Tracing Lifestyle Adaptation in Prokaryotic Genomes

Lifestyle adaptation of microbes due to changes in their ecological niches or acquisition of new environments is a major driving force for genetic changes in their respective genomes. Moving into more specialized niches often results in the acquisition of new gene sets via horizontal gene transfer to utilize previously unavailable metabolites, while genetic ballast is shed by gene loss and/or gene inactivation. In some cases, larger genome rearrangements can be observed, such as the incorporation of whole genetic islands, providing a range of new phenotypic capabilities. Until recently these changes could not be comprehensively followed and identified due to the lack of complete microbial genome sequences. The advent of high-throughput DNA sequencing has dramatically changed the scientific landscape and today microbial genomes have become increasingly abundant. Currently, more than 2,900 genomes are published and more than 11,000 genome projects are listed in the Genomes Online Database‡. Although this wealth of information provides many new opportunities to assess microbial functionality, it also creates a new array of challenges when a comparison between multiple microbial genomes is required. Here, functional genome distribution (FGD) is introduced, analyzing the diversity between microbes based on their predicted ORFeome. FGD is therefore a comparative genomics approach, emphasizing the assessments of gene complements. To further facilitate the comparison between two or more genomes, degrees of amino-acid similarities between ORFeomes can be visualized in the Artemis comparison tool, graphically depicting small and large scale genome rearrangements, insertion and deletion events, and levels of similarity between individual open reading frames. FGD provides a new tool for comparative microbial genomics and the interpretation of differences in the genetic makeup of bacteria

ViromeScan: a new tool for metagenomic viral community profiling

BACKGROUND: Bioinformatics tools available for metagenomic sequencing analysis are principally devoted to the identification of microorganisms populating an ecological niche, but they usually do not consider viruses. Only some software have been designed to profile the viral sequences, however they are not efficient in the characterization of viruses in the context of complex communities, like the intestinal microbiota, containing bacteria, archeabacteria, eukaryotic microorganisms and viruses. In any case, a comprehensive description of the host-microbiota interactions can not ignore the profile of eukaryotic viruses within the virome, as viruses are definitely critical for the regulation of the host immunophenotype. RESULTS: ViromeScan is an innovative metagenomic analysis tool that characterizes the taxonomy of the virome directly from raw data of next-generation sequencing. The tool uses hierarchical databases for eukaryotic viruses to unambiguously assign reads to viral species more accurately and >1000 fold faster than other existing approaches. We validated ViromeScan on synthetic microbial communities and applied it on metagenomic samples of the Human Microbiome Project, providing a sensitive eukaryotic virome profiling of different human body sites. CONCLUSIONS: ViromeScan allows the user to explore and taxonomically characterize the virome from metagenomic reads, efficiently denoising samples from reads of other microorganisms. This implies that users can fully characterize the microbiome, including bacteria and viruses, by shotgun metagenomic sequencing followed by different bioinformatic pipelines. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1186/s12864-016-2446-3) contains supplementary material, which is available to authorized users

Homology inference with specific molecular constraints

Evolutionary processes can be considered at multiple levels of biological organization. The work developed in this thesis focuses on protein molecular evolution. Although proteins are linear polymers composed from a basic set of 20 amino acids, they generate an enormous variety of form and function. Proteins that have arisen by a common descent are classified into families; they often share common properties including similarities in sequence, structure, and function. Multiple methods have been developed to infer evolutionary relationships between proteins and classify them into families. Yet, those generic methods are often inaccurate, especially when specific protein properties limit their applications. In this thesis, we analyse two protein classes that are often difficult for the evolutionary analysis: the coiled-coils – repetitive protein domains defined by a simple widespread peptide motif (chapters 2 and 3) and Rab small GTPases – a large family of closely related proteins (chapters 4 and 5). In both cases, we analyse the specific properties that determine protein structure and function and use them to improve their evolutionary inference

Singled Out: Genomic analysis of uncultured microbes in marine sediments

The vast majority of abundant taxa in marine sediment environments have not yielded to culture, leaving questions about their relationship to other taxa and their functional potential unanswered. However, in the absence of active cultures, careful application of various omics methods can be used to help us make useful inferences about their evolutionary history and how they have continued to survive in environments of extreme energy deprivation. For this dissertation, I have applied comparative genomics methods to members of two uncultured groups, the recently proposed Altiarchaeales order and a cosmopolitan taxon associated with the Actinobacteria phylum. Additionally, I combined transcript recruitment and metabolomic profiles to investigate metabolisms inferred from the single-cell amplified genomes extracted from members of a taxa that thrive in Baltic Sea sediment microbial communities. In Chapter II, I establish a phylogenetic relationship across distantly related members of the order Altiarchaeales and discuss environment-specific adaptations. In Chapter III, transcript recruitment and metabolite profiles support a community-wide focus on microbial persistence with active members of the uncultured Atribacteria phylum playing an important ecological role. In Chapter IV, my analysis leads to the proposal of the new class within the Actinobacteria. Osirisbacteria is a class of Actinobacteria that is specialized for life in anoxic environments. Overall, this work offers new insights into deeply-branching microbial taxa, improved understanding of recently considered branches of the evolutionary tree, and new perspective on metabolisms important for survival in low-energy marine sediment environments

Sulfate reducing communities in aquifer systems can be reliably stimulated by addition of complex nutrients

The disseration presented below is the summation of research into the potential roles of microbial communities associated with aquifers of Bangladesh contaminated with naturally occuring arsenic. These investigations also included experimental microcosm experiments to assess the role of nutrients supplementation of complex carbon sources (molasses), and inorganic sulfate (MgSO4), on both the solubility of arsenic to determine the feasibility of this method for the goal of performing in situ bioremediation. Community structure and functional gene profiling was performed on all samples, as well as detection of community shifts following amendments predicted to encourage the growth of sulfate reducting microorganisms (SRM). This included community profiling via 16S analysis, as well as presence and quantification of a number of genes involved in respiratory sulfate reduction and genes involved in arsenic cycling. Investigation of samples gathered from contaminated aquifers seems to indicate that even in a community with relatively simple distribution of organisms, there is no distinct linkage between examined functional genes and concentrations of any detected elements in the aquifers. Examination of the effects of nutrient supplementation on sediments gathered from one impacted aquifer shows that stimulation of the system with either nutrient tested is sufficient to stimulate growth of sulfate reducing microbes, as indicated by conserved genes in the respiratory sulfate reduction pathway. These shifts can be closely associated with an initial decrease in detectable soluble arsenic levels, as well as a commensurate decrease in soluble metals. However, only the addition of both a complex carbon source and magnesium sulfate in equal molar portions seemed to show prolonged removal of these elements from the soluble phase. Community shifts appear to have occurred by 14 days of incubation, and were coupled with expected changes in the color and consistency of sediment as black particulate can serve as an indicator of sulfidic minerals formed as a result of excess sulfides produced by SRM. Increased SRM numbers were maintained through 96 days of incubation. Due to the ability of any perturbation of a microcosm system to produce increased density of SRM in the samples, a bioinformatic investigation of the identified subsystems encoded by all sequenced and finished bacteria capable of carrying out the most conserved steps in sulfate reduction was performed. These analyses indicated that there are a number of SRM capable of directly reducing complex carbon sources, both in syntrophic communities, as well as without additional aid from the environment. These results indicate that sulfate reducing microbes are present, detectable and easily stimulated to grow in aquifer sediment, and that these communities of SRM are able to create conditions capable of removing arsenic from the soluble phase. The rate of growth and ability to maintain this immobilization supports the theory that SRM detected in the environment are capable of growth on complex nutrients, and require additional nutrients to successfully remediate arsenic for long periods of time

Biogeographic patterns and drivers of soil viromes

Viruses are crucial in shaping soil microbial functions and ecosystems. However, studies on soil viromes have been limited in both spatial scale and biome coverage. Here we present a comprehensive synthesis of soil virome biogeographic patterns using the Global Soil Virome dataset (GSV) wherein we analysed 1,824 soil metagenomes worldwide, uncovering 80,750 partial genomes of DNA viruses, 96.7% of which are taxonomically unassigned. The biogeography of soil viral diversity and community structure varies across different biomes. Interestingly, the diversity of viruses does not align with microbial diversity and contrasts with it by showing low diversity in forest and shrubland soils. Soil texture and moisture conditions are further corroborated as key factors affecting diversity by our predicted soil viral diversity atlas, revealing higher diversity in humid and subhumid regions. In addition, the binomial degree distribution pattern suggests a random co-occurrence pattern of soil viruses. These findings are essential for elucidating soil viral ecology and for the comprehensive incorporation of viruses into soil ecosystem models

The computational analysis of post-translational modifications

The post-translational modification (PTMs) of proteins presents a means to increase the proteome size and diversity of an organism through the inclusion of structural elements not encoded at the sequence-level alone. Their erroneous inclusion or exclusion has been linked to a variety of diseases and disorders thus their characterisation has the potential to present viable drug targets. The proliferation of newer high-throughput methods, such as mass spectrometry, to identify such modifications has led to a rapid increase in the number of databases and tools to display and analyse such vast amounts of data effectively. This study covers the development of one such tool; PTM Browser, and the construction of the underlying database that it is based upon. This new database was initially seeded with annotations from the Swiss-Prot and Phospho.ELM resources. The initial database of PTMs was then expanded to include a large repertoire of previously unannotated proteins for a selection of topical species (e.g. Danio rerio and Tetraodon nigroviridis). Orthologue assignments have also been added to the database – to allow for queries to be performed regarding the conservation of modifications between homologous proteins. The PTM Browser tool allows for a full exploration of this new database of PTMs – with a special focus on allowing users to identify modifications that are both shared between and are specific to particular species. This tool is freely available for non-commercial use at the following URL: http://www.ptmbrowser.org. An analysis is presented on the conservation of modifications between members of the tumour suppressor family, p53, using this new tool. This tool has also been used to analysis the conservation of modifications between super-kingdoms and Eukaryote species