments experiencing anthropogenic stresses.

Swimming organisms can enhance mixing in their natural environments by creating eddies in their wake and by dragging water along. However, these mixing mechanisms are inefficient for microorganisms, because swimming-induced variations in velocity, temperature, and dissolved substances are evened out before they can be advected. In bioconvection, however, microorganisms induce water movement not by propulsion directly but by locally changing the fluid density, which drives convection. Observations of bioconvection have so far mainly been limited to laboratory settings. We report the first observation and quantification of bioconvection within a stratified natural water body. Using in situ measurements, laboratory experiments, and numerical simulations, we demonstrate that the bacterium Chromatium okenii is capable of mixing 0.3 to 1.2 m thick water layers at around 12 m water depth in the Alpine Lake Cadagno (Switzerland). As many species are capable of driving bioconvection, this phenomenon potentially plays a role in species distributions and influences large-scale phenomena like algal blooms.

Im Projekt EcoImpact wurden die ökotoxikologischen und ökologischen Effekte von Mikroverunreinigungen aus Kläranlagen auf aquatische Lebensgemeinschaften untersucht. Chemische und biologische Untersuchungen oberhalb und unterhalb von Kläranlagen deuten auf Effekte dieser Stoffe hin, die von physiologischen Antworten der Organismen bis hin zu veränderten Ökosystemfunktionen wie z. B. dem Laubabbau reichen. Gezielte Rinnenexperimente mit kontrollierter Wasserqualität unterstützen diese Befunde.

The presence of predators can impact a variety of organisms within the ecosystem, including microorganisms. Because the effects of fish predators and their phenotypic differences on microbial communities have not received much attention, we tested how the presence/absence, genotype, and plasticity of the predatory three-spine stickleback (Gasterosteus aculeatus) influence aquatic microbes in outdoor mesocosms. We reared lake and stream stickleback genotypes on contrasting food resources to adulthood, and then added them to aquatic mesocosm ecosystems to assess their impact on the planktonic bacterial community. We also investigated whether the effects of fish persisted following the removal of adults, and the subsequent addition of a homogenous juvenile fish population. The presence of adult stickleback increased the number of bacterial OTUs and altered the size structure of the microbial community, whereas their phenotype affected bacterial community composition. Some of these effects were detectable after adult fish were removed from the mesocosms, and after juvenile fish were placed in the tanks, most of these effects disappeared. Our results suggest that fish can have strong short-term effects on microbial communities that are partially mediated by phenotypic variation of fish.

Antibiotic resistance is a pervasive global health threat. To combat the spread of resistance, it is necessary to consider all possible sources and understand the pathways and mechanisms by which resistance disseminates. Best management practices are urgently needed to provide barriers to the spread of resistance and maximize the lifespan of antibiotics as a precious resource. Herein we advise upon the need for coordinated national and international strategies, highlighting three essential components: 1) Monitoring, 2) Risk Assessment, and 3) Mitigation of antibiotic resistance. Central to all three components is What exactly to monitor, assess, and mitigate? We address this question within an environmental framework, drawing from fundamental microbial ecological processes driving the spread of resistance.

Combined partial nitritation–anammox (PN/A) systems are increasingly being employed for sustainable removal of nitrogen from wastewater, but process instabilities present ongoing challenges for practitioners. The goal of this study was to elucidate differences in process stability between PN/A process variations employing two distinct aggregate types: biofilm [in moving bed biofilm reactors (MBBRs)] and suspended growth biomass. Triplicate reactors for each process variation were studied under baseline conditions and in response to a series of transient perturbations. MBBRs displayed elevated NH₄⁺ removal rates relative to those of suspended growth counterparts over six months of unperturbed baseline operation but also exhibited significantly greater variability in performance. Transient perturbations led to strikingly divergent yet reproducible behavior in biofilm versus suspended growth systems. A temperature perturbation prompted a sharp reduction in NH₄⁺ removal rates with no accumulation of NO₂^– and rapid recovery in MBBRs, compared to a similar reduction in NH₄⁺ removal rates but a high level of accumulation of NO₂^– in suspended growth reactors. Pulse additions of a nitrification inhibitor (allylthiourea) prompted only moderate declines in performance in suspended growth reactors compared to sharp decreases in NH₄⁺ removal rates in MBBRs. Quantitative fluorescence in situ hybridization demonstrated a significant enrichment of anammox in MBBRs compared to suspended growth reactors, and conversely a proportionally higher AOB abundance in suspended growth reactors. Overall, MBBRs displayed significantly increased susceptibility to transient perturbations employed in this study compared to that of suspended growth counterparts (stability parameter), including significantly longer recovery times (resilience). No significant difference in the maximal impact of perturbations (resistance) was apparent. Taken together, our results suggest that aggregate architecture (biofilm vs suspended growth) in PN/A processes exerts an unexpectedly strong influence on process stability.

Hintergrund: Die Häufigkeit antibiotikaresistenter Keime nimmt weltweit seit Jahren zu. Infektionen mit solchen Keimen sind schwieriger, manchmal sogar gar nicht mehr mit Antibiotika behandelbar, was erhebliche medizinische Probleme und zusätzliche Kosten für das Gesundheitswesen mit sich bringt. Die Bakterien in der Umwelt sind ein natürliches Reservoir für Resistenzgene. Aber antibiotikaresistente Bakterien gelangen zum Beispiel auch mit menschlichen oder tierischen Ausscheidungen in die Umwelt. Mit dem Abwasser oder mit dem Oberflächenabfluss gelangen so zusätzlich resistente Organismen in die Gewässer. Die Verbreitung von Resistenzen in der Umwelt wird zunehmend mit Sorge beobachtet. Zur Abschätzung des Risikos solcher Emissionen von resistenten Keimen ist es wichtig die Exposition der Population abzuschätzen. Ein möglicher Weg für die Aufnahme von Resistenten Bakterien durch den Menschen stellt das Trinkwasser dar. Für den SVGW war deshalb die Frage relevant in wie weit resistente Bakterien im Trinkwasser von Schweizer Trinkwasserversorgern vorhanden sind und ob für die Qualität des Trinkwassers in dieser Hinsicht Handlungsbedarf besteht. Das durchgeführte Projekt sollte einen ersten Überblick über das Vorkommen von antibiotikaresistenten Bakterien im Trinkwasser und die Dynamik während der Trinkwasseraufbereitung geben.
Vorgehen: Durchgeführt wurde eine Untersuchung von acht Trinkwasserproduzenten der Schweiz. An jedem Standort wurden Proben von Rohwasser, aufbereitetem Wasser, und Netzwasser auf das Vorkommen von Bakterien mit Resistenzen gegen verschiedene Kombinationen von Antibiotika und auf die Häufigkeit verschiedener Resistenzgene untersucht.
Ergebnisse: Resistente Bakterien waren erwartungsgemäss im Rohwasser häufiger als in aufbereitetem Trinkwasser. Die Belastung war lokal sehr unterschiedlich, Seewasser wies dabei tendenziell höhere und zeitlich variablere Werte auf als Grund- oder Infiltrationswasser. Die Trinkwasseraufbereitung führte in den meisten Fällen zu einer deutlichen Reduktion der Zahl der kultivierbaren resistenten Bakterien. Resistenzen gegen Antibiotika der neueren Generation (Norfloxacin und Ceftazidim) wurden im aufbereiteten Wasser praktisch gar nicht gefunden. Allerdings nahm die Zahl resistenter Bakterien an mehreren Standorten nach der Netzpassage wieder zu. Auch deuten die Ergebnisse für bestimmte Standorte auf eine erhöhte relative Abundanz resistenter Bakterien nach der Aufbereitung hin, also auf eine mögliche Selektion von Resistenzen durch die Aufbereitung. Resistenzgene konnten an allen Standorten nur teilweise nachgewiesen werden, was ebenso wie die geringe Zahl der kultivierbaren antibiotikaresistenten Bakterien die insgesamt geringe Belastung der untersuchten Wasserproben belegt. Die Ergebnisse liefern daher keine Hinweise darauf, dass Trinkwasser in der Schweiz eine bedeutende Rolle für die Exposition der Bevölkerung gegenüber resistenten Bakterien bzw. genetischen Resistenzfaktoren spielen könnte. Allerdings zeigen die Ergebnisse auch, dass es in einigen Trinkwasserbetrieben noch Verbesserungspotential bei der Elimination von resistenten Bakterien oder bei der mikrobiologischen Stabilisierung des Trinkwassers für den Netztransport gibt.

Obwohl in der Schweiz vergleichsweise wenig Antibiotika in der Humanmedizin eingesetzt werden und die Verbrauchszahlen in der Veterinärmedizin seit 2008 stetig sinken, werden auch hierzulande Resistenzen in der aquatischen Umwelt beobachtet. Dieser Artikel stellt Fallstudien zur Verbreitung von Antibiotikaresistenzen in Abwasser und Fliessgewässern in der Schweiz und daraus resultierende Risiken vor. Es wird diskutiert, inwiefern neuartige Technologien in der Abwasserbehandlung (Ozonung, Ultrafiltration) diese biologischen Verunreinigungen reduzieren können.

Ozone, a strong oxidant and disinfectant, seems ideal to cope with future challenges of water treatment, such as micropollutants, multiresistant bacteria (MRB) and even intracellular antibiotic resistance genes (ARG), but information on the latter is scarce. In ozonation experiments we simultaneously determined kinetics and dose-dependent inactivation of Escherichia coli and its plasmid-encoded sulfonamide resistance gene sul1 in different water matrixes. Effects in E. coli were compared to an autochthonous wastewater community. Furthermore, resistance elimination by ozonation and post-treatment were studied in full-scale at a wastewater treatment plant (WWTP). Bacterial inactivation (cultivability, membrane damage) and degradation of sul1 were investigated using plate counts, flow cytometry and quantitative real-time PCR. In experiments with E. coli and the more ozone tolerant wastewater community disruption of intracellular genes was observed at specific ozone doses feasible for full-scale application, but flocs seemed to interfere with this effect. At the WWTP, regrowth during postozonation treatment partly compensated inactivation of MRB, and intracellular sul1 seemed unaffected by ozonation. Our findings indicate that ozone doses relevant for micropollutant abatement from wastewater do not eliminate intracellular ARG.

Microorganisms are vital in mediating the earth’s biogeochemical cycles; yet, despite our rapidly increasing ability to explore complex environmental microbial communities, the relationship between microbial community structure and ecosystem processes remains poorly understood. Here, we address a fundamental and unanswered question in microbial ecology: ‘When do we need to understand microbial community structure to accurately predict function?’ We present a statistical analysis investigating the value of environmental data and microbial community structure independently and in combination for explaining rates of carbon and nitrogen cycling processes within 82 global datasets. Environmental variables were the strongest predictors of process rates but left 44% of variation unexplained on average, suggesting the potential for microbial data to increase model accuracy. Although only 29% of our datasets were significantly improved by adding information on microbial community structure, we observed improvement in models of processes mediated by narrow phylogenetic guilds via functional gene data, and conversely, improvement in models of facultative microbial processes via community diversity metrics. Our results also suggest that microbial diversity can strengthen predictions of respiration rates beyond microbial biomass parameters, as 53% of models were improved by incorporating both sets of predictors compared to 35% by microbial biomass alone. Our analysis represents the first comprehensive analysis of research examining links between microbial community structure and ecosystem function. Taken together, our results indicate that a greater understanding of microbial communities informed by ecological principles may enhance our ability to predict ecosystem process rates relative to assessments based on environmental variables and microbial physiology.

Antibiotic resistance is a threat to human and animal health worldwide, and key measures are required to reduce the risks posed by antibiotic resistance genes that occur in the environment. These measures include the identification of critical points of control, the development of reliable surveillance and risk assessment procedures, and the implementation of technological solutions that can prevent environmental contamination with antibiotic resistant bacteria and genes. In this Opinion article, we discuss the main knowledge gaps, the future research needs and the policy and management options that should be prioritized to tackle antibiotic resistance in the environment.

Methods to estimate microbial diversity have developed rapidly in an effort to understand the distribution and diversity of microorganisms in natural environments. For bacterial communities, the 16S rRNA gene is the phylogenetic marker gene of choice, but most studies select only a specific region of the 16S rRNA to estimate bacterial diversity. Whereas biases derived from from DNA extraction, primer choice and PCR amplification are well documented, we here address how the choice of variable region can influence a wide range of standard ecological metrics, such as species richness, phylogenetic diversity, β-diversity and rank-abundance distributions. We have used Illumina paired-end sequencing to estimate the bacterial diversity of 20 natural lakes across Switzerland derived from three trimmed variable 16S rRNA regions (V3, V4, V5). Species richness, phylogenetic diversity, community composition, β-diversity, and rank-abundance distributions differed significantly between 16S rRNA regions. Overall, patterns of diversity quantified by the V3 and V5 regions were more similar to one another than those assessed by the V4 region. Similar results were obtained when analyzing the datasets with different sequence similarity thresholds used during sequences clustering and when the same analysis was used on a reference dataset of sequences from the Greengenes database. In addition we also measured species richness from the same lake samples using ARISA Fingerprinting, but did not find a strong relationship between species richness estimated by Illumina and ARISA. We conclude that the selection of 16S rRNA region significantly influences the estimation of bacterial diversity and species distributions and that caution is warranted when comparing data from different variable regions as well as when using different sequencing techniques.

Contexte : La résistance des pathogènes aux antibiotiques est une menace grandissante pour la santé dans le monde entier. Or ce phénomène ne se limite pas au domaine médical et hospitalier. Les gènes de résistance peuvent être transmis par les bactéries vivant dans le milieu naturel et les activités humaines peuvent causer l'émission de facteurs de résistance dans l'environnement. La dissémination environnementale des facteurs de résistance est ainsi considérée comme une forme émergente de pollution. L'Eawag a récemment montré que des bactéries résistantes aux antibiotiques étaient libérées avec les eaux usées traitées de la ville de Lausanne et qu'elles affectaient déjà la qualité de l'eau et des sédiments dans la baie de Vidy. Or cette eau est l'une des principales sources d'eau potable de la cité vaudoise et il importe donc de savoir si les bactéries résistantes et les gènes de résistance aux antibiotiques peuvent atteindre les habitants par le réseau de distribution. Alors qu'eauservice effectue une série de filtrations et de désinfections dans des installations ultramodernes pour produire une eau de qualité bactériologique irréprochable, l'effet de ces traitements sur les bactéries environnementales porteuses de facteurs de résistance est mal connu. De même, la question se pose de savoir si une reprise de la croissance bactérienne se produit ensuite dans le réseau de distribution et si l'occurrence et l'abondance des facteurs de résistance s'en trouvent affectées. [...]

Rationale: Pathogens with resistance against antibiotics are on the rise globally, creating a serious threat to public health. Antibiotic resistance is not a phenomenon only of clinical and medical environments. Resistance genes can be acquired from environmental bacteria and human activities release antibiotic resistance factors into the environment. This environmental dissemination of resistance factors has consequently been termed an emerging environmental contaminant. Recent studies conducted by Eawag have shown that antibiotic resistant bacteria are released with the treated wastewater of the city of Lausanne and affect the water and the sediment in the bay of Vidy. This water is one of the main sources of drinking water for Lausanne, raising questions whether antibiotic resistant bacteria or antibiotic resistance genes could be reaching consumers through the drinking water distribution network. While Eauservice Lausanne operates a modern water treatment facility that uses various filtration and disinfection procedures to produce hygienically safe drinking water, it was unclear how these procedures affect environmental bacteria carrying resistance factors. Similarly it was unclear to what extent bacterial regrowth takes place in the distribution system, and how this affects the presence and abundance of resistance factors. [...]

Antibiotic resistance genes (ARGs) are emerging environmental contaminants, known to be continuously discharged into the aquatic environment via human and animal waste. Freshwater aquatic environments represent potential reservoirs for ARG and potentially allow sewage-derived ARG to persist and spread in the environment. This may create increased opportunities for an eventual contact with, and gene transfer to, human and animal pathogens via the food chain or drinking water. However, assessment of this risk requires a better understanding of the level and variability of the natural resistance background and the extent of the human impact. We have analyzed water samples from 21 Swiss lakes, taken at sampling points that were not under the direct influence of local contamination sources and analyzed the relative abundance of ARG using quantitative real-time PCR. Copy numbers of genes mediating resistance to three different broad-spectrum antibiotic classes (sulfonamides: sul1, sul2, tetracyclines: tet(B), tet(M), tet(W) and fluoroquinolones: qnrA) were normalized to copy numbers of bacterial 16S rRNA genes. We used multiple linear regression to assess if ARG abundance is related to human activities in the catchment, microbial community composition and the eutrophication status of the lakes. Sul genes were detected in all sampled lakes, whereas only four lakes contained quantifiable numbers of tet genes, and qnrA remained below detection in all lakes. Our data indicate higher abundance of sul1 in lakes with increasing number and capacity of wastewater treatment plants (WWTPs) in the catchment. sul2 abundance was rather related to long water residence times and eutrophication status. Our study demonstrates the potential of freshwater lakes to preserve antibiotic resistance genes, and provides a reference for ARG abundance from lake systems with low human impact as a baseline for assessing ARG contamination in lake water.

Microbial community assembly and microbial functions are affected by a number of different but coupled drivers such as local habitat characteristics, dispersal rates, and species interactions. In groundwater systems, hydrological flow can introduce spatial structure and directional dependencies among these drivers. We examined the importance of hydrology in structuring bacterial communities and their function within two alpine floodplains during different hydrological states. Piezometers were installed in stream sediments and surrounding riparian zones to assess hydrological flows and also were used as incubation chambers to examine bacterial community structures and enzymatic functions along hydrological flow paths. Spatial eigenvector models in conjunction with models based on physico-chemical groundwater characteristics were used to evaluate the importance of hydrologically-driven processes influencing bacterial assemblages and their enzymatic activities. Our results suggest a strong influence (up to 40% explained variation) of hydrological connectivity on enzymatic activities. The effect of hydrology on bacterial community structure was considerably less strong, suggesting that assemblages demonstrate large functional plasticity/redundancy. Effect size varied between hydrological periods but flow-related mechanisms always had the most power in explaining both bacterial structure and functioning. Changes in hydrology should be considered in models predicting ecosystem functioning and integrated into ecosystem management strategies for floodplains.

Background. The evolution and spread of antibiotic resistance in bacterial pathogens is a serious problem. Both antibiotics and resistant organisms reach the aquatic environment, with wastewater or from agricultural sources. For this reason antibiotic resistance factors have been declared an emerging environmental contaminant.
Literature review. This paper provides a short review on the current state of research on this topic in Switzerland. Switzerland represents one of the countries with the lowest consumption of antibiotics in Europe and has a comparatively low incidence of resistant pathogens. The paper gives an overview on the situation in the medical field and agriculture but puts the main focus on the dissemination of antibiotics and antibiotic resistance into the water cycle. While the available data cannot provide a complete picture, there is solid evidence regarding the dissemination of antibiotics and resistance factors into the environment in Switzerland.
Conclusions. Since even the "best case" Switzerland experiences a considerable level of contamination with resistance factors, other European countries where the general conditions are less favorable are likely to experience more severe contamination. Additional stages for wastewater treatment may, in the context of an integrated strategy, be useful measures against the spread of resistance.

Antibiotic-resistance genes (ARGs) are currently discussed as emerging environmental contaminants. Hospital and municipal sewage are important sources of ARGs for the receiving freshwater bodies. We investigated the spatial distribution of different ARGs (sul1, sul2, tet(B), tet(M), tet(W) and qnrA) in freshwater lake sediments in the vicinity of a point source of treated wastewater. ARG contamination of Vidy Bay, Lake Geneva, Switzerland was quantified using real-time PCR and compared with total mercury (THg), a frequently particle-bound inorganic contaminant with known natural background levels. Two-dimensional mapping of the investigated contaminants in lake sediments with geostatistical tools revealed total and relative abundance of ARGs in close proximity of the sewage discharge point were up to 200-fold above levels measured at a remote reference site (center of the lake) and decreased exponentially with distance. Similar trends were observed in the spatial distribution of different ARGs, whereas distributions of ARGs and THg were only moderately correlated, indicating differences in the transport and fate of these pollutants or additional sources of ARG contamination. The spatial pattern of ARG contamination and supporting data suggest that deposition of particle-associated wastewater bacteria rather than co-selection by, for example, heavy metals was the main cause of sediment ARG contamination.

Glacial alpine landscapes are undergoing rapid transformation due to changes in climate. The loss of glacial ice mass has directly influenced hydrologic characteristics of alpine floodplains. Consequently, hyporheic sediment conditions are likely to change in the future as surface waters fed by glacial water (kryal) become groundwater dominated (krenal). Such environmental shifts may subsequently change bacterial community structure and thus potential ecosystem functioning. We quantitatively investigated the structure of major bacterial groups in glacial and groundwater-fed streams in three alpine floodplains during different hydrologic periods. Our results show the importance of several physico-chemical variables that reflect local geological characteristics as well as water source in structuring bacterial groups. For instance, Alpha-, Betaproteobacteria and Cytophaga-Flavobacteria were influenced by pH, conductivity and temperature as well as by inorganic and organic carbon compounds, whereas phosphorous compounds and nitrate showed specific influence on single bacterial groups. These results can be used to predict future bacterial group shifts, and potential ecosystem functioning, in alpine landscapes under environmental transformation.

Distinct layers of iron(III) and manganese(IV) (Fe/Mn) oxides are found buried within the reducing part of the sediments in Lake Baikal and cause considerable complexity and steep vertical gradients with respect to the redox sequence. For the on-site investigation of the responsible biogeochemical processes, we applied filter tube samplers for the extraction of sediment porewater combined with a portable capillary electrophoresis instrument for the analyses of inorganic cations and anions. On the basis of the new results, the sequence of diagenetic processes leading to the formation, transformation, and dissolution of the Fe/Mn layers was investigated. With two exemplary cores we demonstrate that the dissolution of particulate Fe and Mn is coupled to the anaerobic oxidation of CH₄ (AOM) either via the reduction of sulphate (SO₄²⁻) and the subsequent generation of Fe(II) by S(−II) oxidation, or directly coupled to Fe reduction. Dissolved Fe(II) diffuses upwards to reduce particulate Mn(IV) thus forming a sharp mineral boundary. An alternative dissolution pathway is indicated by the occurrence of anaerobic nitrification of NH₄⁺ observed at locations with Mn(IV). Furthermore, the reasons and consequences of the non-steady-state sediment pattern and the resulting redox discontinuities are discussed and a suggestion for the burial of active Fe/Mn layers is presented.

Sampling methods for characterization of microbial communities in aquifers should target both suspended and attached microorganisms (biofilms). We investigated the effectiveness and reproducibility of low-frequency (200 Hz) sonication pulses on improving extraction efficiency and quality of microorganisms from a petroleum-contaminated aquifer in Studen (Switzerland). Sonication pulses at different power levels (0.65, 0.9, and 1.1 kW) were applied to three different groundwater monitoring wells. Groundwater samples extracted after each pulse were compared with background groundwater samples for cell and adenosine tri-phosphate concentration. Turbidity values were obtained to assess the release of sediment fines and associated microorganisms. The bacterial community in extracted groundwater samples was analyzed by terminal restriction-fragment-length polymorphism and compared with communities obtained from background groundwater samples and from sediment cores. Sonication enhanced the extraction efficiency up to 13-fold, with most of the biomass being associated with the sediment fines extracted with groundwater. Consecutive pulses at constant power were decreasingly effective, while pulses with higher power yielded the best results both in terms of extraction efficiency and quality. Our results indicate that low-frequency sonication may be a viable and cost-effective tool to improve the extraction of microorganisms from aquifers, taking advantage of existing groundwater monitoring wells.

Microbial characterization of aquifers should include samples of both suspended and attached microorganisms (biofilms). We investigated the effect of shear, sonication, and heat on the extraction of microorganisms from water-saturated, packed sediment columns containing established biofilms. Shear was studied by increasing flow velocity of the column eluent, sonication by treating the columns with ultrasound at different power levels, and heat by warming up the column eluent to different temperatures. Effluent cell concentrations were used as a measure of extraction efficiency. Dissolved organic carbon and adenosine tri-phosphate (ATP) concentrations were used to corroborate cell-extraction results. Additionally, ATP was used as an indicator of cell-membrane integrity. Extraction quality was determined by comparing terminal-restriction fragment length polymorphism (T-RFLP) profiles of extracted bacterial communities with destructively sampled sediment-community profiles. Sonication and heat increased the extraction efficiency up to 200-fold and yielded communities comparable to the sediment community. These treatments showed high potential for in-situ application in aquifers.

In diesem Projekt wurde eine publizierte Methode zur Quantifizierung des horizontalen Gentransfers durch konjugativen Transfer von Plasmiden in Umweltproben evaluiert. Es wird ein bakterieller Donor eingesetzt der ein konjugatives Plasmid trägt, welches gfp (green fluorescent protein) Fluoreszenz vermittelt, die im Donororganismus durch ein chromosomal lokalisiertes Repressorsystem unterdrückt wird. Wird das Plasmid an Empfängerzellen anderer Spezies transferiert (sogenannte Transkonjuganten), werden diese durch das gebildete gfp markiert. Die fluoreszenten Zellen werden mittels Durchflusszytometrie detektiert und quantifiziert.
Die Methode würde es erlauben diesen für die Verbreitung von Antibiotikaresistenzgenen in der Umwelt potentiell wichtigen Aspekt semi-quantitativ (Bestimmung der Zahl der Transkonjuganten) und qualitativ (z.B. Zellsortierung und anschliessende Bestimmung der Artenzusammensetzung der markierten Zellen) zu Untersuchen.
Es wurden zahlreiche Versuche durchgeführt um verschiedene Aspekte der Methode zu testen und zu optimieren. Neben methodischen Details wurde vor allem die Überlebensfähigkeit der Donororganismen in verschiedenen Wasserproben überprüft, und verschiedene Experimente durchgeführt um den Nachweis und die Quantifizierung der Bildung von Transkonjuganten mit Reinkulturen und mit natürlichen Bakteriengemeinschaften in verschiedenen Wasserproben zu testen. Ziel war es die Methode zu etablieren und so weit zu entwickeln dass sie für einen semi-quantitativen Vergleich der Häufigkeit von Konjugationsereignissen in verschiedenen Wasserproben (z.B. Abwasser, gereinigtes Abwasser, Seewasser) einsetzbar wäre. [...]

Rationale: Wastewater is increasingly recognized as a source of antibiotic resistant bacteria to receiving aquatic systems like rivers and lakes. The spread of resistance factors into the environment has been recognized as an environmental pollutant. In the context of the global increase of antibiotic resistance among pathogens the release of resistance factors may increase the risk of further dissemination and evolutionary spread of resistance factors. For the WWTP of Lausanne previous research demonstrated that the current treatment allows a considerable number of resistant bacteria to reach Lake Geneva with the treated wastewater, and that elevated resistance factor concentrations are present in the sediment of Vidy Bay around the point of wastewater discharge. The city of Lausanne currently plans the renovation of the WWTP, including additional advanced treatment stages and collaborated with Eawag to obtain data on the effectiveness of the planned facility with regards to the removal of resistance factors.
Purpose: The WWTP of Lausanne tested a pilot scale system for activated charcoal powder (CAP) adsorption and ultrafiltration (UF). In addition to further substantiating results regarding the efficiency of resistance factor removal obtained during a first study conducted in 2012, the purpose of this study was to obtain additional data on the degree of temporal variability in resistance factors in the inflow, biologically treated wastewater, and CAP-UF treated wastewater. [...]

The discovery and introduction of antimicrobial agents to clinical medicine was one of the greatest medical triumphs of the 20th century that revolutionized the treatment of bacterial infections. However, the gradual emergence of populations of antimicrobial-resistant pathogenic bacteria resulting from use, misuse, and abuse of antimicrobials has today become a major global health concern. Antimicrobial resistance (AMR) genes have been suggested to originate from environmental bacteria, as clinically relevant resistance genes have been detected on the chromosome of environmental bacteria. As only a few new antimicrobials have been developed in the last decade, the further evolution of resistance poses a serious threat to public health. Urgent measures are required not only to minimize the use of antimicrobials for prophylactic and therapeutic purposes but also to look for alternative strategies for the control of bacterial infections. This review examines the global picture of antimicrobial resistance, factors that favor its spread, strategies, and limitations for its control and the need for continuous training of all stake-holders i.e., medical, veterinary, public health, and other relevant professionals as well as human consumers, in the appropriate use of antimicrobial drugs.

Glaciated alpine floodplains are responding quickly to climate change through shrinking ice masses. Given the expected future changes in their physicochemical environment, we anticipated variable shifts in structure and ecosystem functioning of hyporheic microbial communities in proglacial alpine streams, depending on present community characteristics and landscape structures. We examined microbial structure and functioning during different hydrologic periods in glacial (kryal) streams and, as contrasting systems, groundwater-fed (krenal) streams. Three catchments were chosen to cover an array of landscape features, including interconnected lakes, differences in local geology and degree of deglaciation. Community structure was assessed by automated ribosomal intergenic spacer analysis and microbial function by potential enzyme activities. We found each catchment to contain a distinct bacterial community structure and different degrees of separation in structure and functioning that were linked to the physicochemical properties of the waters within each catchment. Bacterial communities showed high functional plasticity, although achieved by different strategies in each system. Typical kryal communities showed a strong linkage of structure and function that indicated a major prevalence of specialists, whereas krenal sediments were dominated by generalists. With the rapid retreat of glaciers and therefore altered ecohydrological characteristics, lotic microbial structure and functioning are likely to change substantially in proglacial floodplains in the future. The trajectory of these changes will vary depending on contemporary bacterial community characteristics and landscape structures that ultimately determine the sustainability of ecosystem functioning.

Factors driving bacterial community composition (BCC) and linkages to ecosystem function (EF) are a fundamental interest in microbial ecology. Climate warming is expected to cause a shift from glacial- to groundwater-dominated water sources in alpine catchments due to receding glaciers, which is likely accompanied by a shift in BCC and EF. In this context, we performed a reciprocal transplant experiment of hyporheic sediments within a Swiss alpine floodplain. We assessed the influence of water source (groundwater = krenal, glacial water = kryal) and nutritional state (C, N, and P) on BCC and EF. Experimental response was tested using automated ribosomal intergenic spacer amplification and potential activities of eight different enzymes. BCC from both kryal and krenal systems was highly resistant to changes in water source, yet exhibited pronounced EF flexibility. Major factors determining BCC and EF response were sediment origin followed by seasonal variation in BCC. The gradient in seasonal change in BCC showed different strengths in the two water systems. Krenal BCC was more seasonally stable compared with kryal BCC, although functional plasticity showed the same extent in both. This difference in connectivity between BCC and EF suggests that krenal BCC was dominated by generalists, whereas kryal BCC was dominated by specialists. The weak effect of altered nutritional state on BCC and EF indicates a complex but hierarchically structured relationship among these factors. We conclude that microbial communities in alpine catchments are able to rapidly buffer the effect of shifts in water source on ecosystem functioning.

In order to determine how concentration, composition, and degradation states of particulate organic matter (POM) influence lacustrine bacteria, we analyzed changes in bacterial community (BC) structure and total bacterial cell abundance throughout the water columns of 2 contrasting deep lakes in Switzerland. Lake Brienz is oligotrophic and fully oxic while Lake Zug is eutrophic and partially anoxic. The community composition of the particle-associated (>5 µm) and free-living (>0.2, <5 µm) bacteria was analyzed by automated ribosomal intergenic spacer analysis (ARISA). Cluster analysis showed that the lakes comprised distinct BCs. However, the BCs of both lakes were structured with depth. Although particulate amino compounds appeared to impact the number of ARISA-operational taxonomic units of the particle-associated BCs throughout the lake water columns, the compositional dynamics of this bacterial fraction were affected more strongly by the sampling date and physico-chemical parameters, such as pH. For the free-living BCs, the chlorin index (CI), an indicator for the degradation state of primary produced POM, appeared to significantly impact the vertical community shifts. The vertical changes of the total bacterial cell abundance were also significantly determined by the CI and by shifts in particulate amino compounds. The present study shows that not only bulk environmental parameters but also POM composition and degradation state shape the abundance and composition of lacustrine BCs.

Microbial characterization of aquifers should combine collection of suspended and attached microorganisms (biofilms). This study investigated chemical extraction of microorganisms from water-saturated, packed sediment containing established biofilms. It compares the use of different detachment-promoting agent (DPA) solutions with tap water as eluent in column experiments. Extraction efficiency was determined from cell concentrations in the column effluent. Adenosine triphosphate concentrations were measured to confirm cell extraction and as an indicator of cell membrane integrity. Quality of extracted bacterial communities was assessed by comparing their terminal restriction fragment length polymorphism profiles with destructively sampled sediment-community profiles. Extraction efficiency increased more than 8-fold when deionized water, D-amino acids, or enzymes were used as a DPA. Community profiles recovered by individual DPA solutions showed more pronounced differences at the level of rare microbial groups, whereas abundant groups appeared ubiquitous across treatments. These results suggest that comparison of communities extracted by different DPAs can provide improved information on the occurrence of rare microbial groups in biofilms.

Meromictic Lake Kivu is renowned for its enormous quantity of methane dissolved in the hypolimnion. The methane is primarily of biological origin, and its concentration has been increasing in the past half-century. Insight into the origin of methane production in Lake Kivu has become relevant with the recent commercial extraction of methane from the hypolimnion. This study provides the first culture-independent approach to identifying the archaeal communities present in Lake Kivu sediments at the sediment-water interface. Terminal restriction fragment length polymorphism analysis suggests considerable heterogeneity in the archaeal community composition at varying sample locations. This diversity reflects changes in the geochemical conditions in the sediment and the overlying water, which are an effect of local groundwater inflows. A more in-depth look at the archaeal community composition by clone library analysis revealed diverse phylogenies of Euryarchaeota and Crenarachaeota. Many of the sequences in the clone libraries belonged to globally distributed archaeal clades such as the rice cluster V and Lake Dagow sediment environmental clusters. Several of the determined clades were previously thought to be rare among freshwater sediment Archaea (e.g., sequences related to the SAGMEG-1 clade). Surprisingly, there was no observed relation of clones to known hydrogentrophic methanogens and less than 2 % of clones were related to acetoclastic methanogens. The local variability, diversity, and novelty of the archaeal community structure in Lake Kivu should be considered when making assumptions on the biogeochemical functioning of its sediments.

Real-time quantitative PCR (qPCR) is a widely used technique in microbial community analysis, allowing the quantification of the number of target genes in a community sample. Currently, the standard-curve (SC) method of absolute quantification is widely employed for these kinds of analysis. However, the SC method assumes that the amplification efficiency (E) is the same for both the standard and the sample target template. We analyzed 19 bacterial strains and nine environmental samples in qPCR assays, targeting the nifH and 16S rRNA genes. The E values of the qPCRs differed significantly, depending on the template. This has major implications for the quantification. If the sample and standard differ in their E values, quantification errors of up to orders of magnitude are possible. To address this problem, we propose and test the one-point calibration (OPC) method for absolute quantification. The OPC method corrects for differences in E and was derived from the ΔΔC_T method with correction for E, which is commonly used for relative quantification in gene expression studies. The SC and OPC methods were compared by quantifying artificial template mixtures from Geobacter sulfurreducens (DSM 12127) and Nostoc commune (Culture Collection of Algae and Protozoa [CCAP] 1453/33), which differ in their E values. While the SC method deviated from the expected nifH gene copy number by 3- to 5-fold, the OPC method quantified the template mixtures with high accuracy. Moreover, analyzing environmental samples, we show that even small differences in E between the standard and the sample can cause significant differences between the copy numbers calculated by the SC and the OPC methods.

Hypolimnetic oxygenation systems (HOx) are being increasingly used in freshwater reservoirs to elevate dissolved oxygen levels in the hypolimnion and suppress sediment–water fluxes of soluble metals (e.g. Fe and Mn) which are often microbially mediated. We assessed changes in sediment microbial community structure and corresponding biogeochemical cycling on a reservoir-wide scale as a function of HOx operations. Sediment microbial biomass as quantified by DNA concentration was increased in regions most influenced by the HOx. Following an initial decrease in biomass in the upper sediment while oxygen concentrations were low, biomass typically increased at all depths as the 4-month-long oxygenation season progressed. A distinct shift in microbial community structure was only observed at the end of the season in the upper sediment near the HOx. While this shift was correlated to HOx-enhanced oxygen availability, increased TOC levels and precipitation of Fe- and Mn-oxides, abiotic controls on Fe and Mn cycling, and/or the adaptability of many bacteria to variations in prevailing electron acceptors may explain the delayed response and the comparatively limited changes at other locations. While the sediment microbial community proved remarkably resistant to relatively short-term changes in HOx operations, HOx-induced variation in microbial structure, biomass, and activity was observed after a full season of oxygenation.

Amino sugars (ASs), D-amino acids (D-AAs), and bacterial cell counts were measured in two Swiss lakes to study the contribution of bacterial cells to organic matter (OM) and the fate of ASs and bacterial amino biomarkers during OM degradation. Concentrations of individual ASs (glucosamine, galactosamine, muramic acid, and mannosamine) in the particulate and total OM pools were analyzed in water-column profiles of Lake Brienz (oligotrophic and oxic throughout the entire water column) and Lake Zug (eutrophic, stratified, and permanently anoxic below 170 m) in spring and in fall. Generally, carbon-normalized AS concentrations decreased with water depth, indicating the preferential decomposition of ASs. For Lake Brienz the relative loss of particulate ASs was higher than in Lake Zug, suggesting enhanced AS turnover in an oligotrophic environment. AS ratio changes in the water column revealed a replacement of plankton biomass with OM from heterotrophic microorganisms with increasing water depth. Similar to the ASs, highest carbon normalized D-AA concentrations were found in the upper water column with decreasing concentrations with depth and an increase close to the sediments. In Lake Zug, an increase in the percentage of D-AAs also showed the involvement of bacteria in OM degradation. Estimations of OM derived from bacterial cells using cell counts and the bacterial biomarkers muramic acid and D-AAs gave similar results. For Lake Brienz 0.2–14% of the organic carbon pool originated from bacterial cells, compared to only 0.1–5% in Lake Zug. Based on our estimates, muramic acid appeared primarily associated with bacterial biomass and not with refractory bacterial necromass. Our study underscores that bacteria are not only important drivers of OM degradation in lacustrine systems, they also represent a significant source of OM themselves, especially in oligotrophic lakes.

At present, very little is known about the fate and persistence of multiresistant bacteria (MRB) and their resistance genes in natural aquatic environments. Treated, but partly also untreated sewage of the city of Lausanne, Switzerland is discharged into Vidy Bay (Lake Geneva) resulting in high levels of contamination in this part of the lake. In the present work we have studied the prevalence of MRB and resistance genes in the wastewater stream of Lausanne. Samples from hospital and municipal raw sewage, treated effluent from Lausanne’s wastewater treatment plant (WTP) as well as lake water and sediment samples obtained close to the WTP outlet pipe and a remote site close to a drinking water pump were evaluated for the prevalence of MRB. Selected isolates were identified (16S rRNA gene fragment sequencing) and characterized with regards to further resistances, resistance genes, and plasmids. Mostly, studies investigating this issue have relied on cultivation-based approaches. However, the limitations of these tools are well known, in particular for environmental microbial communities, and cultivation-independent molecular tools should be applied in parallel in order to take non-culturable organisms into account. Here we directly quantified the sulfonamide resistance genes sul1 and sul2 from environmental DNA extracts using TaqMan real-time quantitative PCR. Hospital sewage contained the highest load of MRB and antibiotic resistance genes (ARGs). Wastewater treatment reduced the total bacterial load up to 78% but evidence for selection of extremely multiresistant strains and accumulation of resistance genes was observed. Our data clearly indicated pollution of sediments with ARGs in the vicinity of the WTP outlet. The potential of lakes as reservoirs of MRB and potential risks are discussed.

Microbes, such as heterotrophic bacteria, are crucial in the functional ecology of terrestrial and aquatic ecosystems, being the driving force behind metabolic processes, nutrient retention and cycling, and trophic links with secondary consumers. Bacterial biodiversity, including the actual genetic and functional spectrum, has only recently been investigated and knowledge about alpine lotic ecosystems and their local bacterial communities, in particular, is even scarcer. The ongoing climate-induced change in ecosystems is very likely to influence bacterial communities and consequently ecosystem functioning. To what extent shifts in ecosystem functioning due to future landscape transformation will be buffered by a potential resistance/resilience of bacterial communities is mostly unknown. Glaciated alpine floodplains provide excellent opportunities to test such fundamental ecological questions as they have a high degree of insularity, biotic endemicity and are sensitive to environmental change. In addition, these landscapes harbor structured, hydrologically interconnected and spatio-temporally heterogeneous landscape features such as different stream types or interspersed lakes. These features can provide additional mechanistic understanding of how structures within landscapes drive biodiversity and ecosystem functioning in space and time. Such knowledge could be extrapolated to similar ecosystems that potentially undergo future landscape changes induced by climate change and would improve models of ecosystem services. [...]

Mikroorganismen, wie heterotrophe Bakterien, spielen eine entscheidende Rolle in der funktionellen Ökologie von terrestrischen und aquatischen Ökosystemen. Sie stellen die treibende Kraft hinter metabolischen Prozessen, Nährstoffrückhaltung und Wiederverwertung wie auch die Verbindung mit höheren trophischen Stufen dar. Bakterielle Biodiversität, welche die genetische als auch funktionelle Vielfalt darstellt, ist erst seit relativ kurzer Zeit im Fokus der Wissenschaft. Dies gilt insbesondere für alpine Fliessgewässer und ihre bakteriellen Gemeinschaften.
Die durch den fortschreitenden Klimawandel induzierte Veränderung von Ökosystemen wird höchst wahrscheinlich die bakteriellen Gemeinschaften beeinflussen und konsequenterweise die mit ihnen verbundenen Ökosystemprozesse. Zu welchem Teil Veränderungen von Ökosystemfunktionen durch die zukünftigen Landschaftsveränderungen von einer potentiellen Resistenz/Anpassungsfähigkeit der bakteriellen Gemeinschaft abgefedert wird, ist mehrheitlich unbekannt. Vergletscherte alpine Überschwemmungsebenen bieten eine ausgezeichnete Möglichkeit, solchen fundamentalen ökologischen Fragen nachzugehen, da sie durch ein hohes Mass an Abgeschlossenheit, Endemismus und Sensitivität für Umweltveränderungen charakterisiert sind. Zusätzlich beinhalten sie strukturierte, hydrologisch verbundene, räumlich und zeitlich heterogene Landschaftsmerkmale, wie z.B. verschiedene Flusstypen oder zwischengeschaltete Stillgewässer. Die Erforschung dieser Aspekte kann zusätzliches Verständnis für die landschaftsgebundenen Faktoren generieren, welche die bakterielle und funktionelle Biodiversität beeinflussen. Dieses Wissen könnte man auf ähnliche Ökosysteme übertragen, die durch den Klimawandel beeinflusst werden und dadurch Modelle, welche Ökosystemdienstleistungen voraussagen, verbessern. [...]

Chitin, which is a biopolymer of the amino sugar glucosamine (GlcN), is highly abundant in aquatic ecosystems, and its degradation is assigned a key role in the recycling of carbon and nitrogen. In order to study the significance of chitin decomposition in two temperate freshwater lakes with contrasting trophic and redox conditions, we measured the turnover rate of the chitin analog methylumbelliferyl-N,N′-diacetylchitobioside (MUF-DC) and the presence of chitinase (chiA) genes in zooplankton, water, and sediment samples. In contrast to the eutrophic and partially anoxic lake, chiA gene fragments were detectable throughout the oligotrophic water column and chiA copy numbers per ml of water were up to 15 times higher than in the eutrophic waters. For both lakes, the highest chiA abundance was found in the euphotic zone—the main habitat of zooplankton, but also the site of production of easily degradable algal chitin. The bulk of chitinase activity was measured in zooplankton samples and the sediments, where recalcitrant chitin is deposited. Both, chiA abundance and chitinase activity correlated well with organic carbon, nitrogen, and concentrations of particulate GlcN. Our findings show that chitin, although its overall contribution to the total organic carbon is small (∼0.01 to 0.1%), constitutes an important microbial growth substrate in these temperate freshwater lakes, particularly where other easily degradable carbon sources are scarce.

Microbial communities are at the heart of all ecosystems, and yet microbial community behavior in disturbed environments remains difficult to measure and predict. Understanding the drivers of microbial community stability, including resistance (insensitivity to disturbance) and resilience (the rate of recovery after disturbance) is important for predicting community response to disturbance. Here, we provide an overview of the concepts of stability that are relevant for microbial communities. First, we highlight insights from ecology that are useful for defining and measuring stability. To determine whether general disturbance responses exist for microbial communities, we next examine representative studies from the literature that investigated community responses to press (long-term) and pulse (short-term) disturbances in a variety of habitats. Then we discuss the biological features of individual microorganisms, of microbial populations, and of microbial communities that may govern overall community stability. We conclude with thoughts about the unique insights that systems perspectives – informed by meta-omics data – may provide about microbial community stability.

This chapter summarises the current knowledge on the vertical distribution of methane (CH₄) and its formation in Lake Kivu. Additionally, we review the objectives and restrictions under consideration for sustainable extraction (safe, environmentally acceptable, and economically effective) of the enormous amount of CH₄ from the lake. The harvested CH₄ will be used to produce electricity which is desperately needed in both neighbouring countries: the Democratic Republic of the Congo and Rwanda.
From a system-analysis point of view, the following processes need to be included as the minimum for adequately evaluating the vertical and temporal development of the lake CH₄ during extraction: (1) in situ CH₄ formation occurring in the permanently stratified, anoxic deep-water, (2) CH₄ oxidation in the oxic surface water, (3) natural lake-water upwelling caused by subaquatic springs, (4) artificial lake-water up- and downwelling due to extraction- and reinjection-related flows, and (5) upward diffusion caused by double diffusive convection and weak turbulence. Water density is parameterised as a function of temperature, salinity, and the two gases carbon dioxide and CH₄. For the sake of clarity of the presentation, we use a simplified 4-box analysis and are neglecting the diffusion process (5). This allows for the essence of the CH₄ extraction challenge to be conveyed while avoiding excessive complexities.
The system analysis for different CH₄ extraction concepts clearly reveals that the depth of reinjection of the CH₄-depleted deep-water is critical for the sustainability of the extraction and an optimal CH₄ harvesting plan. Here, the suitability of different reinjection scenarios is compared by evaluating each of them in terms of the objectives "safety" (water column stability), "lake ecological integrity" (nutrient upward fluxes), and "economic viability" (amount of harvestable CH₄). Comparison of model simulations, run over 50 years, revealed that (1) using lake surface (dilution) water for adjusting the density of the reinjection water and (2) reinjecting the nutrient-rich deep-water in the top 190 m are both unacceptable in terms of sustainability.

Methane oxidation is a microbial metabolic process for energy generation and carbon assimilation from methane that is carried out by specific groups of bacteria, the methanotrophs. Methane (CH₄) is oxidized with molecular oxygen (O₂) to carbon dioxide (CO₂).

The microbial population and physicochemical process parameters of a sequencing batch reactor for nitrogen removal from urine were monitored over a 1.5-year period. Microbial community fingerprinting (automated ribosomal intergenic spacer analysis), 16S rRNA gene sequencing, and quantitative PCR on nitrogen cycle functional groups were used to characterize the microbial population. The reactor combined nitrification (ammonium oxidation)/anammox with organoheterotrophic denitrification. The nitrogen elimination rate initially increased by 400%, followed by an extended period of performance degradation. This phase was characterized by accumulation of nitrite and nitrous oxide, reduced anammox activity, and a different but stable microbial community. Outwashing of anammox bacteria or their inhibition by oxygen or nitrite was insufficient to explain reactor behavior. Multiple lines of evidence, e.g., regime-shift analysis of chemical and physical parameters and cluster and ordination analysis of the microbial community, indicated that the system had experienced a rapid transition to a new stable state that led to the observed inferior process rates. The events in the reactor can thus be interpreted to be an ecological regime shift. Constrained ordination indicated that the pH set point controlling cycle duration, temperature, airflow rate, and the release of nitric and nitrous oxides controlled the primarily heterotrophic microbial community. We show that by combining chemical and physical measurements, microbial community analysis and ecological theory allowed extraction of useful information about the causes and dynamics of the observed process instability.

Over half of the bacterioplankton cells in ocean surface waters are capable of carrying out a demethylation of the phytoplankton metabolite dimethylsulfoniopropionate (DMSP) that routes the sulfur moiety away from the climatically active gas dimethylsulfide (DMS). In this study, we tracked changes in dmdA, the gene responsible for DMSP demethylation, over the course of an induced phytoplankton bloom in Gulf of Mexico seawater microcosms. Analysis of >91,000 amplicon sequences indicated 578 different dmdA sequence clusters at a conservative clustering criterion of ≥90% nucleotide sequence identity over the 6-day study. The representation of the major clades of dmdA, several of which are linked to specific taxa through genomes of cultured marine bacterioplankton, remained fairly constant. However, the representation of clusters within these major clades shifted significantly in response to the bloom, including two Roseobacter-like clusters and a SAR11-like cluster, and the best correlate with shifts of the dominant dmdA clades was chlorophyll a concentration. Concurrent 16S rRNA amplification and sequencing indicated the presence of Roseobacter, SAR11, OM60, and marine Rhodospirillales populations, all of which are known to harbor dmdA genes, although the largest taxonomic change was an increase in Flavobacteriaceae, a group not yet demonstrated to have DMSP-demethylating capabilities. Sequence heterogeneity in dmdA and other functional gene populations is becoming increasingly evident with the advent of high-throughput sequencing technologies, and understanding the ecological implications of this heterogeneity is a major challenge for marine microbial ecology.

Unique worldwide, Lake Kivu stores enormous amounts of CH₄ and CO₂. A recent study reported that CH₄ concentrations in the lake have increased by up to 15% in the last 30 years and that accumulation at this rate could lead to catastrophic outgassing by ∼2100. This study investigates the present-day CH₄ formation and oxidation in Lake Kivu. Analyses of ¹⁴C and ¹³C in CH₄ and potential carbon sources revealed that below 260 m, an unusually high ∼65% of the CH₄ originates either from reduction of geogenic CO₂ with mostly geogenic H₂ or from direct inflows of geogenic CH₄. Aerobic CH₄ oxidation, performed by close relatives of type X CH₄-oxidizing bacteria, is the main process preventing CH₄ from escaping to the atmosphere. Anaerobic CH₄ oxidation, carried out by CH₄-oxidizing archaea in the SO₄²⁻-reducing zone, was also detected but is limited by the availability of sulfate. Changes in ¹⁴C_CH4 and ¹³C_CH4 since the 1970s suggest that the amount of CH₄ produced from degrading organic material has increased due to higher accumulation of organic matter. This, as well as the sudden onset of carbonates in the 1960s, has previously been explained by three environmental changes: (1) introduction of nonnative fish, (2) amplified subaquatic inflows following hydrological changes, and (3) increased external inputs due to the fast growing population. The resulting enhancement of primary production and organic matter sedimentation likely caused CH₄ to increase. However, given the large proportion of old CH₄ carbon, we cannot exclude an increased inflow of geogenic H₂ or CH₄.

The fraction of dissolved dimethylsulfoniopropionate (DMSPd) converted by marine bacterioplankton into the climate-active gas dimethylsulfide (DMS) varies widely in the ocean, with the factors that determine this value still largely unknown. One current hypothesis is that the ratio of DMS formation: DMSP demethylation is determined by DMSP availability, with ‘availability’ in both an absolute sense (i.e. concentration in seawater) and in a relative sense (i.e. proportionally to other labile organic S compounds) proposed as the critical factor. We investigated these models during an experimentally induced phytoplankton bloom using a taxon-specific microarray targeting DMSP-related gene transcription in members of the Roseobacter clade, a group hypothesized to play an important role in the surface ocean sulfur cycle and well represented by genome sequences. The array consisted of 1578 probes to 431 genes and was designed to target diverse Roseobacter communities in natural seawater by using hierarchical probe design based on 13 genome sequences. The prevailing pattern of Roseobacter gene transcription showed relative depletion of DMSP-related transcripts during the peak of the bloom, despite increasing absolute concentrations and flux of DMSP-related compounds. DMSPd thus appeared to be assimilated by Roseobacter populations in proportion to its relative abundance in the organic matter pool (the ‘relative sense’ hypothesis), rather than assimilated in preference to other labile organic sulfur or carbon compounds produced during the bloom. The relative investment of the Roseobacter community in DMSP demethylation was not useful for predicting the formation of DMS, however, suggesting a complex regulatory process that may involve multiple taxa and alternative fates of DMSPd.

Variations in influent loads pose major challenges for biological wastewater treatment. Could increased biodiversity allow for more stable operation, and does this mean that biodiversity should also be protected in wastewater treatment plants? Using molecular biological methods, we can now quantify biodiversity and specific groups of microorganisms in complex systems.

Schwankende Zulauffrachten stellen die biologische Abwasserreinigung vor grosse Herausforderungen. Kann hier eine erhöhte Biodiversität helfen und ist Biodiversität also auch in der Kläranlage erstrebenswert? Molekularbiologische Methoden erlauben es uns heute, die Artenvielfalt und spezifische Organismengruppen in komplexen Systemen zu quantifizieren.

Heterotrophic microorganisms are widely recognized as crucial components of ecosystems; yet information on their community structure and dynamics in benthic freshwater habitats is notably scarce. Using denaturing gradient gel electrophoresis (DGGE), we determined the composition of bacterial and fungal communities in a freshwater marsh over four seasons. DGGE revealed diverse bacterial communities in four contrasting microhabitats. The greatest compositional differences emerged between water-column and surface-associated bacteria, although communities associated with sediment also differed from those on plant litter and epiphytic biofilms. Sequences of bacterial clones derived from DGGE bands belonged to the Alphaproteobacteria (31%), Actinobacteria (19%) and Bacteriodetes (19%). Betaproteobacteria were notably absent. Fungal clones obtained from leaf litter were mainly Ascomycota, but two members of the Basidiomycota were also identified. Overall, habitat type was the most important factor explaining variation in bacterial communities among samples, whereas temporal patterns in community composition were less pronounced in spite of large seasonal variation in environmental conditions such as temperature. The observed differences among bacterial communities in different microhabitats were not caused by random variation, but rather appeared to be determined by habitat characteristics, as evidenced by largely congruent community profiles of replicate samples taken at 10–100 m distances within the marsh.

Forefields of receding glaciers are unique and sensitive environments representing natural chronosequences. In such habitats, microbial nitrogen fixation is of particular interest since the low concentration of bioavailable nitrogen is one of the key limitations for growth of plants and soil microorganisms. Asymbiotic nitrogen fixation in the Damma glacier (Swiss Central Alps) forefield soils was assessed using the acetylene reduction assay. Free-living diazotrophic diversity and population structure were resolved by assembling four NifH sequence libraries for bulk and rhizosphere soils at two soil age classes (8- and 70-year ice-free forefield). A total of 318 NifH sequences were analyzed and grouped into 45 unique phylotypes. Phylogenetic analyses revealed a higher diversity as well as a broader distribution of NifH sequences among phylogenetic clusters than formerly observed in other environments. This illustrates the importance of free-living diazotrophs and their potential contribution to the global nitrogen input in this nutrient-poor environment. NifH diversity in bulk soils was higher than in rhizosphere soils. Moreover, the four libraries displayed low similarity values. This indicated that both soil age and the presence of pioneer plants influence diversification and population structure of free-living diazotrophs.

The degassing of volcanic Lake Nyos (Cameroon) provides the opportunity to study the strong isotopic variation of dissolved Fe(II) in a well constrained redox cycle and to identify the governing processes by reaction-transport modeling. Two depth profiles sampled in the lake in March 2004 and 2005 reveal an increase in iron concentrations and δ⁵⁷Fe from around 1 mg L⁻¹ and −1.88‰ at 55 m depth up to 344 mg L⁻¹ and +0.83‰ at the bottom of the lake, respectively. A steep increase in δ⁵⁷Fe was observed across the oxic–anoxic boundary. As many biological and geochemical processes are known to fractionate Fe isotopes, we used a calibrated reaction-transport model to disentangle the processes governing the Fe cycle. The model combines the isotopic signatures of dissolved Fe(II) and settling Fe(III) particles with the concentration profiles and settling fluxes of the Fe particles in the lake. We show that the strong shift in δ⁵⁷Fe is caused by isotopic fractionation via dissimilatory Fe reduction across the oxic–anoxic boundary of Lake Nyos. The shift towards more positive values below the oxic–anoxic interface could be attributed to vertical mixing of a heavier component from the bottom of the lake.

The degradation of chitin by heterotrophic bacteria is an important process in the biogeochemical cycling of carbon and nitrogen. Samples from three Swiss lakes with variable trophic status were amended with dierent types of chitin to enrich their bacterial community utilizing chitin. Bacterial abundance of enrichments over 21 days was analyzed by ow cytometry. Hydrolysis of substrate analogue 4-methylumbelliferyl diacetyl-chitobioside (MUF-diNAG) was measured and chitinase A (chiA) gene copies were enumerated by quantitative polymerase chain reaction (Q-PCR) to assess chitinolytic activity of samples. The bacterial diversity was analyzed by ngerprinting communities with denaturing gradient gel electorphoresis (DGGE). Results show that enrichment of chitinolytic community is possible when amending lake samples with chitin from crab shell, glucosamine or chitin extracted from natural zooplankton. The bacterial abundance increased in all samples, so did the number of chiA gene copies. Hydrolysis of substrate analogue could only be detected in samples enriched on crab shell chitin. Enrichment of bacteria on chitin substrates resulted in bacterial community shifts toward lower species diversity and higher intensity of single species. Similar community composition was found for enrichments on crab shell chitin and glucosamine, but with dierent response rates. Enriched communities from the three dierent lakes shared high similarity even though their initial community compositions varied. Our results suggest that by chitin amendment of lake samples chitinolytic bacteria can be isolated. Their community composition is most likely inuenced by the type of chitin.

Detection and quantification of bacteria related to Dehalococcoides is essential for the development of effective remediation strategies for tetrachloroethene (PCE)-contaminated sites. In this study, the authors applied three methods for quantifying Dehalococcoides-like bacteria in a PCE-contaminated aquifer undergoing natural attenuation in Grenchen, Switzerland: a catalyzed reporter deposition-fluorescence in situ hybridization (CARD-FISH) protocol, a competitive nested polymerase chain reaction (PCR) approach, and a direct PCR end point quantification with external standards. For the investigated aquifer, multiple lines of evidence indicated that reductive dechlorination (and likely dehalorespiration) was an active process. Both PCR-based quantification methods indicated that low numbers of mostly sediment-bound Dehalococcoides were present in the contaminated zone of the Grenchen aquifer. Estimates based on the quantitative PCR methods ranged from 2.1 10⁷ to 1.5 10⁸ sediment-bound Dehalococcoides 16S rRNA gene copies per liter of aquifer volume. In contrast, the liquid phase only contained between 8 and 80 copies per liter aquifer volume. CARD-FISH was not sensitive enough for the quantification of Dehalococcoides cell numbers in this aquifer. Cloning and sequencing of the PCR products revealed the presence of sequences closely related to Dehalococcoides isolates such as D. ethenogenes and Dehalococcoides sp. BAV1. An apparently abundant group (termed “Grenchen Cluster”) of sequences more distantly related to Dehalococcoides was also identified, so far without cultured representatives.

With the beginning of the industrial revolution in the mid 18^th century, the concentrations of greenhouse gases (i.e. carbon dioxide and methane) in the atmosphere have risen dramatically and have resulted in a substantial increase in the global mean surface air temperature. Current scenarios predict a temperature increase of 1.8-3.4 °C by 2100. Combustion of fossil fuel has also increased the release of geologically fixed N into the atmosphere, thus increasing atmospheric N deposition, which is further exacerbated by the intensified use of fertilizers for growing crop and by growing legumes for biological fertilization of agricultural fields. Both global warming and the human alteration of the N cycle are important components of global environmental change, but little is known about the impacts of these factors on ecosystem functioning. Information on aquatic ecosystems, decomposition processes, and microbial communities is particularly scarce and evidence from field-based experiments to elucidate the potential impacts is entirely lacking at present. [...]

Mit dem Beginn der industriellen Revolution Mitte 19. Jahrhundert, sind die Treibhausgaskonzentrationen in der Atmosphäre dramatisch gestiegen. Diese erhöhte Treibhausgaskonzentration hat zu einem starken Anstieg der Lufttemperatur an der Erdoberfläche geführt. Laufende Szenarien prophezeien einen Temperaturanstieg von 1.8-3.4 ºC bis 2100. Verbrennung von fossilem Rohstoff führte zusätzlich zur vermehrten Freisetzung von geologisch fixiertem Stickstoff (N) in die Atmosphäre, was dann wiederum zu einem erhöhten atmosphärischen N-Niederschlag führte. Diese atmosphärische N-Deposition wurde zusätzlich verstärkt durch den vermehrten Gebrauch von N-Düngern in der Landwirtschaft und auch durch den vermehrten Anbau von Leguminosen, um den Boden auf biologische weise zu düngen. Globale Erwärmung und die anthropogene Veränderung des N-Zyklus, sind beides wichtige Komponenten der globalen Umweltveränderung. Es ist jedoch nur wenig darüber bekannt, wie sich diese Veränderungen auf die Ökosystemfunktion auswirken. Zudem gibt es nur wenige Informationen über die Auswirkung solcher Faktoren auf die aquatischen Ökosysteme, den Abbau von organischem Material und die mikrobiellen Lebensgemeinschaften. Ausserdem gibt es bis jetzt keine Feldversuche, die diese Effekte in aquatischen Systemen untersucht haben. [...]

The field of microbial ecology has grown tremendously with the advent of novel molecular techniques, allowing the study of uncultured microbes in the environment, and producing a paradigm shift: now, rather than using bacteria cultures for evaluating cell-specific questions, researchers use RNA and DNA techniques to examine more broad-based ecological and evolutionary constructs such as biogeography and the long-debated biological species concept. Recent work has begun to relate bacteria functional genes to ecosystem processes and functioning, thereby enabling a better understanding of the interactive role of bacteria in different and often-changing environments. The field continues to mature and will most likely make substantial contributions in the future with additional efforts that include metagenomics and genomics. Here we review progress in the application of molecular techniques to study microbial communities in freshwater environments.

Dimethylsufoniopropionate (DMSP) is an abundant organic sulfur compound in the ocean and an important substrate for marine bacterioplankton. The Roseobacter clade of marine alphaproteobacteria, including Silicibacter pomeroyi strain DSS-3, are known to be involved in DMSP degradation in situ. The fate of DMSP has important implications for the global sulfur cycle, but the genes involved in this process and their regulation are largely unknown. S. pomeroyi is capable of performing two major pathways of DMSP degradation, making it an important model organism. Based on the full genome sequence of this strain we designed an oligonucleotide-based microarray for the detection of transcripts of nearly all genes. The array was used to study the transcriptional response of S. pomeroyi cultures to additions of DMSP compared to the non-sulfur compound acetate in a time series experiment. We identified a number of upregulated genes that could be assigned to potential roles in the metabolism of DMSP. DMSP also affected the transcription of genes for transport and metabolism of peptides, amino acids and polyamines. DMSP concentration may thus also play a role as a chemical signal, indicating phytoplankton abundance and eliciting a regulatory response aimed at making maximum use of available nutrients.

This study investigates the small-scale spatial impact of the pioneering plant Leucanthemopsis alpina (L.) Heywood (L. alpina) on biological and chemical–physical parameters in an early successional stage of a glacier forefield. Considering the frequent occurrence of isolated patches of this pioneer plant in the forefield of the Dammaglacier (Switzerland), we hypothesized that the impact of the plant would establish gradients in nutrients, and microbial community structure and activity that may be of importance for the successional processes occurring in the forefield. Our results indicated that, in young successional soils, the rhizosphere effect of L. alpina plant patches can influence bacterial cell numbers and activities not only within the root zone, but even at 20 cm distance from the plant. Microbial cell counts, active cells, and saccharase, glucosidase, and acid phosphatase activities revealed significant distance effects, decreasing from soil directly underneath the plant to soils at 20 and 40 cm distance. Soil chemical and physical parameters did not exhibit significant trends. Fingerprinting analysis of amplified 16S rDNA fragments was used to characterize the microbial community. A selective effect of the plant on the microbial community could not be shown because the bacterial communities were similar regardless of distance to the plant.