Microbial Evolution through the Lens of Metagenomics and Archaeogenetics

Abstract

Microbial evolution is fundamental to fields ranging from industrial bioprocesses to agriculture and medicine, governing everything from pathogen emergence and transmission to environmental ecosystems and biotechnological advancement. However, microbiology’s historical reliance on culture-based methods and primarily clinically focused studies has limited our ability to fully integrate ecological and evolutionary perspectives, leaving critical gaps in knowledge regarding the genomic structure, diversity, and evolutionary dynamics of microbial populations. Host-microbial associations and microbial community interactions in particular drive several key biological events across Earth’s history - from organelle origins to evolutionary arms race dynamics. Recent advances in sequencing technologies have revealed a startling expansive picture of the genomic diversity of symbiotic microbes – including the enigmatic Candidate Phyla Radiation (CPR) – a group of abundant yet largely uncultured, ultra-small bacteria that are found ubiquitously from deep-sea hydrothermal vents to the human microbiome. In this dissertation, I integrate metagenomic, pangenomic, and paleogenomic approaches to investigate microbial eco-evolutionary dynamics. Altogether, this work illuminates how genomic plasticity facilitates microbial adaptation and diversification, with broad implications for the ecological understanding of microbial relationships and insights into host-microbial co-evolution. In my first chapter, I utilize a novel statistical framework that combines longitudinal metagenomic sampling with clonal sequencing to track the strain-level population dynamics of industrial yeast (Saccharomyces cerevisiae) lineages across two Brazilian bioethanol refineries over two industrial seasons. The results show diverging evolutionary trajectories: one plant’s yeast community is characterized by the stable dominance of a domesticated starter lineage, whereas another plant experiences invasion by foreign but closely related strains. These findings highlight how ecological forces, such as competition and invasion, influence microbial communities within industrial processes in relation to industrial operational stability. In my second chapter, I develop a scalable, integrative computational pipeline that combines metagenomic assembly and pangenomics to characterize the genome structure of over two thousand Parcubacteria (OD1) genomes in their environmental context, including novel genomes recently discovered from deep-sea anemones. Utilizing this approach, I demonstrate that Parcubacteria – despite having extremely reduced core genomes with limited metabolic capabilities – retain a flexible and modular accessory genome across clades, structured by phylogeny rather than habitat specificity. With a core genome that encodes primarily informational systems, DNA recombining and repair mechanisms, environmental sensing, and a conserved type IV pili, the Parcubacteria core genome reflects a host-dependent interaction-focused lifestyle. In contrast, the accessory genome exhibits remarkable modularity, with lineage-specific gene clusters encoding diverse specialized functions in secondary metabolism, stress responses, and signal transduction systems, with complete turnover between clades. Strikingly, co-occurrence analyses also show extensive genomic plasticity driven by insertion sequence (IS) elements – especially the IS21 family, and reveal structured mobility of antibiotic resistance genes (ARGs) and accessory gene clusters, highlighting ongoing gene transfer and mobility without disrupting core genomic integrity. Altogether, the results showcase a unique evolutionary strategy for small genomes where genome reduction is coupled with modular genomic innovation. Altogether, these findings redefine genome reduction paradigms by illustrating how Parcubacteria leverage dynamic accessory content and interaction alongside its genomic minimalism for flexible specialization and ecological persistence. My third chapter employs a combination of archaeogenetics and Bayesian tip-calibrated phylogenetics to reconstruct the evolutionary history and habitat transitions of the prominent CPR lineage Saccharimonadia (TM7), a globally distributed bacterial group that is also commonly found in human microbiomes. Leveraging ancient DNA derived from archaeological samples from ancient human populations and Neanderthals spanning a range of 100,000 years, as well as oral microbiome samples from underrepresented traditional farming and hunter-gatherer communities, I curated a dataset of 4,317 genomes, establishing one of the most comprehensive temporal datasets available for a CPR bacterial group. I identified at least seven independent habitat transitions from environmental reservoirs into mammalian hosts, with distinct lineage diversifications driven by host colonization and subsequent specialization in oral or gut biofilms. Bayesian evolutionary analyses were used to calculate substitution rates, with TMRCA (Time to the Most Recent Common Ancestor) dating diversification events into the Pleistocene epoch. Notably, my analyses also uncovered several previously unrecognized human-associated lineages that persist within ancient and non-industrialized human populations, indicating underrepresentation linked to human lifestyle or subsistence differences. Altogether, these findings underscore the significance of ancient DNA approaches in providing high-resolution evolutionary modeling for tracking long-term microbial evolutionary trajectories, and revealed how evolutionary events shape host-specific diversity patterns observed in modern Saccharimonadia populations.