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The advent of next-generation sequencing (NGS) technology has enabled researchers unprecedented access to genomic data that can reveal the evolutionary histories as well as future trajectories of natural populations from microbes to human populations. Due to the large quantity of sequencing data as well as the complexity of evolutionary models, such inference often requires sophisticated computational tools. Hence, advances in computational methods are essential for efficient learning and drawing insights from the wealth of genomic data. In this work, I developed several methods aimed to improve various aspects of existing approaches for making inference from genomic variation data. The first method, donni, is a supervised machine learning (ML) approach based on dadi, a widely used likelihood-based demographic history inference framework. With a more efficient management of the data generated by dadi's underlying models, donni significantly improves the computational efficiency while maintaining similar inference accuracy. Donni also includes an accompanying uncertainty quantification method, which is often lacking in most existing ML-based approaches in the field. The second method, ConfuseNN, focuses on improving the interpretability of inference results generated by existing trained convolutional neural networks (CNNs), which is an increasingly popular approach that has been applied to many population genomic inference tasks. With ConfuseNN, we designed and implemented a suite of tests that disrupt a specific feature in the genomic image data (e.g. allele frequency, linkage disequilibrium, etc.) to assess how each feature affects the CNN performance. I applied these tests to CNNs developed by other groups as well as the CNN we developed in our group for DFE inference (DFEnn), identifying the fundamental population genomic features that drive inference for each network. Finally, our group has recently developed an extension of dadi to improve demographic history i

The taxonomy of viruses is developed and overseen by the International Committee on Taxonomy of Viruses (ICTV), which scrutinizes, approves and ratifies taxonomic proposals, and maintains a list of virus taxa with approved names (https://ictv.global). The ICTV has approximately 180 members who vote by simple majority. Taxon-specific Study Groups established by the ICTV have a combined membership of over 600 scientists from the wider virology community; they provide comprehensive expertise across the range of known viruses and are major contributors to the creation and evaluation of taxonomic proposals. Proposals can be submitted by anyone and will be considered by the ICTV irrespective of Study Group support. Thus, virus taxonomy is developed from within the virology community and realized by a democratic decision-making process. The ICTV upholds the distinction between a virus or replicating genetic element as a physical entity and the taxon category to which it is assigned. This is reflected by the nomenclature of the virus species taxon, which is now mandated by the ICTV to be in a binomial format (genus + species epithet) and is typographically distinct from the names of viruses. Classification of viruses below the rank of species (such as, genotypes or strains) is not within the remit of the ICTV. This article, authored by the ICTV Executive Committee, explains the principles of virus taxonomy and the organization, function, processes and resources of the ICTV, with the aim of encouraging greater understanding and interaction among the wider virology community.

The correction refers to the sentence: Another notable taxonomic change approved in this ratification was the inclusion of gene transfer agents (GTAs) in the classification scheme as viriforms [153]. The sentence should read: Another notable taxonomic change approved in this ratification was the inclusion of gene transfer agents (GTAs) in the classification scheme as viriforms [154]. The original article has been corrected.

Antimicrobial research and development are not of economic priority and has led to few companies bringing new antibiotics to market in recent years. Modern antibiotic usage dates back to the 1940s about a decade after the initial discovery of Penicillin, followed by the “golden era” of discovery of novel antibiotic classes between the 1950s and 1970s. Numerous classes of antibiotics have been discovered since, but only a few have come to market. With antibiotic resistance on the rise not only in human pathogens, but especially in agricultural settings, there is a need for a new class of antimicrobial that can be optimized to be narrow spectrum and combat antibiotic-resistant pathogens. Tailocins, a class of bacteriocins, are proteinaceous antimicrobials produced by a variety of bacteria that kill closely-related strains while also playing a role in shaping the microbiome. Previous studies have pointed to the potential for tailocins to be used prophylactically or therapeutically against bacterial infections, specifically against plant pathogens for healthier crop production. We used what was previously known in the field to better characterize tailocins-producing Pseudomonas syringae strains and uncover the genetic mechanisms behind production, specificity, and killing. Together, this research can be taken further towards the goal of developing new, more efficient antimicrobials.

Zoonotic viral spillover events pose a constant threat to human health. A multitude of factors such as climate change and increased population density have expanded the human-animal interface, leading to increased opportunities for viral spillover from animals to humans. Much of the work in this dissertation was done in response to the COVID-19 pandemic and to understand the human immune response to SARS-CoV-2 infection, however, the advances in knowledge and technology can be applied for future pandemic prevention and management. Initially, widespread testing of the population was difficult and placed a large burden on healthcare workers, who were already overextended due to demands of patient care. We validated a massively scalable alternative to the gold-standard, nasopharyngeal swab sample collection method, called saline gargle. This collection method was implemented as part of the University of Arizona’s Test, Trace, Treat program to facilitate a safe return to in-person instruction. Not only did this expand testing capacity, but it enabled the University to perform comprehensive whole-viral genome sequencing and monitor for the presence of more transmissible and pathogenic variants on campus, and in turn, provided the opportunity for policies to be implemented in response to the shifting pandemic landscape. During the initial mRNA vaccine rollout, we identified immunocompromised individuals as an important population that had been excluded from mRNA clinical trials. We recruited individuals undergoing immunosuppressive therapy for cancer treatment and interrogated their cellular and humoral immune responses to the recommended two-dose mRNA vaccine series. We determined that these individuals derived great benefit from a third mRNA dose, leading to a recommended three-dose primary series for immunocompromised individuals. Immunocompromised people have been shown to be susceptible to prolonged SARS-CoV-2 infections. I present a case-study where an individual ha

Clostridioides difficile is a leading cause of healthcare-associated infections worldwide, and there is considerable debate about the optimal diagnostic method for C. difficile infection (CDI). Multi-step diagnostic algorithms use enzyme immunoassays (EIA) for the detection of C. difficile in stool. According to the literature, patients with a negative specimen toxin EIA result (while positive for the presence of C. difficile toxin gene by nucleic acid amplification test) have less severe disease, shorter diarrhea duration, and fewer adverse events. This work is a series of systematic assessments on the prevalence and virulence of C. difficile isolated from these discrepant specimens. In our retrospective analysis of 968 symptomatic patient stool samples from three Southern Arizona hospitals, 38% were discrepant. Many of the C. difficile isolates recovered from discrepant specimens belong to outbreak-associated molecular types (ribotype RT027; 32%) associated with more severe disease and higher risk of adverse events. RT027 strains isolated from discrepant specimens produce markedly less toxin than RT027 strains isolated from non-discrepant specimens. These “low-toxin” (LT) RT027 (LT-027) strains are equivalently cytotoxic to cultured host cells and exhibit comparable disease trajectories in rodent models compared to the high-toxin (HT) RT027 strains. However, LT-027 strains display robust colonization in vivo. Immunoblotting with serum of animals infected with LT or HT strains reveals that LT-027 display distinct surface proteins. Comparative genomic and proteomic analyses of several LT-027 strains identified unique genes and altered protein expression, respectively, relative to HT RT027 strains. Utilizing these data, we developed a rapid diagnostic assay to detect C. difficile while also providing epidemiological data. Collectively, this work highlights the robust virulence and clinical relevance of LT C. difficile strains.

Innate immune responses are crucial for limiting virus infection. However, viruses often hijack our best defenses for viral objectives. Human Cytomegalovirus (HCMV) is a beta herpesvirus which establishes a life-long latent infection. Defining the virus-host interactions controlling latency and reactivation is vital to the control of viral disease risk posed by virus reactivation. We defined an interaction between UL138, a pro-latency HCMV gene, and the host deubiquintase complexes, UAF1-USP. UAF1 is a scaffold protein pivotal for the activity of ubiquitin specific peptidases (USP), including USP1, USP12, and USP46. UAF1-USP1 sustains an innate immune response through the phosphorylation and activation of signal transducer and activator of transcription-1 (pSTAT1), as well as regulates the DNA damage response. UAF1-USP12 prevents dephosphorylation of pSTAT1 to sustain signaling. After the onset of viral DNA synthesis, pSTAT1 levels are elevated in infection and this depends upon UL138, USP1, and USP12. Upstream kinases, JAK1 and TYK2 are activated regardless of sustained pSTAT1 suggesting UL138-UAF1-USP1 are maintaining pSTAT1 downstream of the kinases. The most likely mechanism of UL138-UAF1-USP1 in maintaining pSTAT1 is deubiquitination of two regulatory monoubiquitin on STAT1 to allow for STAT1 interaction with IFNAR and JAK1 for phosphorylation and activation. Meanwhile, UL138-UAF1-USP12 complexes sustain pSTAT1 by preventing acetylation of pSTAT1 and subsequent dephosphorylation by the phosphatase, PTPN2. Early in infection, pSTAT1 induces specific ISGs but at late times post infection, localizes to viral centers of replication, binds to the viral genome, and influences viral gene expression. Inhibition of USP1 results in a failure to establish latency, marked by increased viral genome replication and production of viral progeny. Inhibition of Jak-STAT signaling also results in increased viral genome synthesis, consistent with a role for UL138-USP1-mediated reg

Streptococcus pneumoniae is a Gram-positive opportunistic pathogen that typically lives asymptomatically in the human nasopharynx but can cause pneumoniae, meningitis, otitis media, and sepsis. As with any organism living within a host, S. pneumoniae must acquire all its nutrients from the host. One group of essential nutrients is metals. Metals are vital to many cellular processes where they serve as structural components or catalytic cofactors of proteins. While metals are essential for life, they can also become toxic if their concentration is not tightly regulated. Bombardment of bacteria with toxic metals is one of the strategies of the mammalian innate immune system. Two metals that are known for their antimicrobial activity are copper and zinc. The metal environment will change as the bacteria move to different host tissues or experience the host immune response. In order to overcome stresses imposed by excess metal, bacteria encode metal efflux proteins. A notable example is the cop operon, a dedicated copper export system under the control of CopY. CopY is a metal-sensitive repressor that binds more tightly to DNA in a zinc-bound state and releases from DNA upon copper binding. Due to the role of these two metals in CopY regulation and their use as antimicrobials, we studied how changes in their concentrations impact S. pneumoniae. This dissertation explores the transcriptional and metabolic profile under a range of zinc and copper concentrations. In addition, we focused on CopY as we had predicted its regulon would likely expand beyond the cop operon. However, this was not the case, and we now propose an updated consensus operator sequence for CopY. Lastly, we look at how dysregulation of the cop operon affects the cell and we present the early stages of work to predict the CopY regulon across bacteria for which a full annotated genome is available.