Microbial life in preferential flow paths in subsurface clayey till revealed by metataxonomy and metagenomics.

BACKGROUND: Subsurface microorganisms contribute to important ecosystem services, yet little is known about how the composition of these communities is affected by small scale heterogeneity such as in preferential flow paths including biopores and fractures. This study aimed to provide a more complete characterization of microbial communities from preferential flow paths and matrix sediments of a clayey till to a depth of 400 cm by using 16S rRNA gene and fungal ITS2 amplicon sequencing of environmental DNA. Moreover, shotgun metagenomics was applied to samples from fractures located 150 cm below ground surface (bgs) to investigate the bacterial genomic adaptations resulting from fluctuating exposure to nutrients, oxygen and water. RESULTS: The microbial communities changed significantly with depth. In addition, the bacterial/archaeal communities in preferential flow paths were significantly different from those in the adjacent matrix sediments, which was not the case for fungal communities. Preferential flow paths contained higher abundances of 16S rRNA and ITS gene copies than the corresponding matrix sediments and more aerobic bacterial taxa than adjacent matrix sediments at 75 and 150 cm bgs. These findings were linked to higher organic carbon and the connectivity of the flow paths to the topsoil as demonstrated by previous dye tracer experiments. Moreover, bacteria, which were differentially more abundant in the fractures than in the matrix sediment at 150 cm bgs, had higher abundances of carbohydrate active enzymes, and a greater potential for mixotrophic growth. CONCLUSIONS: Our results demonstrate that the preferential flow paths in the subsurface are unique niches that are closely connected to water flow and the fluctuating ground water table. Although no difference in fungal communities were observed between these two niches, hydraulically active flow paths contained a significantly higher abundance in fungal, archaeal and bacterial taxa. Metagenomic analysis suggests that bacteria in tectonic fractures have the genetic potential to respond to fluctuating oxygen levels and can degrade organic carbon, which should result in their increased participation in subsurface carbon cycling. This increased microbial abundance and activity needs to be considered in future research and modelling efforts of the soil subsurface.

Snow Microorganisms Colonise Arctic Soils Following Snow Melt.

Arctic soils are constantly subjected to microbial invasion from either airborne, marine, or animal sources, which may impact local microbial communities and ecosystem functioning. However, in winter, Arctic soils are isolated from outside sources other than snow, which is the sole source of microorganisms. Successful colonisation of soil by snow microorganisms depends on the ability to survive and compete of both, the invading and resident community. Using shallow shotgun metagenome sequencing and amplicon sequencing, this study monitored snow and soil microbial communities throughout snow melt to investigate the colonisation process of Arctic soils. Microbial colonisation likely occurred as all the characteristics of successful colonisation were observed. The colonising microorganisms originating from the snow were already adapted to the local environmental conditions and were subsequently subjected to many similar conditions in the Arctic soil. Furthermore, competition-related genes (e.g. motility and virulence) increased in snow samples as the snow melted. Overall, one hundred potentially successful colonisers were identified in the soil and, thus, demonstrated the deposition and growth of snow microorganisms in soils during melt.

Toward a Universal Unit for Quantification of Antibiotic Resistance Genes in Environmental Samples.

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.

Effect of Contact Area and Shape of Anode Current Collectors on Bacterial Community Structure in Microbial Fuel Cells.

Low electrical conductivity of carbon materials is a source of potential loss for large carbonaceous electrode surfaces of MFCs due to the long distance traveled by electrons to the collector. In this paper, different configurations of titanium current collectors were used to connect large surfaces of carbon cloth anodes. The current collectors had different distances and contact areas to the anode. For the same anode surface (490 cm2), increasing the contact area from 28 cm2 to 70 cm2 enhanced power output from 58 mW·m-2 to 107 mW·m-2. For the same contact area (28 cm2), decreasing the maximal distance of current collectors to anodes from 16.5 cm to 7.75 cm slightly increased power output from 50 mW·m-2 to 58 mW·m-2. Molecular biology characterization (qPCR and 16S rRNA gene sequencing) of anodic bacterial communities indicated that the Geobacter number was not correlated with power. Moreover, Geobacter and Desulfuromonas abundance increased with the drop in potential on the anode and with the presence of fermentative microorganisms. Electrochemical impedance spectroscopy (EIS) showed that biofilm resistance decreased with the abundance of electroactive bacteria. All these results showed that the electrical gradient arising from collectors shapes microbial communities. Consequently, current collectors influence the performance of carbon-based anodes for full-scale MFC applications.

A Modified Approach for in Situ Chemical Oxidation Coupled to Biodegradation Enhances Light Nonaqueous Phase Liquid Source-Zone Remediation.

Field and batch experiments were conducted to assess whether a modified approach for in situ chemical oxidation (ISCO) (with MgO2 and Fe2O3 particles recovered from acid mine drainage treatment) can enhance LNAPL (light nonaqueous phase liquid) dissolution and produce bioavailable soluble compounds. This modified ISCO approach was coupled to biodegradation to further remove residual compounds by microbially mediated processes. Pure palm biodiesel (B100) was chosen to represent a poorly water-soluble compound that behaves like LNAPLs, and 100 L was released to a 2 m2 area excavated down to the water table. A past adjacent B100-field experiment under natural attenuation was conducted as a baseline control. Results demonstrated the enhancement of organic compound dissolution and production of soluble compounds due to the modified in situ chemical oxidation. The slow release of H2O2 by MgO2 decomposition (termed partial chemical oxidation) and production of soluble compounds allowed the stimulation of microbial growth and promoted a beneficial response in microbial communities involved in oxidized biodiesel compound biodegradation. This is the first field experiment to demonstrate that this modified ISCO approach coupled to biodegradation could be a feasible strategy for the removal of poorly water-soluble compounds (e.g., biodiesel) and prevent the long-term effects generally posed in source zones.

Functional Basis of Microorganism Classification.

Correctly identifying nearest "neighbors" of a given microorganism is important in industrial and clinical applications where close relationships imply similar treatment. Microbial classification based on similarity of physiological and genetic organism traits (polyphasic similarity) is experimentally difficult and, arguably, subjective. Evolutionary relatedness, inferred from phylogenetic markers, facilitates classification but does not guarantee functional identity between members of the same taxon or lack of similarity between different taxa. Using over thirteen hundred sequenced bacterial genomes, we built a novel function-based microorganism classification scheme, functional-repertoire similarity-based organism network (FuSiON; flattened to fusion). Our scheme is phenetic, based on a network of quantitatively defined organism relationships across the known prokaryotic space. It correlates significantly with the current taxonomy, but the observed discrepancies reveal both (1) the inconsistency of functional diversity levels among different taxa and (2) an (unsurprising) bias towards prioritizing, for classification purposes, relatively minor traits of particular interest to humans. Our dynamic network-based organism classification is independent of the arbitrary pairwise organism similarity cut-offs traditionally applied to establish taxonomic identity. Instead, it reveals natural, functionally defined organism groupings and is thus robust in handling organism diversity. Additionally, fusion can use organism meta-data to highlight the specific environmental factors that drive microbial diversification. Our approach provides a complementary view to cladistic assignments and holds important clues for further exploration of microbial lifestyles. Fusion is a more practical fit for biomedical, industrial, and ecological applications, as many of these rely on understanding the functional capabilities of the microbes in their environment and are less concerned with phylogenetic descent.

Microbial ecology of chlorinated solvent biodegradation.

This study focused on the microbial ecology of tetrachloroethene (PCE) degradation to trichloroethene, cis-1,2-dichloroethene and vinyl chloride to evaluate the relationship between the microbial community and the potential accumulation or degradation of these toxic metabolites. Multiple soil microcosms supplied with different organic substrates were artificially contaminated with PCE. A thymidine analogue, bromodeoxyuridine (BrdU), was added to the microcosms and incorporated into the DNA of actively replicating cells. We compared the total and active bacterial communities during the 50-day incubations by using phylogenic microarrays and 454 pyrosequencing to identify microorganisms and functional genes associated with PCE degradation to ethene. By use of this integrative approach, both the key community members and the ecological functions concomitant with complete PCE degradation could be determined, including the presence and activity of microbial community members responsible for producing hydrogen and acetate, which are critical for Dehalococcoides-mediated PCE degradation. In addition, by correlation of chemical data and phylogenic microarray data, we identified several bacteria that could potentially oxidize hydrogen. These results demonstrate that PCE degradation is dependent on some microbial community members for production of appropriate metabolites, while other members of the community compete for hydrogen in soil at low redox potentials.

Mastering methodological pitfalls for surviving the metagenomic jungle.

Metagenomics is a culture- and PCR-independent approach that is now widely exploited for directly studying microbial evolution, microbial ecology, and developing biotechnologies. Observations and discoveries are critically dependent on DNA extraction methods, sequencing technologies, and bioinformatics tools. The potential pitfalls need to be understood and, to some degree, mastered if the resulting data are to survive scrutiny. In particular, methodological variations appear to affect results from different ecosystems differently, thus increasing the risk of biological and ecological misinterpretation. Part of the difficulty is derived from the lack of knowledge concerning the true microbial diversity and because no approach can guarantee accessing microorganisms in the same proportion in which they exist in the environment. However, the variation between different approaches (e.g. DNA extraction techniques, sequence annotation systems) can be used to evaluate whether observations are meaningful. These methodological variations can be integrated into the error analysis before comparing microbial communities.

Microbial defluorination of TFA, PFOA, and HFPO-DA by a native microbial consortium under anoxic conditions.

In this study, the biodegradability of trifluoroacetate (TFA), perfluorooctanoic acid (PFOA), and perfluoro-2-methyl-3-oxahexanoic acid (HFPO-DA) by a native microbial community was evaluated over a 10-month incubation period. The observed microbial defluorination ratios and removal efficiency were 3.46 ( ± 2.73) % and 8.03 ( ± 3.03) %, 8.44 ( ± 1.88) % and 13.52 ( ± 4.96) %, 3.02 ( ± 0.62) % and 5.45 ( ± 2.99) % for TFA, PFOA and HFPO-DA, respectively. The biodegradation intermediate products, TFA and pentafluoropropionic acid (PFA), of PFOA and HFPO-DA were detected in their biodegradation treatment groups. Furthermore, the concentrations of the PFOA metabolites, perfluorohexanoic acid (PFHxA) and perfluoroheptanoic acid (PFHpA), in the aqueous solutions after incubation were quantified to be 0.21 and 4.14 µg/L. TFA, PFOA and HFPO-DA significantly reduced the microbial diversity and changed the structure of the community. The co-occurrence network analysis showed that low abundance species, such as Flexilinea flocculi, Bacteriovorax stolpii, and g_Sphingomonas, are positively correlated with the generation of fluoride ion, implying their potential collaborative functions contributing to the observed biodefluorination. The findings in this study can provide insights for the biodegradation of perfluoroalkyl carboxylic acids and their emerging alternatives by indigenous microorganisms in the environment.

Emerging contaminants: A One Health perspective.

Environmental pollution is escalating due to rapid global development that often prioritizes human needs over planetary health. Despite global efforts to mitigate legacy pollutants, the continuous introduction of new substances remains a major threat to both people and the planet. In response, global initiatives are focusing on risk assessment and regulation of emerging contaminants, as demonstrated by the ongoing efforts to establish the UN's Intergovernmental Science-Policy Panel on Chemicals, Waste, and Pollution Prevention. This review identifies the sources and impacts of emerging contaminants on planetary health, emphasizing the importance of adopting a One Health approach. Strategies for monitoring and addressing these pollutants are discussed, underscoring the need for robust and socially equitable environmental policies at both regional and international levels. Urgent actions are needed to transition toward sustainable pollution management practices to safeguard our planet for future generations.

Ethyl tert-butyl ether (ETBE) biodegradation by a syntrophic association of Rhodococcus sp. IFP 2042 and Bradyrhizobium sp. IFP 2049 isolated from a polluted aquifer.

Ethyl tert-butyl ether (ETBE) enrichment was obtained by adding contaminated groundwater to a mineral medium containing ETBE as the sole carbon and energy source. ETBE was completely degraded to biomass and CO2 with a transient production of tert-butanol (TBA) and a final biomass yield of 0.37 ± 0.08 mg biomass (dry weight).mg(-1) ETBE. Two bacterial strains, IFP 2042 and IFP 2049, were isolated from the enrichment, and their 16S rRNA genes (rrs) were similar to Rhodococcus sp. (99 % similarity to Rhodococcus erythropolis) and Bradyrhizobium sp. (99 % similarity to Bradyrhizobium japonicum), respectively. Rhodococcus sp. IFP 2042 degraded ETBE to TBA, and Bradyrhizobium sp. IFP 2049 degraded TBA to biomass and CO2. A mixed culture of IFP 2042 and IFP 2049 degraded ETBE to CO2 with a biomass yield similar to the original ETBE enrichment (0.31 ± 0.02 mg biomass.mg(-1) ETBE). Among the genes previously described to be involved in ETBE, MTBE, and TBA degradation, only alkB was detected in Rhodococcus sp. IFP 2042 by PCR, and none were detected in Bradyrhizobium sp. IFP 2049.

Influence of Hydrodynamic Forces on Electroactive Bacterial Adhesion in Microbial Fuel Cell Anodes.

This investigation examined the role of shear stress on the dynamic development of microbial communities within anodic biofilms in single-chamber microbial fuel cells (MFCs). Bacterial attachment to surfaces, often regarded as a crucial step in biofilm formation, may significantly contribute to the selection of electroactive bacteria (EAB). It is well established that hydrodynamic forces, particularly shear forces, have a profound influence on bacterial adhesion. This study postulates that shear stress could select EAB on the anode during the adhesion phase by detaching non-EAB. To examine this hypothesis, MFC reactors equipped with a shear stress chamber were constructed, creating specific shear stress on the anode. The progression of adhesion under various shear stress conditions (1, 10, and 50 mPa) was compared with a control MFC lacking shear stress. The structure of the microbial community was assessed using 16S rRNA gene (rrs) sequencing, and the percentage of biofilm coverage was analyzed using fluorescence microscopy. The results indicate a significant impact of shear stress on the relative abundance of specific EAB, such as Geobacter, which was higher (up to 30%) under high shear stress than under low shear stress (1%). Furthermore, it was noted that shear stress decreased the percentage of biofilm coverage on the anodic surface, suggesting that the increase in the relative abundance of specific EAB occurs through the detachment of other bacteria. These results offer insights into bacterial competition during biofilm formation and propose that shear stress could be utilized to select specific EAB to enhance the electroactivity of anodic biofilms. However, additional investigations are warranted to further explore the effects of shear stress on mature biofilms.

Impact of in situ solar irradiation on snow bacterial communities and functional potential.

Polar regions are increasingly exposed to ultraviolet light due to ozone depletion. Snowpacks contain photochemically active particles that, when irradiated, can lead to the production and accumulation of reactive species that can induce oxidative stress on snow microorganisms. This could generate a selective pressure on snowpack bacteria. In this study, snow microcosms were buried in a snowpack at Ny-Ålesund (Svalbard), either exposed to solar irradiation or incubated in the dark for 10 days, and the bacterial response to solar irradiation was evaluated in situ using a metagenomics approach. Solar irradiation induced a significant decrease in bacterial abundance and richness. Genes involved in glutathione synthesis, sulphur metabolism, and multidrug efflux were significantly enriched in the light, whereas genes related to cell wall assembly and nutrient uptake were more abundant in the dark. This is the first study demonstrating the response of snow bacterial communities to solar irradiation in situ and providing insights into the mechanisms involved. Our research shows that polar sun irradiation is sufficiently intense to impose a selective pressure on snow bacteria and supports the concern that increased ultraviolet exposure due to anthropogenic activities and climatic change could drive critical changes in the structure and functioning of snow bacterial communities.

Metaproteomic and Metagenomic-Coupled Approach to Investigate Microbial Response to Electrochemical Conditions in Microbial Fuel Cells.

MFCs represent a promising sustainable biotechnology that enables the direct conversion of organic matter from wastewater into electricity using bacterial biofilms as biocatalysts. A crucial aspect of MFCs is how electroactive bacteria (EAB) behave and their associated mechanisms during extracellular electron transfer to the anode. A critical phase in the MFC start-up process is the initial colonization of the anode by EAB. Two MFCs were operated with an external resistance of 1000 ohms, one with an applied electrical voltage of 500 mV during the initial four days of biofilm formation and the other without any additional applied voltage. After stabilization of electricity production, total DNA and protein were extracted and sequenced from both setups. The combined metaproteomic/metagenomic analysis revealed that the application of voltage during the colonization step predominantly increased direct electron transfer via cytochrome c, mediated primarily by Geobacter sp. Conversely, the absence of applied voltage during colonization resulted in a broader diversity of bacteria, including Pseudomonas and Aeromonas, which participated in electricity production via mediated electron transfer involving flavin family members.

Sub-inhibitory gentamicin pollution induces gentamicin resistance gene integration in class 1 integrons in the environment.

Antibiotics at sub-inhibitory concentrations are often found in the environment. Here they could impose selective pressure on bacteria, leading to the selection and dissemination of antibiotic resistance, despite being under the inhibitory threshold. The goal of this study was to evaluate the effects of sub-inhibitory concentrations of gentamicin on environmental class 1 integron cassettes in natural river microbial communities. Gentamicin at sub-inhibitory concentrations promoted the integration and selection of gentamicin resistance genes (GmRG) in class 1 integrons after only a one-day exposure. Therefore, sub-inhibitory concentrations of gentamicin induced integron rearrangements, increasing the mobilization potential of gentamicin resistance genes and potentially increasing their dissemination in the environment. This study demonstrates the effects of antibiotics at sub-inhibitory concentrations in the environment and supports concerns about antibiotics as emerging pollutants.

Evolution and interaction of microbial communities in mangrove microbial fuel cells and first description of Shewanella fodinae as electroactive bacterium.

Understanding exoelectrogenic bacteria mechanisms and their interactions in complex biofilm is critical for the development of microbial fuel cells (MFCs). In this article, assumptions concerning the benefits of the complex sediment microbial community for electricity production were explored with both the complex microbial community and isolates identified as Shewanella. Analysis of the microbial community revealed a strong influence of the sediment community on anodes and electrolytes compared to that of only water. Moreover, while Pelobacteraceae-related genera were dominant in our MFCs instead of Desulfuromonas and Geobacter as usually reported, the electroactive Shewanella algae and Shewanella fodinae were isolated and cultivated from the anodic biofilm. S. fodinae, described for the first time as an electroactive bacterium to the best of our knowledge, led to a maximal current density of 3.6 A/m2 set as 0.3 V/SCE in a three-electrode set-up fed with lactate. S. algae, in a complex medium containing several available substrates, showed several preferential oxidative behaviors including a diauxic behavior. In pure culture and under our conditions, S. fodinae and S. algae were not able to use acetate as a sole electron donor. However, their presence in our acetate-fed MFCs and the adaptive behavior of S. algae hint a syntrophic interaction between the bacteria to optimize the use of the substrate in a complex environment.

Selection processes of Arctic seasonal glacier snowpack bacterial communities.

BACKGROUND: Arctic snowpack microbial communities are continually subject to dynamic chemical and microbial input from the atmosphere. As such, the factors that contribute to structuring their microbial communities are complex and have yet to be completely resolved. These snowpack communities can be used to evaluate whether they fit niche-based or neutral assembly theories. METHODS: We sampled snow from 22 glacier sites on 7 glaciers across Svalbard in April during the maximum snow accumulation period and prior to the melt period to evaluate the factors that drive snowpack metataxonomy. These snowpacks were seasonal, accumulating in early winter on bare ice and firn and completely melting out in autumn. Using a Bayesian fitting strategy to evaluate Hubbell's Unified Neutral Theory of Biodiversity at multiple sites, we tested for neutrality and defined immigration rates at different taxonomic levels. Bacterial abundance and diversity were measured and the amount of potential ice-nucleating bacteria was calculated. The chemical composition (anions, cations, organic acids) and particulate impurity load (elemental and organic carbon) of the winter and spring snowpack were also characterized. We used these data in addition to geographical information to assess possible niche-based effects on snow microbial communities using multivariate and variable partitioning analysis. RESULTS: While certain taxonomic signals were found to fit the neutral assembly model, clear evidence of niche-based selection was observed at most sites. Inorganic chemistry was not linked directly to diversity, but helped to identify predominant colonization sources and predict microbial abundance, which was tightly linked to sea spray. Organic acids were the most significant predictors of microbial diversity. At low organic acid concentrations, the snow microbial structure represented the seeding community closely, and evolved away from it at higher organic acid concentrations, with concomitant increases in bacterial numbers. CONCLUSIONS: These results indicate that environmental selection plays a significant role in structuring snow microbial communities and that future studies should focus on activity and growth. Video Abstract.

Gentamicin at sub-inhibitory concentrations selects for antibiotic resistance in the environment.

Antibiotics released into the environment at low (sub-inhibitory) concentrations could select for antibiotic resistance that might disseminate to the human microbiome. In this case, low-level anthropogenic sources of antibiotics would have a significant impact on human health risk. In order to provide data necessary for the evaluation of this risk, we implemented river water microcosms at both sub-inhibitory and inhibitory concentrations of gentamicin as determined previously based on bacterial growth in enriched media. Using metagenomic sequencing and qPCR/RT-qPCR, we assessed the effects of gentamicin on water bacterial communities and their resistome. A change in the composition of total and active communities, as well as a gentamicin resistance gene selection identified via mobile genetic elements, was observed during a two-day exposure. We demonstrated the effects of sub-inhibitory concentrations of gentamicin on bacterial communities and their associated resistome in microcosms (simulating in situ conditions). In addition, we established relationships between antibiotic dose and the magnitude of the community response in the environment. The scope of resistance selection under sub-inhibitory concentrations of antibiotics and the mechanisms underlying this process might provide the basis for understanding resistance dispersion and associated risks in relatively low impacted ecosystems.

Sequencing Depth Has a Stronger Effect than DNA Extraction on Soil Bacterial Richness Discovery.

Although Next-Generation Sequencing techniques have increased our access to the soil microbiome, each step of soil metagenomics presents inherent biases that prevent the accurate definition of the soil microbiome and its ecosystem function. In this study, we compared the effects of DNA extraction and sequencing depth on bacterial richness discovery from two soil samples. Four DNA extraction methods were used, and sequencing duplicates were generated for each DNA sample. The V3-V4 region of the 16S rRNA gene was sequenced to determine the taxonomical richness measured by each method at the amplicon sequence variant (ASV) level. Both the overall functional richness and antibiotic resistance gene (ARG) richness were evaluated by metagenomics sequencing. Despite variable DNA extraction methods, sequencing depth had a greater influence on bacterial richness discovery at both the taxonomical and functional levels. Sequencing duplicates from the same sample provided access to different portions of bacterial richness, and this was related to differences in the sequencing depth. Thus, the sequencing depth introduced biases in the comparison of DNA extraction methods. An optimisation of the soil metagenomics workflow is needed in order to sequence at a sufficient and equal depth. This would improve the accuracy of metagenomic comparisons and soil microbiome profiles.

Environmental and Anthropogenic Factors Shape the Snow Microbiome and Antibiotic Resistome.

Winter tourism can generate environmental pollution and affect microbial ecology in mountain ecosystems. This could stimulate the development of antibiotic resistance in snow and its dissemination through the atmosphere and through snow melting. Despite these potential impacts, the effect of winter tourism on the snow antibiotic resistome remains to be elucidated. In this study, snow samples subjected to different levels of anthropogenic activities and surrounding forest were obtained from the Sudety Mountains in Poland to evaluate the impact of winter tourism on snow bacteria using a metagenomic approach. Bacterial community composition was determined by the sequencing of the V3-V4 hypervariable region of the 16S rRNA gene and the composition of the antibiotic resistome was explored by metagenomic sequencing. Whereas environmental factors were the main drivers of bacterial community and antibiotic resistome composition in snow, winter tourism affected resistome composition in sites with similar environmental conditions. Several antibiotic resistance genes (ARGs) showed a higher abundance in sites subjected to human activities. This is the first study to show that anthropogenic activities may influence the antibiotic resistome in alpine snow. Our results highlight the need to survey antibiotic resistance development in anthropogenically polluted sites.