Julien Tremblay

Defining the core Arabidopsis thaliana root microbiome

Land plants associate with a root microbiota distinct from the complex microbial community present in surrounding soil. The microbiota colonizing the rhizosphere (immediately surrounding the root) and the endophytic compartment (within the root) contribute to plant growth, productivity, carbon sequestration and phytoremediation. Colonization of the root occurs despite a sophisticated plant immune system, suggesting finely tuned discrimination of mutualists and commensals from pathogens. Genetic principles governing the derivation of host-specific endophyte communities from soil communities are poorly understood. Here we report the pyrosequencing of the bacterial 16S ribosomal RNA gene of more than 600 Arabidopsis thaliana plants to test the hypotheses that the root rhizosphere and endophytic compartment microbiota of plants grown under controlled conditions in natural soils are sufficiently dependent on the host to remain consistent across different soil types and developmental stages, and sufficiently dependent on host genotype to vary between inbred Arabidopsis accessions. We describe different bacterial communities in two geochemically distinct bulk soils and in rhizosphere and endophytic compartments prepared from roots grown in these soils. The communities in each compartment are strongly influenced by soil type. Endophytic compartments from both soils feature overlapping, low-complexity communities that are markedly enriched in Actinobacteria and specific families from other phyla, notably Proteobacteria. Some bacteria vary quantitatively between plants of different developmental stage and genotype. Our rigorous definition of an endophytic compartment microbiome should facilitate controlled dissection of plant–microbe interactions derived from complex soil communities.

Primer and platform effects on 16S rRNA tag sequencing

Sequencing of 16S rRNA gene tags is a popular method for profiling and comparing microbial communities. The protocols and methods used, however, vary considerably with regard to amplification primers, sequencing primers, sequencing technologies; as well as quality filtering and clustering. How results are affected by these choices, and whether data produced with different protocols can be meaningfully compared, is often unknown. Here we compare results obtained using three different amplification primer sets (targeting V4, V6–V8, and V7–V8) and two sequencing technologies (454 pyrosequencing and Illumina MiSeq) using DNA from a mock community containing a known number of species as well as complex environmental samples whose PCR-independent profiles were estimated using shotgun sequencing. We find that paired-end MiSeq reads produce higher quality data and enabled the use of more aggressive quality control parameters over 454, resulting in a higher retention rate of high quality reads for downstream data analysis. While primer choice considerably influences quantitative abundance estimations, sequencing platform has relatively minor effects when matched primers are used. Beta diversity metrics are surprisingly robust to both primer and sequencing platform biases.

Genome-wide selective sweeps and gene-specific sweeps in natural bacterial populations

Multiple models describe the formation and evolution of distinct microbial phylogenetic groups. These evolutionary models make different predictions regarding how adaptive alleles spread through populations and how genetic diversity is maintained. Processes predicted by competing evolutionary models, for example, genome-wide selective sweeps vs gene-specific sweeps, could be captured in natural populations using time-series metagenomics if the approach were applied over a sufficiently long time frame. Direct observations of either process would help resolve how distinct microbial groups evolve. Here, from a 9-year metagenomic study of a freshwater lake (2005–2013), we explore changes in single-nucleotide polymorphism (SNP) frequencies and patterns of gene gain and loss in 30 bacterial populations. SNP analyses revealed substantial genetic heterogeneity within these populations, although the degree of heterogeneity varied by >1000-fold among populations. SNP allele frequencies also changed dramatically over time within some populations. Interestingly, nearly all SNP variants were slowly purged over several years from one population of green sulfur bacteria, while at the same time multiple genes either swept through or were lost from this population. These patterns were consistent with a genome-wide selective sweep in progress, a process predicted by the ‘ecotype model’ of speciation but not previously observed in nature. In contrast, other populations contained large, SNP-free genomic regions that appear to have swept independently through the populations prior to the study without purging diversity elsewhere in the genome. Evidence for both genome-wide and gene-specific sweeps suggests that different models of bacterial speciation may apply to different populations coexisting in the same environment.

Transplanting Soil Microbiomes Leads to Lasting Effects on Willow Growth, but not on the Rhizosphere Microbiome.

Plants interact closely with microbes, which are partly responsible for plant growth, health, and adaptation to stressful environments. Engineering the plant-associated microbiome could improve plant survival and performance in stressful environments such as contaminated soils. Here, willow cuttings were planted into highly petroleum-contaminated soils that had been gamma-irradiated and subjected to one of four treatments: inoculation with rhizosphere soil from a willow that grew well (LA) or sub-optimally (SM) in highly contaminated soils or with bulk soil in which the planted willow had died (DE) or no inoculation (CO). Samples were taken from the starting inoculum, at the beginning of the experiment (T0) and after 100 days of growth (TF). Short hypervariable regions of archaeal/bacterial 16S rRNA genes and the fungal ITS region were amplified from soil DNA extracts and sequenced on the Illumina MiSeq. Willow growth was monitored throughout the experiment, and plant biomass was measured at TF. CO willows were significantly smaller throughout the experiment, while DE willows were the largest at TF. Microbiomes of different treatments were divergent at T0, but for most samples, had converged on highly similar communities by TF. Willow biomass was more strongly linked to overall microbial community structure at T0 than to microbial community structure at TF, and the relative abundance of many genera at T0 was significantly correlated to final willow root and shoot biomass. Although microbial communities had mostly converged at TF, lasting differences in willow growth were observed, probably linked to differences in T0 microbial communities.

H2-saturation of high affinity H2-oxidizing bacteria alters the ecological niche of soil microorganisms unevenly among taxonomic groups

Soil microbial communities are continuously exposed to H2 diffusing into the soil from the atmosphere. N2-fixing nodules represent a peculiar microniche in soil where H2 can reach concentrations up to 20,000 fold higher than in the global atmosphere (0.530 ppmv). In this study, we investigated the impact of H2 exposure on soil bacterial community structure using dynamic microcosm chambers simulating soil H2 exposure from the atmosphere and N2-fixing nodules. Biphasic kinetic parameters governing H2 oxidation activity in soil changed drastically upon elevated H2 exposure, corresponding to a slight but significant decay of high affinity H2-oxidizing bacteria population, accompanied by an enrichment or activation of microorganisms displaying low-affinity for H2. In contrast to previous studies that unveiled limited response by a few species, the relative abundance of 958 bacterial ribotypes distributed among various taxonomic groups, rather than a few distinct taxa, was influenced by H2 exposure. Furthermore, correlation networks showed important alterations of ribotype covariation in response to H2 exposure, suggesting that H2 affects microbe-microbe interactions in soil. Taken together, our results demonstrate that H2-rich environments exert a direct influence on soil H2-oxidizing bacteria in addition to indirect effects on other members of the bacterial communities.

The tale of a neglected energy source: Elevated hydrogen exposure affects both microbial diversity and function in soil.

ABSTRACT The enrichment of H2-oxidizing bacteria (HOB) by H2 generated by nitrogen-fixing nodules has been shown to have a fertilization effect on several different crops. The benefit of HOB is attributed to their production of plant growth-promoting factors, yet their interactions with other members of soil microbial communities have received little attention. Here we report that the energy potential of H2, when supplied to soil, alters ecological niche partitioning of bacteria and fungi, with multifaceted consequences for both generalist and specialist microbial functions. We used dynamic microcosms to expose soil to the typical atmospheric H2 mixing ratio (0.5 ppmv) permeating soils, as well as mixing ratios comparable to those found at the soil-nodule interface (10,000 ppmv). Elevated H2 exposure exerted direct effects on two HOB subpopulations distinguished by their affinity for H2 while enhancing community level carbon substrate utilization potential and lowering CH4 uptake activity in soil. We found that H2 triggered changes in the abundance of microorganisms that were reproducible yet inconsistent across soils at the taxonomic level and even among HOB. Overall, H2 exposure altered microbial process rates at an intensity that depends upon soil abiotic and biotic features. We argue that further examination of direct and indirect effects of H2 on soil microbial communities will lead to a better understanding of the H2 fertilization effect and soil biogeochemical processes. IMPORTANCE An innovative dynamic microcosm chamber system was used to demonstrate that H2 diffusing in soil triggers changes in the distribution of HOB and non-HOB. Although the response was uneven at the taxonomic level, an unexpected coordinated response of microbial functions was observed, including abatement of CH4 oxidation activity and stimulation of carbon turnover. Our work suggests that elevated H2 rewires soil biogeochemical structure through a combination of direct effects on the growth and persistence of HOB and indirect effects on a variety of microbial processes involving HOB and non-HOB.

The Willow Microbiome Is Influenced by Soil Petroleum-Hydrocarbon Concentration with Plant Compartment-Specific Effects

The interaction between plants and microorganisms, which is the driving force behind the decontamination of petroleum hydrocarbon (PHC) contamination in phytoremediation technology, is poorly understood. Here, we aimed at characterizing the variations between plant compartments in the microbiome of two willow cultivars growing in contaminated soils. A field experiment was set-up at a former petrochemical plant in Canada and after two growing seasons, bulk soil, rhizosphere soil, roots, and stems samples of two willow cultivars (Salix purpurea cv. FishCreek, and Salix miyabeana cv. SX67) growing at three PHC contamination concentrations were taken. DNA was extracted and bacterial 16S rRNA gene and fungal internal transcribed spacer (ITS) regions were amplified and sequenced using an Ion Torrent Personal Genome Machine (PGM). Following multivariate statistical analyses, the level of PHC-contamination appeared as the primary factor influencing the willow microbiome with compartment-specific effects, with significant differences between the responses of bacterial, and fungal communities. Increasing PHC contamination levels resulted in shifts in the microbiome composition, favoring putative hydrocarbon degraders, and microorganisms previously reported as associated with plant health. These shifts were less drastic in the rhizosphere, root, and stem tissues as compared to bulk soil, probably because the willows provided a more controlled environment, and thus, protected microbial communities against increasing contamination levels. Insights from this study will help to devise optimal plant microbiomes for increasing the efficiency of phytoremediation technology.

Chemical dispersants enhance the activity of oil- and gas condensate-degrading marine bacteria

Application of chemical dispersants to oil spills in the marine environment is a common practice to disperse oil into the water column and stimulate oil biodegradation by increasing its bioavailability to indigenous bacteria capable of naturally metabolizing hydrocarbons. In the context of a spill event, the biodegradation of crude oil and gas condensate off eastern Canada is an essential component of a response strategy. In laboratory experiments, we simulated conditions similar to an oil spill with and without the addition of chemical dispersant under both winter and summer conditions and evaluated the natural attenuation potential for hydrocarbons in near-surface sea water from the vicinity of crude oil and natural gas production facilities off eastern Canada. Chemical analyses were performed to determine hydrocarbon degradation rates, and metagenome binning combined with metatranscriptomics was used to reconstruct abundant bacterial genomes and estimate their oil degradation gene abundance and activity. Our results show important and rapid structural shifts in microbial populations in all three different oil production sites examined following exposure to oil, oil with dispersant and dispersant alone. We found that the addition of dispersant to crude oil enhanced oil degradation rates and favored the abundance and expression of oil-degrading genes from a Thalassolituus sp. (that is, metagenome bin) that harbors multiple alkane hydroxylase (alkB) gene copies. We propose that this member of the Oceanospirillales group would be an important oil degrader when oil spills are treated with dispersant.

Metagenomic survey of the taxonomic and functional microbial communities of seawater and sea ice from the Canadian Arctic

Climate change has resulted in an accelerated decline of Arctic sea ice since 2001 resulting in primary production increases and prolongation of the ice-free season within the Northwest Passage. The taxonomic and functional microbial community composition of the seawater and sea ice of the Canadian Arctic is not very well known. Bacterial communities from the bottom layer of sea ice cores and surface water from 23 locations around Cornwallis Island, NU, Canada, were extensively screened. The bacterial 16S rRNA gene was sequenced for all samples while shotgun metagenomics was performed on selected samples. Bacterial community composition showed large variation throughout the sampling area both for sea ice and seawater. Seawater and sea ice samples harbored significantly distinct microbial communities, both at different taxonomic levels and at the functional level. A key difference between the two sample types was the dominance of algae in sea ice samples, as visualized by the higher relative abundance of algae and photosynthesis-related genes in the metagenomic datasets and the higher chl a concentrations. The relative abundance of various OTUs and functional genes were significantly correlated with multiple environmental parameters, highlighting many potential environmental drivers and ecological strategies.

Broth versus Surface-Grown Cells: Differential Regulation of RsmY/Z Small RNAs in Pseudomonas aeruginosa by the Gac/HptB System

Two-component systems are capable of profoundly affecting genetic regulation in bacteria by detecting environmental stimuli, allowing them to quickly adapt. In Pseudomonas aeruginosa, the small RNAs (sRNAs) RsmY and RsmZ are under the control of the GacS/A system. They have been described as ones of the major key players in the control of planktonic and surface-associated behaviors. Genetic regulation by these sRNAs is achieved by the titration of the negative post-transcriptional regulator RsmA which affects the expression of over 500 genes. There is increasing evidence pinpointing the importance of RsmY and RsmZ in the planktonic-sessile P. aeruginosa lifestyles switch control. Using swarming motility as a model, we show here that these sRNA are differentially regulated depending on the selected growth conditions (i.e., planktonic versus surface grown-cells). Also, we report that opposite to planktonically grown cells, rsmZ regulation does not implicate the response regulator GacA in swarming cells. Furthermore, we present data indicating that RsmY/Z expression influence swarming motility via the protein HptB which acts as a negative regulator of these sRNAs and that they do not strictly converge to RsmA as previously reported.