Eurythermalism of a deep-sea symbiosis system from an enzymological aspect

The recently proposed and experimentally validated Equilibrium Model provides the most detailed description of temperature's effect on enzyme catalytic activity to date. By introducing an equilibrium between Eact, the active form of enzyme, and Einact, a reversibly inactivated form of enzyme, the Equilibrium Model explains apparent enzyme activity loss at high temperatures that cannot be accounted for by irreversible thermal denaturation. The Equilibrium Model describes enzyme behavior in the presence of substrates and under assay conditions; thus its associated parameters, deltaHeq and Teq, may have physiological significance. The Equilibrium Model parameters have been determined for twenty-one enzymes of diverse origins. The results demonstrated the wide applicability of the Equilibrium Model to enzymes of different types and temperature affinity. The study has also established deltaHeq as the first quantitative measure of enzyme eurythermalism and demonstrated the relationship between Teq and optimal growth temperature of organisms. The Equilibrium Model is therefore a useful tool for studying enzyme temperature adaptation and its role in adaptations to thermophily and eurythermalism. Moreover, it potentially enables a description of the originating environment from the properties of the enzymes. The Equilibrium Model has been employed to characterize enzymes isolated from bacterial episymbionts of Alvinella pompejana. A. pompejana inhabits one of the most extreme environments known to science and has been proposed as an extremely eurythermal organism. A metagenomic study of the A. pompejana episymbionts has unveiled new information related to the adaptive and metabolic properties of the bacterial consortium; the availability of metagenomic sequences has also enabled targeted retrieval and heterologous expression of A. pompejana episymbiont genes. By inspecting enzymes derived from the unique episymbiotic microbial consortium intimately associated with A. pompejana, the study has shed light on temperature adaptations in this unique symbiotic relationship. The findings suggested that eurythermal enzymes are one of the mechanisms used by the microbial consortium to achieve its adaptations. By combining metagenomic and enzymological studies, the research described in this thesis has lead to insights on the eurythermalism of a complex microbial system from an enzymological aspect. The findings have enhanced our knowledge on how life adapts to extreme environments, and the validation of the Equilibrium Model as a tool for studying enzyme temperature adaptation paves the way for future studies

Evidence of global-scale aeolian dispersal and endemism in isolated geothermal microbial communities of Antarctica

New evidence in aerobiology challenges the assumption that geographical isolation is an effective barrier to microbial transport. However, given the uncertainty with which aerobiological organisms are recruited into existing communities, the ultimate impact of microbial dispersal is difficult to assess. To evaluate the ecological significance of global-scale microbial dispersal, molecular genetic approaches were used to examine microbial communities inhabiting fumarolic soils on Mt. Erebus, the southernmost geothermal site on Earth. There, hot, fumarolic soils provide an effective environmental filter to test the viability of organisms that have been distributed via aeolian transport over geological time. We find that cosmopolitan thermophiles dominate the surface, whereas endemic Archaea and members of poorly understood Bacterial candidate divisions dominate the immediate subsurface. These results imply that aeolian processes readily disperse viable organisms globally, where they are incorporated into pre-existing complex communities of endemic and cosmopolitan taxa

The equilibrium model for the effect of temperature on enzymes: Insights and implications

A new, experimentally-validated “Equilibrium Model” describes the effect of temperature on enzymes, and provides a new mechanism for the reversible loss of enzyme activity with temperature. It incorporates two new, fundamental parameters that allow a complete description of the effect of temperature on enzyme activity: ΔHeq and Teq. ΔHeq emerges as an intrinsic and quantitative measure of enzyme eurythermal adaptation, while Teq, the equilibrium temperature, has fundamental and technological significance for our understanding of the effect of temperature on enzymatic reactions. For biotechnological purposes, these parameters need to be considered when enzymes are applied or engineered for activity at high temperatures

Benthic microbial communities of coastal terrestrial and ice shelf Antarctic meltwater ponds.

The numerous perennial meltwater ponds distributed throughout Antarctica represent diverse and productive ecosystems central to the ecological functioning of the surrounding ultra oligotrophic environment. The dominant taxa in the pond benthic communities have been well described however, little is known regarding their regional dispersal and local drivers to community structure. The benthic microbial communities of 12 meltwater ponds in the McMurdo Sound of Antarctica were investigated to examine variation between pond microbial communities and their biogeography. Geochemically comparable but geomorphologically distinct ponds were selected from Bratina Island (ice shelf) and Miers Valley (terrestrial) (<40 km between study sites), and community structure within ponds was compared using DNA fingerprinting and pyrosequencing of 16S rRNA gene amplicons. More than 85% of total sequence reads were shared between pooled benthic communities at different locations (OTU0.05), which in combination with favorable prevailing winds suggests aeolian regional distribution. Consistent with previous findings Proteobacteria and Bacteroidetes were the dominant phyla representing over 50% of total sequences; however, a large number of other phyla (21) were also detected in this ecosystem. Although dominant Bacteria were ubiquitous between ponds, site and local selection resulted in heterogeneous community structures and with more than 45% of diversity being pond specific. Potassium was identified as the most significant contributing factor to the cosmopolitan community structure and aluminum to the location unique community based on a BEST analysis (Spearman's correlation coefficient of 0.632 and 0.806, respectively). These results indicate that the microbial communities in meltwater ponds are easily dispersed regionally and that the local geochemical environment drives the ponds community structure

Influence of soil properties on archaeal diversity and distribution in the McMurdo Dry Valleys, Antarctica

Archaea are the least understood members of the microbial community in Antarctic mineral soils. Although their occurrence in Antarctic coastal soils has been previously documented, little is known about their distribution in soils across the McMurdo Dry Valleys, Victoria Land. In this study, terminal-restriction fragment length polymorphism (t-RFLP) analysis and 454 pyrosequencing were coupled with a detailed analysis of soil physicochemical properties to characterize archaeal diversity and identify environmental factors that might shape and maintain archaeal communities in soils of the three southern most McMurdo Dry Valleys (Garwood, Marshall, and Miers Valley). Archaea were successfully detected in all inland and coastal mineral soils tested, revealing a low overall richness (mean of six operational taxonomic units [OTUs] per sample site). However, OTU richness was higher in some soils and this higher richness was positively correlated with soil water content, indicating water as a main driver of archaeal community richness. In total, 18 archaeal OTUs were detected, predominately Thaumarchaeota affiliated with Marine Group 1.1b (> 80% of all archaeal sequences recovered). Less abundant OTUs (2% of all archaeal sequences) were loosely related to members of the phylum Euryarchaeota. This is the first comprehensive study showing a widespread presence and distribution of Archaea in inland Antarctic soils

Diffuse flow environments within basalt- and sediment-based hydrothermal vent ecosystems harbor specialized microbial communities

Hydrothermal vents differ both in surface input and subsurface geochemistry. The effects of these differences on their microbial communities are not clear. Here, we investigated both alpha and beta diversity of diffuse flow-associated microbial communities emanating from vents at a basalt-based hydrothermal system along the East Pacific Rise (EPR) and a sediment-based hydrothermal system, Guaymas Basin. Both Bacteria and Archaea were targeted using high throughput 16S rRNA gene pyrosequencing analyses. A unique aspect of this study was the use of a universal set of 16S rRNA gene primers to characterize total and diffuse flow-specific microbial communities from varied deep-sea hydrothermal environments. Both surrounding seawater and diffuse flow water samples contained large numbers of Marine Group I (MGI) Thaumarchaea and Gammaproteobacteria taxa previously observed in deep-sea systems. However, these taxa were geographically distinct and segregated according to type of spreading center. Diffuse flow microbial community profiles were highly differentiated. In particular, EPR dominant diffuse flow taxa were most closely associated with chemolithoautotrophs, and off axis water was dominated by heterotrophic-related taxa, whereas the opposite was true for Guaymas Basin. The diversity and richness of diffuse flow-specific microbial communities were strongly correlated to the relative abundance of Epsilonproteobacteria, proximity to macrofauna, and hydrothermal system type. Archaeal diversity was higher than or equivalent to bacterial diversity in about one third of the samples. Most diffuse flow-specific communities were dominated by OTUs associated with Epsilonproteobacteria, but many of the Guaymas Basin diffuse flow samples were dominated by either OTUs within the Planctomycetes or hyperthermophilic Archaea. This study emphasizes the unique microbial communities associated with geochemically and geographically distinct hydrothermal diffuse flow environments

Groundtruthing next-gen sequencing for microbial ecology-biases and errors in community structure estimates from PCR amplicon pyrosequencing

Analysis of microbial communities by high-throughput pyrosequencing of SSU rRNA gene PCR amplicons has transformed microbial ecology research and led to the observation that many communities contain a diverse assortment of rare taxa-a phenomenon termed the Rare Biosphere. Multiple studies have investigated the effect of pyrosequencing read quality on operational taxonomic unit (OTU) richness for contrived communities, yet there is limited information on the fidelity of community structure estimates obtained through this approach. Given that PCR biases are widely recognized, and further unknown biases may arise from the sequencing process itself, a priori assumptions about the neutrality of the data generation process are at best unvalidated. Furthermore, post-sequencing quality control algorithms have not been explicitly evaluated for the accuracy of recovered representative sequences and its impact on downstream analyses, reducing useful discussion on pyrosequencing reads to their diversity and abundances. Here we report on community structures and sequences recovered for in vitro-simulated communities consisting of twenty 16S rRNA gene clones tiered at known proportions. PCR amplicon libraries of the V3-V4 and V6 hypervariable regions from the in vitro-simulated communities were sequenced using the Roche 454 GS FLX Titanium platform. Commonly used quality control protocols resulted in the formation of OTUs with >1% abundance composed entirely of erroneous sequences, while over-aggressive clustering approaches obfuscated real, expected OTUs. The pyrosequencing process itself did not appear to impose significant biases on overall community structure estimates, although the detection limit for rare taxa may be affected by PCR amplicon size and quality control approach employed. Meanwhile, PCR biases associated with the initial amplicon generation may impose greater distortions in the observed community structure

A core phyllosphere microbiome exists across distant populations of a tree species indigenous to New Zealand

The phyllosphere microbiome is increasingly recognised as an influential component of plant physiology, yet it remains unclear whether stable host-microbe associations generally exist in the phyllosphere. Leptospermum scoparium (mānuka) is a tea tree indigenous to New Zealand, and honey derived from mānuka is widely known to possess unique antimicrobial properties. However, the host physiological traits associated with these antimicrobial properties vary widely, and the specific cause of such variation has eluded scientists despite decades of research. Notably, the mānuka phyllosphere microbiome remains uncharacterised, and its potential role in mediating host physiology has not been considered. Working within the prevailing core microbiome conceptual framework, we hypothesise that the phyllosphere microbiome of mānuka exhibits specific host association patterns congruent with those of a microbial community under host selective pressure (null hypothesis: the mānuka phyllosphere microbiome is recruited stochastically from the surrounding environment). To examine our hypothesis, we characterised the phyllosphere and associated soil microbiomes of five distinct and geographically distant mānuka populations across the North Island of New Zealand. We identified a habitat-specific and relatively abundant core microbiome in the mānuka phyllosphere, which was persistent across all samples. In contrast, non-core phyllosphere microorganisms exhibited significant variation across individual host trees and populations that was strongly driven by environmental and spatial factors. Our results demonstrate the existence of a dominant and ubiquitous core microbiome in the phyllosphere of mānuka, supporting our hypothesis that phyllosphere microorganisms of mānuka exhibit specific host association and potentially mediate physiological traits of this nationally and culturally treasured indigenous plant. In addition, our results illustrate biogeographical patterns in mānuka phyllosphere microbiomes and offer insight into factors contributing to phyllosphere microbiome assembly

Evidence of plant and animal communities at exposed and subglacial (cave) geothermal sites in Antarctica

Geothermal areas, such as volcanoes, might have acted as glacial microrefugia for a wide range of species. The heavily glaciated but volcanically active Antarctic continent presents an ideal system for assessing this hypothesis. Ice-free terrain around volcanoes in Antarctica is, however, often restricted to small patches, whereas subglacial cave systems, formed by vented volcanic steam, can be extensive and interconnected. No observations of macrobiota have yet been made for subglacial geothermal environments in Antarctica, but these organisms are often patchily distributed and can be difficult to find. We carried out metabarcoding (eDNA) analyses of soil samples taken from exposed areas on three volcanoes in Victoria Land, and subglacial caves on Mount Erebus. We found evidence of numerous eukaryotic groups, including mosses, algae, arthropods, oligochaetes and nematodes, at both exposed and subglacial sites. Our findings support the notion that geothermal areas—including subglacial environments—can nurture biodiversity in glaciated regions

A pilot randomised controlled trial of oral doxycycline after endoscopic sinus surgery and its effects on the sinonasal microbiome

Oral antibiotics are commonly prescribed after endoscopic sinus surgery (ESS) despite minimal clinical data supporting this practice. We aim to assess the effect of post-ESS doxycycline on clinical outcomes and on the diversity and composition of the sinonasal microbiome. Methods: Samples from the middle meatus were collected from twelve patients undergoing ESS to treat chronic rhinosinusitis. Patients were double-blind randomised to receive either oral doxycycline or placebo in the post-operative period. Further samples were collected at two weeks and three months post-operatively. The sinonasal microbiome was characterized using 16S ribosomal RNA (rRNA) gene amplicon sequencing. SNOT-22 scores, Lund Mackay scores, and Modified Lund Mackay Endoscopic Scores (MLMES) were collected. Results: After ESS, bacterial diversity increased while SNOT-22 score decreased for both treatments. Microbiome composition diverged between treatments, and random forest analysis identified nine taxa that may distinguish treatment groups. There was no significant difference in SNOT-22 score, 3-month MLMES or bacterial diversity between the placebo and doxycycline groups. The trends for all of these measures favour placebo. Conclusion: In this pilot study, we detected no significant difference between placebo and antibiotic treatments in clinical outcome. As patient symptoms improved after ESS, we detected a concurrent increase in the diversity of the sinonasal microbiome. Our data highlight the need for and facilitate the design of future larger studies to explore the relationship between prophylactic antibiotics and post-ESS recovery