This thesis examines the potential interface that exists between health information, specifically electronic health record (EHR) systems, and notifiable disease surveillance in Canada. It aims to highlight the benefits and barriers experienced by the current national notifiable disease surveillance strategy, as well as to highlight the successes and roadblocks to the successful implementation and adoption of EHR technologies in Canada. Qualitative methodologies, which include the 16 semi-structured interviews conducted with four key stakeholder groups, including public health experts, physicians, health administrators and academics that are concerned with EHR adoption and public health were used to obtain data. Data from interviews was analysed using grounded theory methodology and then verified using member checking and other data validation methods. Emergent themes from obtained data indicated that there is a large potential for the improvement of the current notifiable disease through the use of EHR technologies: however, the barriers currently faced by both the notifiable disease surveillance system and the state of implementation and adoption of EHR technologies prevent this from occurring. These barriers include political, financial, human, security/privacy, and technology barriers. Differences between stakeholder groups were explored, and potential solutions and insights into existing barriers were provided. The information gained from this study provides insight into the efficiency of the current infectious disease surveillance system and the progress of and need for the implementation of EHRs nationwide. In addition, the results of this study provide stakeholders with a deeper understanding of the barriers facing the use of EHR technologies for infectious disease surveillance and provide a starting place to address these issues. The results of this study can help to inform policy regarding public health surveillance and EHR implementation and adoption.

The onset of immune response against microbial stimuli activates induction of many anti- inflammatory genes and ISGs for effective clearance of the pathogen. This response includes transcriptional activation of several non-coding transcripts such as miRNAs and long non-coding RNAs (lncRNAs). LncRNAs constitutes the largest class of non-coding genome and are arbitrarily described as transcripts greater than 200 base pairs. Similar to protein coding mRNAs, lncRNAs are RNA polymerase II transcripts and undergo mRNA processing such as capping, splicing and polyadenylation. In recent years, high throughput sequencing has enabled an in-depth exploration of the human genome and subsequent discovery of lncRNAs. Several studies have highlighted the crucial role of lncRNAs in many biological processes including as regulators of gene expression as well as molecular effectors of host-pathogen driven immune responses. To date, majority of lncRNAs have been studied in murine models with limited understanding in human cells. In order to elucidate the role of lncRNAs in human immune cell regulation, the goal of this thesis is to identify and characterize novel lncRNAs critical to host-pathogen innate immune responses. RNA sequencing in LPS, IAV and HSV stimulated cells revealed lncRNA LUCAT1 as most differentially regulated lncRNA. CRISPR-cas9 and shRNA mediated depletion of LUCAT1 showed enhanced IFN-I genes signature, which was suppressed upon overexpression of LUCAT1. Additionally, LPS stimulated hDCs showed enrichment of LUCAT1 in the nucleus and its association with the chromatin markers. Further, LUCAT1 depletion contributed to enhanced occupancy of transcriptional coactivators at the promoters of IFN-I genes. Global identification of RNA associated proteins revealed LUCAT1 association with STAT1 in the nucleus thus emphasizing its role in transcriptional regulation of Type I IFN genes in inflammatory responses. This thesis furthers the understanding about the molecular factors affecting immune regulation and describes the novel role of LUCAT1 as an attenuator of immune cell response to pathogens.

Chlamydia is the most commonly reported sexually transmitted infection in the US, with an estimated 4 million new cases in 2018 alone. In addition to humans, Chlamydia infects other animals including mice, and mice have become a popular model for the study of Chlamydia infection. Female sex hormones (FSH) estrogen (E2) and progesterone (P4) rise and fall in a cyclic fashion in both humans and mice, and it is well established that these hormones affect the establishment and progression of genital chlamydial infection. Prior studies that used a co-culture model of human endometrial epithelial cells (IK cells) grown on extracellular matrix-coated inserts over human stromal cells (SHT cells) showed that E2 treatment enhanced initial chlamydial infection and production of progeny Chlamydia compared to hormone free (HF), P4 or combination E2’E2/P4 treatment. This led to the hypothesis that the treatment of ovariectomized (OVX) mice with E2 would enhance chlamydial infection compared to mice treated with no hormone, P4, or a combination of E2 and P4. We ordered OVX mice from Jackson Laboratories and surgically implanted silastic capsules that contained E2, P4, E2/P4, or no hormone diluted in sesame oil. A gas chromatography method was developed to test E2 and P4 concentration in mouse serum, ensuring that hormone levels were physiologically relevant. 8 days after the implantation of the capsules, mice were vaginally-inoculated with C. muridarum¸ a chlamydial species that mimics human chlamydial infection in mice. Every 3 days post infection (pi), for 21 days, we vaginally swabbed mice to determine how much C. muridarum each mouse shed and created a graphical representation of chlamydial shedding. A subset of mice were sacrificed on day 10pi so that presence and identity of immune cells could be analyzed by flow cytometry. Surprisingly, E2 alone and E2/P4 treatment completely protected mice from chlamydial infection. HF-treated mice peaked in chlamydial shedding on day 3pi, and P4-treated mice peaked on day 9pi. Flow cytometry data showed that E2-treated mice had a significantly reduced T cell presence in the genital tract. Thus far, our data suggest that FSH affect chlamydial infection in mice differently than in humans. This observation could have important implications for a field that is heavily reliant on murine studies.

Numerous factors that affect herpes simplex virus 1 (HSV1)-mediated pathogenesis have been identified. Such factors directly impact the replication of HSV1 as well as modulate host immune responses following HSV1 infection. In this work, I characterize how HSV1 replication is impacted by expression of the protein encoded by C21orf91, or “Cold Sore Susceptibility Gene” (CSSG1), that has been linked to HSV1 reactivation in humans. I investigated expression of CSSG1 mRNA expression in various tissues and found that CSSG1 mRNA was present in several tissues of importance in HSV1 disease, including brain, trigeminal ganglia (TG), cornea and spleen. Western blot analysis demonstrated that CSSG1 protein is expressed in human cells. Subcellular fractionation analysis reveals that CSSG1 is predominantly found in the cell nucleus, where it colocalizes with chromatin and with Tip60, a chromatin-binding histone modifying protein that has been shown to be essential for the replication of herpesviruses. I also discovered that CSSG1 is present in the cytosol of cells where it forms large cytosolic aggregates in presence of TRAF6, a downstream adapter that plays an important role in innate immune receptor signaling. To determine if CSSG1 directly impacts viral replication, I generated CSSG1 knockdown human cell lines. I found that HSV1 replication was reduced in CSSG1 knockdown cells compared to control cells, whereas replication of the unrelated virus, vesicular stomatitis virus (VSV), was not affected by knockdown of CSSG1. I demonstrate that CSSG1 was necessary for efficient expression of HSV1 viral proteins during infection. Western blot analysis and measurement of expression of HSV1 proteins expressed at various stages of viral replication illustrates that CSSG1 was required for HSV1 replication at very early stage of infection. I also noted that CSSG1 expression impacted the DNA damage response in HSV1 infected cells. Levels of H2AX phosphorylation, a marker of the DNA damage response, were increased in HSV1-infected CSSG1 knockdown cells compared to control cells. DNA damage responses are thought to promote HSV1 reactivation from latency and HSV1 gene expression, indicating a potential mechanism for role of CSSG1 in HSV1 replication through modulating the DNA damage response. Overall, my work demonstrates that CSSG1 affects HSV1 replication and provides insight on how CSSG1 polymorphisms in humans could affect HSV1 reactivation and replication to promote cold sores. These discoveries may also lead to a better understanding of pathogenesis of other herpesviruses in humans.

Many communicable diseases can be prevented with the use of immunizations. The issue this study addressed was the use of vaccinations and the rise of continued preventable communicable disease. Researchers have recommended that children be vaccinated, a recommendation that is bolstered by the readily available supply of vaccinations worldwide. However, even with vaccine promotions, availability, and education, children continue to be unvaccinated and outbreaks have continued to occur. For this study, data will be collected from the Iowa Registry of Immunization Records and a local community health center pediatric clinic. Examples of data that will be collected are vaccine records, race, and language spoken. Also, data from questionnaires given to parents, caregivers, and medical staff will be obtained to discover their beliefs, misconceptions, and thoughts on vaccines. The sample size will be 35, with recruitment occurring when the parent brings a child into the clinic for a well child or sick visit. The questionnaires will be collected and analyzed by totaling the responses from a pre-existing Likert scale questionnaire.The purposes of the project are to (a) discuss with health care providers a project, that consists of reasons children are not vaccinated; (b) develop a plan to determine those reasons and how to educate parents, the community being served by this health care facility and health care staff on the importance of immunizations; (c) and promote vaccinations for the this community.

Influenza is a seasonal acute respiratory infection, causing millions of illnesses worldwide on a yearly basis. A common subtype, the influenza A virus (IAV), is a single stranded RNA virus, that similarly to other subtypes, targets epithelial cells. The best way to protect against the virus is through vaccination. Vaccine induced protection is mediated through the generation of adaptive CD4 and CD8 T cells, as well as antibody producing B Cells. Although generally thought of as helper cells, previous research has highlighted additional roles of memory CD4 T cells in mediating protection against IAV beyond their helper function. More specifically they have been shown to enhance innate inflammatory responses and facilitate the recruitment of innate cell populations; including a recently discovered population of Innate Lymphoid Cells (ILC). Previous research has shown ILCs to have a key role in tissue repair and limiting tissue damage following infection. Whether memory cell response, during protective recall, modulates ILC repair function is currently not well understood. To better understand the possible regulation of ILCs by memory cells, we utilized a molecular technique called reverse transcription polymerase chain reaction (RT-qPCR), to assess select innate lymphoid cell associated protein expression following IAV challenge and rechallenge. We hypothesize, that memory cells drive differential expression of ILC associated repair proteins to assist in a faster and more efficient mobilization of repair processes following pathogenic invasion. Our goal is to highlight and better understand the regulatory and inflammatory responses memory cells provide following viral infection, as these may lead to key steps in the development of long lasting and efficacious vaccines.

Helminths are parasitic worms that can establish long-lived infections by modulating host immune responses. Helminth infection has been associated with a reduced prevalence of allergic disease in human populations, and impaired immunity to co-infecting pathogens. Several human and mouse studies suggest that helminths may impair host responses to concurrent bacterial infection. In this thesis, we study the extent to which helminth infection affects Salmonella colonization and how anthelmintic treatment (deworming) impacts immunity to Salmonella in a mouse model of co-infection. We find that helminth co-infection allows Salmonella to establish in the lumen of the small intestine. Further, we find that deworming prior to bacterial infection restores impaired immunity to Salmonella in the small intestine, however, deworming after Salmonella has established during helminth co-infection does not revert elevated bacterial burdens. To ensure their longevity in the host, helminths release immunomodulatory molecules, and modulate immunity through changes in the gut microbiota. The microbiota is known to influence mucosal immunity through the production of metabolites, but metabolites have not received much attention in the context of helminth modification of immune responses. This thesis uncovers the impact of helminth infection on levels of short-chain fatty acids (SCFAs) and bile acids, two groups of metabolites with immunomodulatory potential. We found that helminth infection increases small intestinal levels of the branched-chain SCFA isovalerate and lowers the small intestinal bile acid concentration. We next explored the consequences of these metabolite shifts on susceptibility to bacterial infection, helminth fitness and regulatory T cells. Collectively, these results contribute to the understanding of host-pathogen interactions in a co-infection scenario, which ultimately, will help to inform strategies for disease control in helminth-endemic areas. Further, our data contributes to the characterization of the helminth-modified intestinal metabolome, which future work can build on to reveal novel immunomodulatory pathways that can be targeted to relieve symptoms in inflammatory diseases such as allergic asthma. / Graduate / 2022-12-06

Cell-mediated lysis of cultured tumor target cells by nonspecific cytotoxic cells (NCC) was examined in the oyster toadfish (Opsanus tau), an estuarine teleost. NCC activity was evaluated in cells taken from the head kidney, peripheral blood, spleen and peritoneal cavity. NCC activity was a property of plastic nonadherent cells which lacked phagocytic activity, indicating that in terms of their functional capacity, the cells which mediate nonspecific cytoxic immune responses in oyster toadfish do not appear to be monocytes or macrophages. However, light and electron microscopic examination of these cells revealed that morphologically, they resemble monocytes and macrophages. A new technique to assess in vitro phagocytic function of fish macrophages is described. This assay involves the spectrophotometric measurement of congo red-stained yeast cells that have been phagocytized by macrophages. The assay is simple, rapid and reproducible. Furthermore, it is less subjective than previously described methods that employ microscopic examination of cells. Using this technique with oyster toadfish, phagocytosis of yeast cells was found to increase with time, reaching a maximum between 60 and 90 minutes (as determined by absorbance at 510 nm). Sampling sites selected for this field investigation had a sediment-bound polycyclic aromatic hydrocarbon (PAH) concentration gradient of 55 ppb to 96,000 ppb total PAH. Results of this study suggest that varying levels of PAH contamination did not lead to significant between-site differences in terms of simple ability of oyster toadfish macrophages to phagocytize foreign particles. This technique has been used in studies designed to characterize the functioning of oyster toadfish NCC and in toxicological investigations comparing the effects of PAH and NCC and macrophage function in oyster toadfish. Studies were undertaken to determine the effect of the chemical carcinogen 7,12-dimethylbenz (a) anthracene (DMBA) on the function of oyster toadfish peritoneal nonspecific cytotoxic cells (NCC) and macrophages. The functioning of these two cell populations was assessed in terms of their ability to lyse cultured tumor target cells and to phagocytize yeast cells, respectively. In a time-course study, the effects of DMBA on NCC activity were found to be highly persistent. Following a single intraperitoneal injection of 10 mg/kg, NCC activity was again virtually eliminated, and did not recover throughout the 28 days of the exposure study. (Abstract shortened by UMI.).

<p>The main body of this thesis deals with three related concepts pertaining to vaccination strategies for childhood infectious disease. Chapter 2 deals with the implications of reversion in the Oral Polio Vaccine on global polio eradication programs. Chapter 3 explores the phenomenon of contact or secondary vaccination in the use of live-attenuated virus vaccines. Chapter 4 explores the importance of demographic stochasticity in pulse vaccination campaigns. largely focusing on measles dynamics. Abstracts for each chapter are given below.</p><p>Poliomyelitis vaccination via live Oral Polio Vaccine (OPV) suffers from the inherent problem of reversion: the vaccine may, upon replication in the human gut, mutate back to virulence and transmissibility resulting in circulating vaccine derived polio viruses (cVDPVs). We formulate a general mathematical model to assess the impact of cVDPVs on prospects for polio eradication. We find that for OPV coverage levels below a certain threshold, cVDPVs have a small impact in comparison to the expected endemic level of the disease in the absence of reversion. Above this threshold, the model predicts a small but significant endemic level of the disease, even where standard models predict eradication. In light of this, we consider and analyze three alternative eradication strategies involving a transition from continuous OPV vaccination to either continuous Inactivated Polio Vaccine (IPV), pulsed OPV vaccination, or a one-time IPV pulse vaccination. Stochastic modeling shows continuous IPV vaccination is effective at achieving eradication for moderate coverage levels, while pulsed OPV is effective if higher coverage levels are maintained. The one-time pulse IPV method may also be a viable strategy, especially in terms of the number of vaccinations required and time to eradication, provided that a sufficiently large pulse is practically feasible. More investigation is needed rq?;arding the frequency of revertant virus infection resulting directly from vaccination, the ability of IPV to induce gut immunity, and the potential role of spatial transmission dynamics in eradication efforts.</p> <p>Viruses contained in live-attenuated vims vaccines (LAVV) can be transmitted between individuals, resulting in secondary or contact vaccinations. This fact has been exploited successfully in the use of the Oral Polio Vaccine (OPV) to better control wild polio viruses. In this work we analyze general LAVV vaccination models for infections that confer lifelong immunity. We consider both standard (continuous) vaccination strategies and pulse vaccination programs (where mass vaccination is carried out at regular intervals). For continuous vaccination, we provide a complete global analysis of a very general compartmental ordinary differential equation LAVV model. We find that the threshold vaccination level required for eradication of wild virus depends on the basic reproduction numbers of both the wild and vaccine viruses, but is otherwise independent of the distributions of the durations in each of the sequence of stages of disease progression (e.g., latent, infectious, etc.). Furthermore, even for vaccine viruses with reproduction numbers below one. which would naturally fade from the population upon cessation of vaccination, there can be a significant reduction in the threshold vaccination level. The dependence of the threshold vaccination level on the virus reproduction numbers largely generalizes to the pulse vaccination model. For shorter pulsing periods there is negligible difference in threshold vaccination level as compared to continuous vaccination campaigns. Thus, we conclude that current policy in many countries to employ annual pulsed OPV vaccination does not significantly diminish the benefits of contact vaccination.</p><p> In the last two decades, many countries have implemented pulse vaccination for infectious diseases (mass vaccination campaigns repeated annually or at other regular intervals). Based on deterministic mathematical models, previous work has shown that the total expected cost of control or eradication (measured by the number of vaccine doses required) is identical for pulse vaccination (with any pulse interval) and for traditional, continuous vaccination. We reconsider this problem using stochastic epidemic models (both by direct simulation and by employing a moment closure approximation). We focus on measles and show that demographic stochasticity has a large impact on the relative success of pulse and continuous vaccination programs, even for well-mixed populations as large as 10 million.</p> / Thesis / Doctor of Philosophy (PhD)

Background: ​Infectious disease outbreaks are amongst the most threatening of disasters, capable of affecting the health and economies of millions of people around the world in a single occurrence. Mathematical models are a tool that can be used to synthesize information from different disciplines into a comprehensive model, which can further be used to guide public health in making appropriate economic and social decisions. However, the integration of modelling within public health is not maximized. This study aims to explore perceptions of key stakeholders in planning, producing and applying infectious disease mathematical models in public health. Methodology: ​Data was collected using semi-structured key informant interviews with key stakeholder groups (n=19), academic modellers (n=6), government modellers (n=5), government end-users (n=5) and professionals and practitioners who are end-users (n=4). Data was analyzed with thematic analysis with NVivo 11 (QSR International). A stakeholder analysis was used to map out the interrelatedness of key stakeholder issues, and a thematic analysis was used to abstract themes of collaboration between stakeholders, challenges with data and perceptions of predictive modelling. Results and Conclusion:​ The findings of this study identify and organize important insights and recommendations required to optimize the utilization of infectious disease mathematical models in public health decision-making. The findings suggest that models that are most applicable to public health problems often go through iterative collaborations between end-users and modellers. The findings also suggest that there are growing challenges when it comes to the collection and interpretation of sources of infectious disease data and that mathematical models are valuable when used for understanding infectious disease outbreaks and/or interventions, rather than projecting the course of a specific outbreak. This study recommends actions be taken in education, practice and research to minimize the existing gap between mathematical models of infectious disease and their application for public health decision-making. / Thesis / Master of Science (MSc)