The compositional and evolutionary logic of metabolism

Metabolism displays striking and robust regularities in the forms of modularity and hierarchy, whose composition may be compactly described. This renders metabolic architecture comprehensible as a system, and suggests the order in which layers of that system emerged. Metabolism also serves as the foundation in other hierarchies, at least up to cellular integration including bioenergetics and molecular replication, and trophic ecology. The recapitulation of patterns first seen in metabolism, in these higher levels, suggests metabolism as a source of causation or constraint on many forms of organization in the biosphere. We identify as modules widely reused subsets of chemicals, reactions, or functions, each with a conserved internal structure. At the small molecule substrate level, module boundaries are generally associated with the most complex reaction mechanisms and the most conserved enzymes. Cofactors form a structurally and functionally distinctive control layer over the small-molecule substrate. Complex cofactors are often used at module boundaries of the substrate level, while simpler ones participate in widely used reactions. Cofactor functions thus act as "keys" that incorporate classes of organic reactions within biochemistry. The same modules that organize the compositional diversity of metabolism are argued to have governed long-term evolution. Early evolution of core metabolism, especially carbon-fixation, appears to have required few innovations among a small number of conserved modules, to produce adaptations to simple biogeochemical changes of environment. We demonstrate these features of metabolism at several levels of hierarchy, beginning with the small-molecule substrate and network architecture, continuing with cofactors and key conserved reactions, and culminating in the aggregation of multiple diverse physical and biochemical processes in cells.Comment: 56 pages, 28 figure

Phylogenomic analysis of metabotrophic P2Y receptor family and its expression in zebrafish, Danio rerio

G-protein-coupled receptors (GPCRs) constitute the largest and most divergent class of cell surface proteins. GPCRs can be activated by extracellular signals as diverse as light, peptides, proteins, lipids, odorants, tastants, nucleotides and nucleosides. Currently, all the known chemosensory receptor genes, such as odorant, taste and pheromone receptors belong to the GPCR family, with many of them being class A or class-A related genes. Metabotropic P2Y receptors belong to class A GPCRs and are activated by extracellular nucleotides. Nucleotides are released to the environment when the organism is injured and therefore serve as one of the food stimuli. As such, it would be interesting to see if P2Y receptors play any role in olfaction in zebrafish. To date, eight functional human P2Y receptors and 25 related orphan receptors have been found. I performed extensive data mining in drosophila, ascidian, jawless, cartilaginous and bony fish, frog and human genomes to delineate the P2Y family and investigate its evolutionary origin. The P2Y family originates early in the vertebrate lineage, reflected by the presence of lamprey and the absence of ascidian orthologues. Consistent with these findings, no P2Y receptor is found in invertebrates. In total, 38 subfamilies can be distinguished within the P2Y family, at least two third of which are already present in the shark genome. Two subfamilies, p2yl-3 and p2yl-4, are lost in the human lineage and only GPR 87 subfamily is lost in all teleost species. Zebrafish has 68 P2Y receptor genes, the most of any fish species, and almost double as many than mammals. The teleost P2Y genes are widely distributed in the genome as small cluster and singletons. The vast majority of P2Y genes are intronless while the remaining genes contain up to five introns. In the teleost lineage, the genomic arrangement of P2Y genes is preserved to a large extent, and some synteny is found even with the elephant shark and human genome, possibly reflecting the functional importance of these genes. Selective pressure on teleost P2Y genes generally is high, as evidenced by a preponderance of negative selection. However, a few genes exhibit positive selection at individual sites. In early development, P2Y genes are expressed in many tissues and organs, notably the central and peripheral nervous system, pharyngeal arches, otic vesicle and kidney, suggesting an important role in the development of many tissues. However, no expression is detected in larval olfactory epithelium. In contrast, olfactory epithelium of adult fish does express several P2Y genes as shown by RT-PCR. A possible explanation would be a late onset of expression in the olfactory epithelium. In situ hybridisation of adult olfactory epithelium established an ubiquitous distribution, both in the sensory and non-sensory region, which seems to argue against a role of P2Y genes in nucleotide odor detection. However, further studies will be necessary to give a definitive answer to that question

Comparative microbial genome analysis of lactobacilli

The genus Lactobacillus is a diverse group with a combined species count of over 200. They are the largest group within the lactic acid bacteria and one of the most important bacterial groups involved in food microbiology and human nutrition because of their fermentative and probiotic properties. Exploiting lactobacilli has been complicated by metabolic diversity, unclear species identity and uncertain relationships between them and other commercially important lactic acid bacteria. Comparative Genomics is becoming increasingly important as a method of investigating the diversity of function and phylogeny of all microbial taxa, including Lactobacillus. In Chapter 1, I give an overview of relevant concepts in comparative microbial genomics, focusing on relevant literature for lactobacilli and related genera. In Chapter 2, we (Sun et al. 2015) carried out comparative genomic analyses of type strains of 213 Lactobacillus strains and associated genera, describing their encoded genetic catalogue for modifying carbohydrates and proteins. In addition, we describe a broad and diverse presence of CRISPR-Cas immune systems, host interaction factors and bacteriocins that affect their natural and industrial environments, and mechanisms to withstand stress during technological processes. We present a robust phylogenomic framework of existing species and for classifying new species. In Chapter 3, we (Harris et al. 2017) we carried out comparative genomic analyses of the functional and phylogenomic diversity of 42 genomes of strains of L. salivarius. We found that multiple phylogenomic and non-phylogenomic methods used for reconstructing trees all converge on similar tree topologies, showing that different metrics largely agree on the evolutionary history of the species. The greatest genomic variation lies on the small plasmids, followed by the repA-type circular megaplasmid, with the chromosome varying least of all. Glycosyl hydrolases, bacteriocins and proteases vary considerably on all replicons while two exopolysaccharide clusters show a lot of variation on the chromosome. In Chapter 4, I focussed on the evolutionary rates of lactobacilli, divided into 10 phylogenetic sub-clades based on an ongoing study (Salvetti et al; in prep). Several multiple alignment strategies were compared, and I showed that conclusions about purifying versus directional selection across subclades was influenced by alignment strategy. In Chapter 5, I give a general discussion of how my thesis contributes to current literature, including future perspectives in the comparative genomics of lactobacilli

An investigation of antimicrobial susceptibility and genotypes of Mycoplasma bovis isolates derived from western Canadian feedlot cattle

Mycoplasma bovis poses a significant threat to the Canadian beef industry, particularly the feedlot sector where it is associated with bovine respiratory disease (BRD) and chronic pneumonia and polyarthritis syndrome (CPPS). Furthermore, its characteristic chronic infections which are refractory to antimicrobials results in animal welfare and economic concerns. Currently, antimicrobials are the primary therapeutic option for treatment and control of M. bovis infections due the absence of a vaccine. This is salient given the global concern regarding antimicrobial use (AMU), antimicrobial resistance (AMR), and increased levels of AMR reported for M. bovis worldwide. Due to the current reliance on antimicrobials for prevention, control, and treatment, continual surveillance of antimicrobial susceptibility is crucial for antimicrobial stewardship and therapeutic treatment of M. bovis related disease. Not only is there a reduced arsenal of antimicrobials for the prevention, control and treatment of M. bovis, but its fastidious nature and difficulties associated with culturing emphasize a need for antimicrobial susceptibility testing (AST) using rapid, accurate, molecular methods. Whole genome sequencing (WGS) provides a plethora of information that can be interrogated to investigate different aspects of M. bovis pathogenesis, including antimicrobial susceptibility and virulence, such as multilocus sequence typing (MLST) for classifying bacterial strains. In conclusion, an epidemiological, genotypic and phenotypic investigation into the antimicrobial susceptibility of M. bovis will contribute to the body of knowledge needed to assist in evidence-based decisions for the treatment and control of M. bovis infections in cattle and provide the rationale for the studies outlined in this thesis. M. bovis isolates derived from western Canadian feedlot cattle sampled over a 12-year period (2006 - 2018) were used to address three objectives: 1), describe the AMU and AMR profiles of isolates derived from M. bovis mortalities in feedlot cattle (Chapter 2); 2), investigate the genotypic basis for macrolide resistance by assessing single nucleotide polymorphisms (SNPs) in 23S rRNA gene alleles and ribosomal proteins L4 and L22 (Chapter 3); and 3), assess the application of four genotyping methods for M. bovis (Chapter 4). M. bovis was cultured from deep nasopharyngeal swabs as well as lung and joint tissue from western Canadian feedlot cattle (cattle, n = 134; isolates, n = 183). Antimicrobial susceptibility testing (AST) was performed using a microbroth dilution assay and a customized panel of nine antimicrobials, representing four drug classes, most commonly administered to feedlot cattle in western Canada. Furthermore, M. bovis isolates (n = 129) underwent WGS utilizing Illumina technology. Although M. bovis isolates were derived from western Canadian feedlot cattle, Chapter 2 was not intended as a representative study of western Canadian feedlots but rather to provide context on AMR and AMU in feedlots and the background of the cattle and isolates represented in this thesis. In chapter 2, over 90% of cattle had received antimicrobial metaphylaxis, with tulathromycin accounting for 94.2 % of treatments. On average, cattle received three antimicrobial classes prior to dying of a mycoplasma-related pneumonia. The most commonly administered classes were macrolides (93.2%), phenicols (78.4%), and fluoroquinolones (67.6%). Isolates had the least resistance to florfenicol, with 89.9% classified as susceptible. Nearly all isolates were resistant to all five macrolides (gamithromycin, tildipirosin, tilmicosin, tulathromycin, tylosin) assessed. The study described in Chapter 3 found that mutations in both domains II and V of the 23S rRNA gene alleles were found to be associated with resistance to all five macrolides. Isolates with a mutation in domain II and the L4 and L22 ribosomal proteins were also resistant to all macrolides, except tulathromycin. Lastly, in Chapter 4, four in silico genotyping methods were applied to M. bovis isolates and the Simpson’s Diversity Index (D) was used to assess resolution of each method. MLST had the lowest resolution (D=0.932) but was the easiest to implement and apply; contrastingly, whole genome single nucleotide variant (wgSNV) yielded the highest resolution (D=1.000), but also involved the most complicated analysis. Application of core genome MLST (cgMLST) and core genome SNV (cgSNV) had a similar resolution of 0.987 and 0.984, respectively. No association between genotype and phenotype was resolved. Overall, M. bovis isolates in western Canadian feedlot cattle were predominately susceptible to a single antimicrobial, florfenicol, and commonly administered macrolides for BRD metaphylaxis. AMR was observed to all macrolides tested, and the accumulation of SNPs in genes associated with macrolide resistance correlated to a macrolide resistant phenotype using AST. Despite being unable to associate genotype and phenotype, the typing methods yielded comparable phylogenetic relationships. Furthermore, the diversity of strain types highlighted the structure of the Canadian cattle industry and how cattle are procured for western Canadian feedlots with some strain types being dominant over geographical areas and time

Ab initio machine learning in chemical compound space

Chemical compound space (CCS), the set of all theoretically conceivable combinations of chemical elements and (meta-)stable geometries that make up matter, is colossal. The first principles based virtual sampling of this space, for example in search of novel molecules or materials which exhibit desirable properties, is therefore prohibitive for all but the smallest sub-sets and simplest properties. We review studies aimed at tackling this challenge using modern machine learning techniques based on (i) synthetic data, typically generated using quantum mechanics based methods, and (ii) model architectures inspired by quantum mechanics. Such Quantum mechanics based Machine Learning (QML) approaches combine the numerical efficiency of statistical surrogate models with an {\em ab initio} view on matter. They rigorously reflect the underlying physics in order to reach universality and transferability across CCS. While state-of-the-art approximations to quantum problems impose severe computational bottlenecks, recent QML based developments indicate the possibility of substantial acceleration without sacrificing the predictive power of quantum mechanics

The Material Theory of Induction

The fundamental burden of a theory of inductive inference is to determine which are the good inductive inferences or relations of inductive support and why it is that they are so. The traditional approach is modeled on that taken in accounts of deductive inference. It seeks universally applicable schemas or rules or a single formal device, such as the probability calculus. After millennia of halting efforts, none of these approaches has been unequivocally successful and debates between approaches persist. The Material Theory of Induction identifies the source of these enduring problems in the assumption taken at the outset: that inductive inference can be accommodated by a single formal account with universal applicability. Instead, it argues that that there is no single, universally applicable formal account. Rather, each domain has an inductive logic native to it.The content of that logic and where it can be applied are determined by the facts prevailing in that domain. Paying close attention to how inductive inference is conducted in science and copiously illustrated with real-world examples, The Material Theory of Induction will initiate a new tradition in the analysis of inductive inference

Molecular Level Characterization of Dissolved Organic Matter Integrating Trapped Ion Mobility Spectrometry and Fourier Transform Ion Cyclotron Resonance Mass Spectrometry

Dissolved organic matter (DOM) is an extremely complex mixture of organic molecules ubiquitous in aquatic systems and a critical component of the global carbon cycle. Little is known about DOM structural composition at the molecular level. The work presented in this dissertation summarizes the development of a novel analytical toolbox based on trapped ion mobility spectrometry and Fourier transform ion cyclotron resonance mass spectrometry (TIMS-FT-ICR MS) that has significantly contributed to expand our knowledge of DOM molecular complexity and diversity. The TIMS-FT-ICR MS/MS analysis provided for the first-time lower and upper estimation of the molecular isomeric diversity. The TIMS-FT-ICR MS/MS methodology was further developed to allow for chemical formula-based isomeric and neutral loss fragmentation structural description and database validation. This novel procedure enabled the unambiguous assignment of candidate isomeric structures based on accurate mass, database MS/MS matching scores, and ion mobility. A fast and routine structural characterization DOM workflow method was developed: GraphDOM. The method utilizes neutral loss fragmentation patterns acquired using continuous accumulation of selected ions (CASI)-collision induced dissociation (CID) FT-ICR MS/MS. The neutral mass loss patterns are used to define structural families leading to the identification and visualization of the DOM transformational processes. The GraphDOM methodology was successfully applied to the characterization of DOM along a salinity transect of the Harney River, Florida Everglades. The GraphDOM method was further implemented with isomeric content description at the molecular level and applied to four common aquatic systems. The application of the GraphDOM methodology allowed for the first time identification of common and unique DOM transformational networks across aquatic ecosystems

Abstracts of Papers, 87th Annual Meeting of the Virginia Academy of Science, 2009

Full proceedings for the 87th Annual Meeting of the Virginia Academy of Science, May 27-29, 2009, Virginia Commonwealth University, Richmond V