Project description:Thiovulum spp. (Campylobacterota) are large sulfur bacteria that form veil-like structures in aquatic environments. The sulfidic Movile Cave (Romania), sealed from the atmosphere for ~5 million years, has several aqueous chambers, some with low atmospheric O2 (~7%). The cave's surface-water microbial community is dominated by bacteria we identified as Thiovulum. We show that this strain, and others from subsurface environments, are phylogenetically distinct from marine Thiovulum. We assembled a closed genome of the Movile strain and confirmed its metabolism using RNAseq. We compared the genome of this strain and one we assembled from public data from the sulfidic Frasassi caves to four marine genomes, including Candidatus Thiovulum karukerense and Ca. T. imperiosus, whose genomes we sequenced. Despite great spatial and temporal separation, the genomes of the Movile and Frasassi Thiovulum were highly similar, differing greatly from the very diverse marine strains. We concluded that cave Thiovulum represent a new species, named here Candidatus Thiovulum stygium. Based on their genomes, cave Thiovulum can switch between aerobic and anaerobic sulfide oxidation using O2 and NO3- as electron acceptors, the latter likely via dissimilatory nitrate reduction to ammonia. Thus, Thiovulum is likely important to both S and N cycles in sulfidic caves. Electron microscopy analysis suggests that at least some of the short peritrichous structures typical of Thiovulum are type IV pili, for which genes were found in all strains. These pili may play a role in veil formation, by connecting adjacent cells, and in the motility of these exceptionally fast swimmers.
Project description:We determined a significant fraction of the genome sequence of a representative of Thiovulum, the uncultivated genus of colorless sulfur Epsilonproteobacteria, by analyzing the genome sequences of four individual cells collected from phototrophic mats from Elkhorn Slough, California. These cells were isolated utilizing a microfluidic laser-tweezing system, and their genomes were amplified by multiple-displacement amplification prior to sequencing. Thiovulum is a gradient bacterium found at oxic-anoxic marine interfaces and noted for its distinctive morphology and rapid swimming motility. The genomic sequences of the four individual cells were assembled into a composite genome consisting of 221 contigs covering 2.083 Mb including 2,162 genes. This single-cell genome represents a genomic view of the physiological capabilities of isolated Thiovulum cells. Thiovulum is the second-fastest bacterium ever observed, swimming at 615 ?m/s, and this genome shows that this rapid swimming motility is a result of a standard flagellar machinery that has been extensively characterized in other bacteria. This suggests that standard flagella are capable of propelling bacterial cells at speeds much faster than typically thought. Analysis of the genome suggests that naturally occurring Thiovulum populations are more diverse than previously recognized and that studies performed in the past probably address a wide range of unrecognized genotypic and phenotypic diversities of Thiovulum. The genome presented in this article provides a basis for future isolation-independent studies of Thiovulum, where single-cell and metagenomic tools can be used to differentiate between different Thiovulum genotypes.
Project description:Primary objectives: The primary objective is to investigate circulating tumor DNA (ctDNA) via deep sequencing for mutation detection and by whole genome sequencing for copy number analyses before start (baseline) with regorafenib and at defined time points during administration of regorafenib for treatment efficacy in colorectal cancer patients in terms of overall survival (OS).
Primary endpoints: circulating tumor DNA (ctDNA) via deep sequencing for mutation detection and by whole genome sequencing for copy number analyses before start (baseline) with regorafenib and at defined time points during administration of regorafenib for treatment efficacy in colorectal cancer patients in terms of overall survival (OS).
Project description:The ecology and dynamics of many microbial systems, particularly in mats and soils, are shaped by how bacteria respond to evolving nutrient gradients and microenvironments. Here we show how the response of the sulfur-oxidizing bacterium Thiovulum majus to changing oxygen gradients causes cells to organize into large-scale fronts. To study this phenomenon, we develop a technique to isolate and enrich these bacteria from the environment. Using this enrichment culture, we observe the formation and dynamics of T. majus fronts in oxygen gradients. We show that these dynamics can be understood as occurring in two steps. First, chemotactic cells moving up the oxygen gradient form a front that propagates with constant velocity. We then show, through observation and mathematical analysis, that this front becomes unstable to changes in cell density. Random perturbations in cell density create oxygen gradients. The response of cells magnifies these gradients and leads to the formation of millimeter-scale fluid flows that actively pull oxygenated water through the front. We argue that this flow results from a nonlinear instability excited by stochastic fluctuations in the density of cells. Finally, we show that the dynamics by which these modes interact can be understood from the chemotactic response of cells. These results provide a mathematically tractable example of how collective phenomena in ecological systems can arise from the individual response of cells to a shared resource.
Project description:The naked mole-rat (NMR; Heterocephalus glaber) has recently gained considerable attention in the scientific community for its unique potential to unveil novel insights in the fields of medicine, biochemistry, and evolution. NMRs exhibit unique adaptations that include protracted fertility, cancer resistance, eusociality, and anoxia. This suite of adaptations is not found in other rodent species, suggesting that interrogating conserved and accelerated regions in the NMR genome will find regions of the NMR genome fundamental to their unique adaptations. However, the current NMR genome assembly has limits that make studying structural variations, heterozygosity, and non-coding adaptations challenging. We present a complete diploid naked-mole rat genome assembly by integrating long-read and 10X-linked read genome sequencing of a male NMR and its parents, and Hi-C sequencing in the NMR hypothalamus (N=2). Reads were identified as maternal, paternal or ambiguous (TrioCanu). We then polished genomes with Flye, Racon and Medaka. Assemblies were then scaffolded using the following tools in order: Scaff10X, Salsa2, 3d-DNA, Minimap2-alignment between assemblies, and the Juicebox Assembly Tools. We then subjected the assemblies to another round of polishing, including short-read polishing with Freebayes. We assembled the NMR mitochondrial genome with mitoVGP. Y chromosome contigs were identified by aligning male and female 10X linked reads to the paternal genome and finding male-biased contigs not present in the maternal genome. Contigs were assembled with publicly available male NMR Fibroblast Hi-C-seq data (SRR820318). Both assemblies have their sex chromosome haplotypes merged so that both assemblies have a high-quality X and Y chromosome. Finally, assemblies were evaluated with Quast, BUSCO, and Merqury, which all reported the base-pair quality and contiguity of both assemblies as high-quality. The assembly will next be annotated by Ensembl using public RNA-seq data from multiple tissues (SRP061363). Together, this assembly will provide a high-quality resource to the NMR and comparative genomics communities.
Project description:The naked mole-rat (NMR; Heterocephalus glaber) has recently gained considerable attention in the scientific community for its unique potential to unveil novel insights in the fields of medicine, biochemistry, and evolution. NMRs exhibit unique adaptations that include protracted fertility, cancer resistance, eusociality, and anoxia. This suite of adaptations is not found in other rodent species, suggesting that interrogating conserved and accelerated regions in the NMR genome will find regions of the NMR genome fundamental to their unique adaptations. However, the current NMR genome assembly has limits that make studying structural variations, heterozygosity, and non-coding adaptations challenging. We present a complete diploid naked-mole rat genome assembly by integrating long-read and 10X-linked read genome sequencing of a male NMR and its parents, and Hi-C sequencing in the NMR hypothalamus (N=2). Reads were identified as maternal, paternal or ambiguous (TrioCanu). We then polished genomes with Flye, Racon and Medaka. Assemblies were then scaffolded using the following tools in order: Scaff10X, Salsa2, 3d-DNA, Minimap2-alignment between assemblies, and the Juicebox Assembly Tools. We then subjected the assemblies to another round of polishing, including short-read polishing with Freebayes. We assembled the NMR mitochondrial genome with mitoVGP. Y chromosome contigs were identified by aligning male and female 10X linked reads to the paternal genome and finding male-biased contigs not present in the maternal genome. Contigs were assembled with publicly available male NMR Fibroblast Hi-C-seq data (SRR820318). Both assemblies have their sex chromosome haplotypes merged so that both assemblies have a high-quality X and Y chromosome. Finally, assemblies were evaluated with Quast, BUSCO, and Merqury, which all reported the base-pair quality and contiguity of both assemblies as high-quality. The assembly will next be annotated by Ensembl using public RNA-seq data from multiple tissues (SRP061363). Together, this assembly will provide a high-quality resource to the NMR and comparative genomics communities.