How tree traits modulate tree methane fluxes: A review.

Trees can play different roles in the regulation of fluxes of methane (CH4), a greenhouse gas with a warming potential 83 times greater than that of carbon dioxide. Forest soils have the greatest potential for methane uptake compared to other land uses. In addition to their influence on soil CH4 fluxes, trees can act directly as a source or sink of CH4, by transporting CH4 produced in the soil and harbouring the key microorganisms involved in CH4 production and consumption (methanogens and methanotrophs). Tree CH4 fluxes can vary between species characterized by different traits that influence transport and modify the availability of CH4 reaction substrates as well as the habitat for methanogens and methanotrophs. Despite their important role in modulating CH4 fluxes from forest ecosystems, the identity and role of tree traits influencing these fluxes are poorly consolidated in the literature. The objectives of this paper are to 1) Review the functional traits of trees associated with their role in the regulation of CH4 emissions; 2) Assess the importance of inter-specific variability in CH4 fluxes via a global analysis of tree methane fluxes in the literature. Our review highlights that differences in CH4 fluxes between tree species and individuals can be explained by a diversity of traits influencing CH4 transport and microbial production of CH4 such as wood density and secondary metabolites. We propose a functional classification for trees based on the key traits associated with a function in CH4 emissions. We identified the fast-growing species with low wood density, species adapted to flood and species vulnerable to rot as functional groups which can be net sources of CH4 in conditions favorable to CH4 production. The global analysis further demonstrated the importance of taxonomy, with other factors such as land type and season in explaining variability in tree CH4 fluxes.

Disinfection of sink drains to reduce a source of three opportunistic pathogens, during Serratia marcescens clusters in a neonatal intensive care unit.

OBJECTIVE: Evaluate the effects of five disinfection methods on bacterial concentrations in hospital sink drains, focusing on three opportunistic pathogens (OPs): Serratia marcescens, Pseudomonas aeruginosa and Stenotrophomonas maltophilia. DESIGN: Over two years, three sampling campaigns were conducted in a neonatal intensive care unit (NICU). Samples from 19 sink drains were taken at three time points: before, during, and after disinfection. Bacterial concentration was measured using culture-based and flow cytometry methods. High-throughput short sequence typing was performed to identify the three OPs and assess S. marcescens persistence after disinfection at the genotypic level. SETTING: This study was conducted in a pediatric hospitals NICU in Montréal, Canada, which is divided in an intensive and intermediate care side, with individual rooms equipped with a sink. INTERVENTIONS: Five treatments were compared: self-disinfecting drains, chlorine disinfection, boiling water disinfection, hot tap water flushing, and steam disinfection. RESULTS: This study highlights significant differences in the effectiveness of disinfection methods. Chlorine treatment proved ineffective in reducing bacterial concentration, including the three OPs. In contrast, all other drain interventions resulted in an immediate reduction in culturable bacteria (4-8 log) and intact cells (2-3 log). Thermal methods, particularly boiling water and steam treatments, exhibited superior effectiveness in reducing bacterial loads, including OPs. However, in drains with well-established bacterial biofilms, clonal strains of S. marcescens recolonized the drains after heat treatments. CONCLUSIONS: Our study supports thermal disinfection (>80°C) for pathogen reduction in drains but highlights the need for additional trials and the implementation of specific measures to limit biofilm formation.

Carbon amendments in soil microcosms induce uneven response on H2 oxidation activity and microbial community composition.

High-affinity H2-oxidizing bacteria (HA-HOB) thriving in soil are responsible for the most important sink of atmospheric H2. Their activity increases with soil organic carbon content, but the incidence of different carbohydrate fractions on the process has received little attention. Here we tested the hypothesis that carbon amendments impact HA-HOB activity and diversity differentially depending on their recalcitrance and their concentration. Carbon sources (sucrose, starch, cellulose) and application doses (0, 0.1, 1, 3, 5% Ceq soildw-1) were manipulated in soil microcosms. Only 0.1% Ceq soildw-1 cellulose treatment stimulated the HA-HOB activity. Sucrose amendments induced the most significant changes, with an abatement of 50% activity at 1% Ceq soildw-1. This was accompanied with a loss of bacterial and fungal alpha diversity and a reduction of high-affinity group 1 h/5 [NiFe]-hydrogenase gene (hhyL) abundance. A quantitative classification framework was elaborated to assign carbon preference traits to 16S rRNA gene, ITS and hhyL genotypes. The response was uneven at the taxonomic level, making carbon preference a difficult trait to predict. Overall, the results suggest that HA-HOB activity is more susceptible to be stimulated by low doses of recalcitrant carbon, while labile carbon-rich environment is an unfavorable niche for HA-HOB, inducing catabolic repression of hydrogenase.

Serratia marcescens Colonization in a Neonatal Intensive Care Unit Has Multiple Sources, with Sink Drains as a Major Reservoir.

Compelling evidence suggests a contribution of the sink environment to the transmission of opportunistic pathogens from the hospital environment to patients in neonatal intensive care units (NICU). In this study, the distribution of the opportunistic pathogen Serratia marcescens in the sink environment and newborns in a NICU was investigated. More than 500 sink drain and faucet samples were collected over the course of five sampling campaigns undertaken over 3 years. Distribution and diversity of S. marcescens were examined with a modified MacConkey medium and a high-throughput short-sequence typing (HiSST) method. Sink drains were an important reservoir of S. marcescens, with an average of 44% positive samples, whereas no faucet sample was positive. The genotypic diversity of S. marcescens was moderate, with an average of two genotypes per drain, while the spatial distribution of S. marcescens was heterogeneous. The genotypic profiles of 52 clinical isolates were highly heterogeneous, with 27 unique genotypes, of which 71% of isolates were found in more than one patient. S. marcescens acquisition during the first outbreaks was mainly caused by horizontal transmissions. HiSST analyses revealed 10 potential cases of patient-to-patient transmission of S. marcescens, five cases of patient-to-sink transmission, and one bidirectional transfer between sink and patient. Environmental and clinical isolates were found in sink drains up to 1 year after the first detection, supporting persisting drain colonization. This extensive survey suggests multiple reservoirs of S. marcescens within the NICU, including patients and sink drains, but other external sources should also be considered. IMPORTANCE The bacterium Serratia marcescens is an important opportunistic human pathogen that thrives in many environments, can become multidrug resistant, and is often involved in nosocomial outbreaks in neonatal intensive care units (NICU). We evaluated the role of sinks during five suspected S. marcescens outbreaks in a NICU. An innovative approach combining molecular and culture methods was used to maximize the detection and typing of S. marcescens in the sink environment. Our results indicate multiple reservoirs of S. marcescens within the NICU, including patients, sink drains, and external sources. These results highlight the importance of sinks as a major reservoir of S. marcescens and potential sources of future outbreaks.

The forecasting power of the microbiome.

Microorganisms are informative biological integrators of past and present environmental abiotic and biotic conditions. At the same time, they are directly involved in ecosystem processes. Unfortunately, the complexity of microbial communities has so far resulted in most studies being descriptive. Here, we suggest that signals in the microbiome data can be used to forecast future ecosystem processes. The combination of omics with various statistical learning approaches, selected based on accuracy-interpretability and bias-variance trade-offs, will be key to attain this goal, as exemplified by recent studies. The time is ripe for microbial ecologists to fully exploit the forecasting power of microbiomes.

High-Throughput Short Sequence Typing Schemes for Pseudomonas aeruginosa and Stenotrophomonas maltophilia Pure Culture and Environmental DNA.

Molecular typing techniques are utilized to determine genetic similarities between bacterial isolates. However, the use of environmental DNA profiling to assess epidemiologic links between patients and their environment has not been fully explored. This work reports the development and validation of two high-throughput short sequence typing (HiSST) schemes targeting the opportunistic pathogens Pseudomonas aeruginosa and Stenotrophomonas maltophilia, along with a modified SM2I selective medium for the specific isolation of S. maltophilia. These HiSST schemes are based on four discriminative loci for each species and demonstrate high discriminating power, comparable to pairwise whole-genome comparisons. Each scheme includes species-specific PCR primers for precise differentiation from closely related taxa, without the need for upstream culture-dependent methods. For example, the primers targeting the bvgS locus make it possible to distinguish P. aeruginosa from the very closely related Pseudomonas paraeruginosa sp. nov. The selected loci included in the schemes are adapted to massive parallel amplicon sequencing technology. An R-based script implemented in the DADA2 pipeline was assembled to facilitate HiSST analyses for efficient and accurate genotyping of P. aeruginosa and S. maltophilia. We demonstrate the performance of both schemes through in silico validations, assessments against reference culture collections, and a case study involving environmental samples.

Early season soil microbiome best predicts wheat grain quality.

Previous studies have shown that it is possible to accurately predict wheat grain quality and yields using microbial indicators. However, it is uncertain what the best timing for sampling is. For optimal usefulness of this modeling approach, microbial indicators from samples taken early in the season should have the best predictive power. Here, we sampled a field every two weeks across a single growing season and measured a wide array of microbial parameters (amplicon sequencing, abundance of N-cycle related functional genes, and microbial carbon usage) to find the moment when the microbial predictive power for wheat grain baking quality is highest. We found that the highest predictive power for wheat grain quality was for microbial data derived from samples taken early in the season (May-June), which coincides roughly with the seedling and tillering growth stages, that are important for wheat N nutrition. Our models based on LASSO regression also highlighted a set of microbial parameters highly coherent with our previous surveys, including alpha- and beta-diversity indices and N-cycle genes. Taken together, our results suggest that measuring microbial parameters early in the wheat growing season could help farmers better predict wheat grain quality.

The biological sink of atmospheric H2 is more sensitive to spatial variation of microbial diversity than N2O and CO2 emissions in a winter cover crop field trial.

The integration of winter cover crop (WCC) in culture rotations promotes multiple ecosystem services, but concomitant microbial diversity and functioning responses in soil have received less attention. A field trial was established to test the hypothesis that enhanced crop diversity with the integration of WCC in a conventional maize-soy rotation promotes microbial diversity and the biological sink of H2 in soil, while reducing N2O emissions to the atmosphere. Vicia villosa (hairy vetch), Avena sativa (oat), and Raphanus sativus (Daikon radish) were cultivated alone or in combinations and flux measurements were performed throughout two subsequent growing seasons. Soil acted as a net sink for H2 and as a net source for CO2 and N2O. CO2 flux was the most sensitive to WCC whereas a significant spatial variation was observed for H2 flux with soil uptake rates observed in the most productive area two-fold greater than the baseline level. Sequencing and quantification of taxonomic and functional genes were integrated to explain variation in trace gas fluxes with compositional changes in soil microbial communities. Fungal communities were the most sensitive to WCC, but neither community abundance nor beta diversity were found to be indicative of fluxes. The alpha diversity of taxonomic and functional genes, expressed as the number of effective species, was integrated into composite variables extracted from multivariate analyses. Only the composite variable computed with the inverse Simpson's index displayed a reproducible pattern throughout both growing seasons, with functional genes and bacterial 16S rRNA gene defining the two most contrasting gradients. The composite variable was decoupled from WCC treatment and explained 19-20% spatial variation of H2 fluxes. The coupling of composite alpha diversity metrics derived from multiple genes with soil processes warrants further investigations to implement novel indicators of soil health in response to changing management practices at the local scale.

Pseudomonas aeruginosa isolates defective in function of the LasR quorum sensing regulator are frequent in diverse environmental niches.

The saprophyte Pseudomonas aeruginosa is a versatile opportunistic pathogen causing infections in immunocompromised individuals. To facilitate its adaptation to a large variety of niches, this bacterium exploits population density-dependent gene regulation systems called quorum sensing (QS). In P. aeruginosa, three distinct but interrelated QS systems (las, rhl and pqs) regulate the production of many survival and virulence functions. In prototypical strains, the las system, through its transcriptional regulator LasR, is important for the full activation of the rhl and pqs systems. Still, LasR-deficient isolates have been reported, mostly sampled from the lungs of people with cystic fibrosis, where they are considered selected by the chronic infection environment. In this study, we show that a defect in LasR activity appears to be an actually widespread mechanism of adaptation in this bacterium. Indeed, we found abundant LasR-defective isolates sampled from hydrocarbon-contaminated soils, hospital sink drains and meat/fish market environments, using an approach based on phenotypic profiling, supported by gene sequencing. Interestingly, several LasR-defective isolates maintain an active rhl system or are deficient in pqs system signalling. The high prevalence of a LasR-defective phenotype among environmental P. aeruginosa isolates questions the role of QS in niche adaptation.

Predictive microbial-based modelling of wheat yields and grain baking quality across a 500 km transect in Québec.

Crops yield and quality are difficult to predict using soil physico-chemical parameters. Because of their key roles in nutrient cycles, we hypothesized that there is an untapped predictive potential in the soil microbial communities. To test our hypothesis, we sampled soils across 80 wheat fields of the province of Quebec at the beginning of the growing season in May-June. We used a wide array of methods to characterize the microbial communities, their functions and activities, including: (1) amplicon sequencing, (2) real-time PCR quantification and (3) community-level substrate utilization. We also measured grain yield and quality at the end of the growing season, and key soil parameters at sampling. The diversity of fungi, the abundance of nitrification genes and the use of specific organic carbon sources were often the best predictors for wheat yield and grain quality. Using 11 or less parameters, we were able to explain 64-90% of the variation in wheat yield and grain and flour quality across the province of Quebec. Microbial-based regression models outperformed basic soil-based models for predicting wheat quality indicators. Our results suggest that the measurement of microbial parameters early in the season could help predict accurately grain quality and quantity.

A High-Throughput Short Sequence Typing Scheme for Serratia marcescens Pure Culture and Environmental DNA.

Molecular typing methods are used to characterize the relatedness between bacterial isolates involved in infections. These approaches rely mostly on discrete loci or whole-genome sequencing (WGS) analyses of pure cultures. On the other hand, their application to environmental DNA profiling to evaluate epidemiological relatedness among patients and environments has received less attention. We developed a specific, high-throughput short sequence typing (HiSST) method for the opportunistic human pathogen Serratia marcescens. Genes displaying the highest polymorphism were retrieved from the core genome of 60 S. marcescens strains. Bioinformatics analyses showed that use of only three loci (within bssA, gabR, and dhaM) distinguished strains with a high level of efficiency. This HiSST scheme was applied to an epidemiological survey of S. marcescens in a neonatal intensive care unit (NICU). In a first case study, a strain responsible for an outbreak in the NICU was found in a sink drain of this unit, by using HiSST scheme and confirmed by WGS. The HiSST scheme was also applied to environmental DNA extracted from sink-environment samples. Diversity of S. marcescens was modest, with 11, 6, and 4 different sequence types (ST) of gabR, bssA, and dhaM loci among 19 sink drains, respectively. Epidemiological relationships among sinks were inferred on the basis of pairwise comparisons of ST profiles. Further research aimed at relating ST distribution patterns to environmental features encompassing sink location, utilization, and microbial diversity is needed to improve the surveillance and management of opportunistic pathogens. IMPORTANCE Serratia marcescens is an important opportunistic human pathogen, often multidrug resistant and involved in outbreaks of nosocomial infections in neonatal intensive care units. Here, we propose a quick and user-friendly method to select the best typing scheme for nosocomial outbreaks in relating environmental and clinical sources. This method, named high-throughput short sequence typing (HiSST), allows to distinguish strains and to explore the diversity profile of nonculturable S. marcescens. The application of HiSST profile analysis for environmental DNA offers new possibilities to track opportunistic pathogens, identify their origin, and relate their distribution pattern with environmental features encompassing sink location, utilization, and microbial diversity. Adaptation of the method to other opportunistic pathogens is expected to improve knowledge regarding their ecology, which is of significant interest for epidemiological risk assessment and elaborate outbreak mitigation strategies.

The phyllosphere microbiome of host trees contributes more than leaf phytochemicals to variation in the Agrilus planipennis Fairmaire gut microbiome structure.

The microbiome composition of living organisms is closely linked to essential functions determining the fitness of the host for thriving and adapting to a particular ecosystem. Although multiple factors, including the developmental stage, the diet, and host-microbe coevolution have been reported to drive compositional changes in the microbiome structures, very few attempts have been made to disentangle their various contributions in a global approach. Here, we focus on the emerald ash borer (EAB), an herbivorous pest and a real threat to North American ash tree species, to explore the responses of the adult EAB gut microbiome to ash leaf properties, and to identify potential predictors of EAB microbial variations. The relative contributions of specific host plant properties, namely bacterial and fungal communities on leaves, phytochemical composition, and the geographical coordinates of the sampling sites, to the EAB gut microbial community was examined by canonical analyses. The composition of the phyllosphere microbiome appeared to be a strong predictor of the microbial community structure in EAB guts, explaining 53 and 48% of the variation in fungi and bacteria, respectively. This study suggests a potential covariation of the microorganisms associated with food sources and the insect gut microbiome.

Natural Variation in Physicochemical Profiles and Bacterial Communities Associated with Aedes aegypti Breeding Sites and Larvae on Guadeloupe and French Guiana.

Aedes aegypti develop in aquatic habitats in which mosquito larvae are exposed to physicochemical elements and microorganisms that may influence their life cycle and their ability to transmit arboviruses. Little is known about the natural bacterial communities associated with A. aegypti or their relation to the biotic and abiotic characteristics of their aquatic habitats. We characterized the physicochemical properties and bacterial microbiota of A. aegypti breeding sites and larvae on Guadeloupe and in French Guiana. In addition, we explored whether geographic location, the type of breeding site and physicochemical parameters influenced the microbiota associated with this mosquito species. We used large-scale 16S rRNA gene sequencing of 160 breeding sites and 147 pools of A. aegypti larvae and recorded 12 physicochemical parameters at the sampled breeding sites. Ordination plots and multiple linear regression were used to assess the influence of environmental factors on the bacterial microbiota of water and larvae. We found territory-specific differences in physicochemical properties (dissolved oxygen, conductivity) and the composition of bacterial communities in A. aegypti breeding sites that influenced the relative abundance of several bacteria genera (e.g., Methylobacterium, Roseoccocus) on the corresponding larvae. A significant fraction of the bacterial communities identified on larvae, dominated by Herbiconiux and Microvirga genera, were consistently enriched in mosquitoes regardless the location. In conclusion, territory-specific differences observed in the biotic and abiotic properties of A. aegypti breeding sites raise concern about the impact of these changes on pathogen transmission by different A. aegypti populations.

Gut microbiome of the emerald ash borer, Agrilus planipennis Fairmaire, and its relationship with insect population density.

The gut microbial communities of beetles play crucial roles in their adaptive capacities. Environmental factors such as temperature or nutrition naturally affect the insect microbiome, but a shift in local conditions like the population density on a host tree could also lead to changes in the microbiota. The emerald ash borer (EAB), Agrilus planipennis Fairmaire, is an exotic wood borer that causes environmental and economic damage to ash trees in North America. This study aimed to describe the taxonomic structure of the EAB gut microbiome and explore its potential relationship with borer population size. The number of EAB adults collected per tree through a 75 km transect from an epicenter allowed the creation of distinct classes of population density. The Gammaproteobacteria and Ascomycota predominated in bacterial and fungal communities respectively, as determined by sequencing of the bacterial 16S rRNA gene and the fungal internal transcribed spacer ITS2. Species richness and diversity of the bacterial community showed significant dependence on population density. Moreover, α-diversity and ß-diversity analysis revealed some indicator amplicon sequence variants suggesting that the plasticity of the gut microbiome could be related to the EAB population density in host trees.

A Gaseous Milieu: Extending the Boundaries of the Rhizosphere.

Plant root activities shape microbial community functioning in the soil, making the rhizosphere the epicenter of soil biogeochemical processes. With this opinion article, we argue to rethink the rhizosphere boundaries: as gases can diffuse several centimeters away from the roots into the soil, the portion of soil influenced by root activities is larger than the strictly root-adhering soil. Indeed, gases are key drivers of biogeochemical processes due to their roles as energy sources or communication molecules, which has the potential to modify microbial community structure and functioning. In order to get a more holistic perspective on this key environment, we advocate for interdisciplinarity in rhizosphere research by combining knowledge of soluble compounds with gas dynamics.

Gamma irradiation triggers a global stress response in Escherichia coli O157:H7 including base and nucleotides excision repair pathways.

Shiga toxin-producing Escherichia coli O157:H7, one of the most severe human foodborne pathogens, can withstand several stresses, including some levels of γ-irradiation. In this study, the response of E. coli O157:H7 to a sensitization irradiation dose of 0.4 kGy was assessed using RNA-seq transcriptomic at 10 (t10) and 60 (t60) min post-irradiation, combined with an isobaric tags for relative and absolute quantitation (iTRAQ) proteomic analysis at 60 min post-irradiation. Several functions were induced by the treatment, such as base excision repair and nucleotide excision repair pathways; sulfur and histidine metabolism, and virulence mechanisms. Additionally, the sulA gene, coding for the cell division repressor, together with other genes involved in SOS response and repair mechanism (including recA, recN, recJ, recQ, mutM and uvrB) were up-regulated at t60. As the early response to irradiation stress (t10), dnaK, groEL, ibpA, sulfur metabolism genes, as well as those related to oxidative stress were up-regulated, while histidine biosynthesis genes were down-regulated. Acid stress, heat shock, UV resistance and several virulence genes, especially stx2A/stx2b which code for the Shiga toxins characteristic of O157:H7, were upregulated at 60 min post-irradiation. The treatment was also found to increase the levels of CysN, MutM, DinG and DnaC in the cells, proteins involved respectively in sulfur metabolism, base excision repair, recombinational DNA repair and chromosome replication. Our results provide insights into the resistance response of E. coli O157:H7 to a non-lethal irradiation dose. Our findings indicate that E. coli O157:H7 can resist to γ-irradiation through important modifications in genes expression and proteins profiles.

Biological H2 and CO oxidation activities are sensitive to compositional change of soil microbial communities.

Trace gas uptake by microorganisms controls the oxidative capacity of the troposphere, but little is known about how this important function is affected by changes in soil microbial diversity. This article bridges that knowledge gap by examining the response of the microbial community-level physiological profiles (CLPPs), carbon dioxide (CO2) production, and molecular hydrogen (H2) and carbon monoxide (CO) oxidation activities to manipulation of microbial diversity in soil microcosms. Microbial diversity was manipulated by mixing nonsterile and sterile soil with and without the addition of antibiotics. Nonsterile soil without antibiotics was used as a reference. Species composition changed significantly in soil microcosms as a result of dilution and antibiotic treatments, but there was no difference in species richness, according to PCR amplicon sequencing of the bacterial 16S rRNA gene. The CLPP was 15% higher in all dilution and antibiotic treatments than in reference microcosms, but the dilution treatment had no effect on CO2 production. Soil microcosms with dilution treatments had 58%-98% less H2 oxidation and 54%-99% lower CO oxidation, relative to reference microcosms, but did not differ among the antibiotic treatments. These results indicate that H2 and CO oxidation activities respond to compositional changes of microbial community in soil.

Physiological traits and relative abundance of species as explanatory variables of co-occurrence pattern of cultivable bacteria associated with chia seeds.

Deciphering the rules defining microbial community assemblage is envisioned as a promising strategy to improve predictions of pathogens colonization and proliferation in food. Despite the increasing number of studies reporting microbial co-occurrence patterns, only a few attempts have been made to challenge them in experimental or theoretical frameworks. Here, we tested the hypothesis that observed variations in co-occurrence patterns can be explained by taxonomy, relative abundance, and physiological traits of microbial species. We used PCR amplicon sequencing of taxonomic markers to assess distribution and co-occurrence patterns of bacterial and fungal species found in 25 chia (Salvia hispanica L.) samples originating from eight different sources. The use of nutrient-rich and oligotrophic media enabled isolation of 71 strains encompassing 16 bacterial species, of which five corresponded to phylotypes represented in the molecular survey. Tolerance to different growth inhibitors and antibiotics was tested to assess the physiological traits of these isolates. Divergence of physiological traits and relative abundance of each pair of species explained 69% of the co-occurrence profile displayed by cultivable bacterial phylotypes in chia. Validation of this ecological network conceptualization approach to more food products is required to integrate microbial species co-occurrence patterns in predictive microbiology.

Molecular Hydrogen, a Neglected Key Driver of Soil Biogeochemical Processes.

The atmosphere of the early Earth is hypothesized to have been rich in reducing gases such as hydrogen (H2). H2 has been proposed as the first electron donor leading to ATP synthesis due to its ubiquity throughout the biosphere as well as its ability to easily diffuse through microbial cells and its low activation energy requirement. Even today, hydrogenase enzymes enabling the production and oxidation of H2 are found in thousands of genomes spanning the three domains of life across aquatic, terrestrial, and even host-associated ecosystems. Even though H2 has already been proposed as a universal growth and maintenance energy source, its potential contribution as a driver of biogeochemical cycles has received little attention. Here, we bridge this knowledge gap by providing an overview of the classification, distribution, and physiological role of hydrogenases. Distribution of these enzymes in various microbial functional groups and recent experimental evidence are finally integrated to support the hypothesis that H2-oxidizing microbes are keystone species driving C cycling along O2 concentration gradients found in H2-rich soil ecosystems. In conclusion, we suggest focusing on the metabolic flexibility of H2-oxidizing microbes by combining community-level and individual-level approaches aiming to decipher the impact of H2 on C cycling and the C-cycling potential of H2-oxidizing microbes, via both culture-dependent and culture-independent methods, to give us more insight into the role of H2 as a driver of biogeochemical processes.

Strain-level genetic diversity of Methylophaga nitratireducenticrescens confers plasticity to denitrification capacity in a methylotrophic marine denitrifying biofilm.

BACKGROUND: The biofilm of a methanol-fed, fluidized denitrification system treating a marine effluent is composed of multi-species microorganisms, among which Hyphomicrobium nitrativorans NL23 and Methylophaga nitratireducenticrescens JAM1 are the principal bacteria involved in the denitrifying activities. Strain NL23 can carry complete nitrate (NO[Formula: see text]) reduction to N2, whereas strain JAM1 can perform 3 out of the 4 reduction steps. A small proportion of other denitrifiers exists in the biofilm, suggesting the potential plasticity of the biofilm in adapting to environmental changes. Here, we report the acclimation of the denitrifying biofilm from continuous operating mode to batch operating mode, and the isolation and characterization from the acclimated biofilm of a new denitrifying bacterial strain, named GP59. METHODS: The denitrifying biofilm was batch-cultured under anoxic conditions. The acclimated biofilm was plated on Methylophaga specific medium to isolate denitrifying Methylophaga isolates. Planktonic cultures of strains GP59 and JAM1 were performed, and the growth and the dynamics of NO[Formula: see text], nitrite (NO[Formula: see text]) and N2O were determined. The genomes of strains GP59 and JAM1 were sequenced and compared. The transcriptomes of strains GP59 and JAM1 were derived from anoxic cultures. RESULTS: During batch cultures of the biofilm, we observed the disappearance of H. nitrativorans NL23 without affecting the denitrification performance. From the acclimated biofilm, we isolated strain GP59 that can perform, like H. nitrativorans NL23, the complete denitrification pathway. The GP59 cell concentration in the acclimated biofilm was 2-3 orders of magnitude higher than M. nitratireducenticrescens JAM1 and H. nitrativorans NL23. Genome analyses revealed that strain GP59 belongs to the species M. nitratireducenticrescens. The GP59 genome shares more than 85% of its coding sequences with those of strain JAM1. Based on transcriptomic analyses of anoxic cultures, most of these common genes in strain GP59 were expressed at similar level than their counterparts in strain JAM1. In contrast to strain JAM1, strain GP59 cannot reduce NO[Formula: see text] under oxic culture conditions, and has a 24-h lag time before growth and NO[Formula: see text] reduction start to occur in anoxic cultures, suggesting that both strains regulate differently the expression of their denitrification genes. Strain GP59 has the ability to reduce NO[Formula: see text] as it carries a gene encoding a NirK-type NO[Formula: see text] reductase. Based on the CRISPR sequences, strain GP59 did not emerge from strain JAM1 during the biofilm batch cultures but rather was present in the original biofilm and was enriched during this process. DISCUSSION: These results reinforce the unique trait of the species M. nitratireducenticrescens among the Methylophaga genus as facultative anaerobic bacterium. These findings also showed the plasticity of denitrifying population of the biofilm in adapting to anoxic marine environments of the bioreactor.