The global population is rapidly aging. It is estimated that over the next thirty years, the number of individuals >60 years of age will increase by over a billion, and the number of individuals over age 80 may increase by 300 million. As humans age, our immune system becomes progressively weaker through a process called immune senescence. This age-related decrease in immune function increases susceptibility to infection and chronic diseases. Sepsis is a leading cause of death worldwide. Over the past two decades, there has been an increased incidence of sepsis which is due, in part, to our aging population and immune senescence. The gut microbiome, which plays an essential role in health and disease, is altered in aging and sepsis. Specifically, the commensal microorganisms of the gut microbiota are replaced with potentially pathogenic bacteria. This contributes to immune dysfunction and worsened outcomes in critical illness. The innate immune system can be “trained” to respond more effectively to pathogens. We examined trained immunity as an approach to modulating immunosenescence and microbiome diversity in aging. We investigated the effect of trained immunity on: i) immune cells from healthy aging subjects and sepsis patients and ii) the diversity of the microbiome in aging and sepsis. Our results indicate that trained immunity is effective in combatting age-related immunosenescence. We found that β-glucan induced trained immunity enhances monocyte metabolism, increases functionality as well as alters the transcriptome and epigenome in aging individuals and sepsis patients. We also found that trained immunity induced the expansion of a unique population of myeloid cells in sepsis. These cells are defined as FSChi, CD11b+, GR-1hi and express high levels of immunosuppressive PD-L1. In addition, we found that trained immunity reversed age-related changes to the microbiome and prevented alterations to the microbiome in septic mice. We found that the Firmicutes/Bacteroidetes ratio increased in aging; however, trained immunity reversed this increase and increased Clostridia in aged mice. In sepsis, trained immunity prevented expansion of Proteobacteria observed in control mice. Thus, our results indicate that trained immunity may be effective in modulating immune senescence and the microbiome in aging and sepsis.

Tuberculosis (TB) is primarily a pulmonary disease caused by Mycobacterium tuberculosis (Mtb). Mtb is highly infectious, but studies have shown that only 5–15% of Mtb-infected individuals develop TB disease. The Bacille Calmette-Gu.rin (BCG) vaccine is the only commercially available Mtb vaccine, but its efficacy varies based on the strain used. The Mtb PhoPR-mutant variant, MTBVAC, has been tested as a possible attenuated live vaccine against Mtb. Although it has successfully conferred durable CD4+ T-cell responses in infants, it has also resulted in adverse effects. Our goal is to identify PhoPR-regulated gene(s) that mediate Mtb-induced burst size necrosis in infected cells. PhoPR is a two-component system in mycobacteria. PhoR responds to environmental cues, such as changes in pH, and phosphorylates the PhoP transcription factor, which then activates or suppresses the expression of approximately 40 Mtb genes. The Mtb PhoPR-mutant strain is able to replicate in infected macrophages, but it does not induce the horizontal spread of Mtb to other immune cells. Our lab has previously shown that virulent, cytopathic strains of Mtb, such as H37Rv, suppress early apoptosis, have faster replication rates in macrophages, and trigger cell death at a lethal load threshold of approximately 25 bacteria. Cell death of infected macrophages primarily occurs via necrosis, which involves nuclear pyknosis without DNA fragmentation and general disruption of lipid bilayer membranes. Viable bacilli are released to infect other macrophages and neutrophils recruited to the developing TB lesion. Here, we show that PhoP contributes to burst size necrosis in macrophages and that the PhoP-regulated genes, fadD21 and pks3, are potential drivers of this necrosis. FadD21 and pks3 are involved in the generation of diacyl trehalose/penta-acyl trehalose (DAT/PAT) for cell wall synthesis, suggesting that Mtb cell wall composition may determine virulence. Therefore, we have uncovered potential targets for early intervention or vaccinations to avoid granuloma formation or tissue damage in response to Mtb-induced macrophage necrosis.

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Decentralised detection and monitoring of hearing loss can be supported by new mHealth technologies using automated testing, which can be facilitated by minimally trained persons. These technologies may prove particularly useful in an infectious disease (ID) clinic setting where patients are at high risk for hearing loss. The current study aimed to evaluate the clinical utility of mobile and automated audiometry hearing health technology in an ID clinic setting. The current study was exploratory as it aimed to determine whether smartphone automated audiometry and South African English Digits-In-Noise (SA Eng DIN) smartphone applications could be utilised in an infectious disease clinic setting to monitor an HIV-related hearing loss in a feasible and time efficient way. Smartphone automated audiometry (hearTest™) and speech-in-noise testing (SA English Digits-In-Noise (DIN) test) were compared with manual audiometry at 2, 4, and 8 kHz. Smartphone automated audiometry and the DIN test were repeated to determine the test re-test reliability. Two hundred subjects (73% female and 27% male) were enrolled. Fifty participants were re-tested with the smartphone applications. Participants’ ages ranged from 18 to 55 years with a mean age of 44.4 (8.7 SD). Threshold comparisons were made between smartphone audiometry testing and manual audiometry. Smartphone automated audiometry, manual audiometry, and test re-test measures were compared to determine the statistical significance of any differences observed using the Wilcoxon signed-ranked test. Spearman rank correlation test was used to determine the relationship between the smartphone applications and manual audiometry, as well as for test re-test measurements. For all participants, 88.2% of thresholds corresponded within 10 dB or less between smartphone audiometry and manual audiometry. There was a significant difference (p>0.05) between smartphone and manual audiometry for the right ear at 4 and 8 kHz and the left ear at 2 and 4 kHz respectively. No significant difference was noted (p>0.05) between test and re-test measures of smartphone technology except at 4kHz in the right ear in smartphone automated audiometry. The absolute average difference between the initial and re-test of DIN testing was 1.2 dB (1.5 SD). No significant difference was noted in the test re-test measures of the DIN test (p < vii 0.05). A correlation coefficient of 0.56 was present in the DIN test re-test measures when the Spearman rank correlation test was administered. Smartphone audiometry with calibrated headphones provides reliable results and can be used as a baseline and monitoring tool at ID clinics. / Dissertation (MA)--University of Pretoria, 2017. / Speech-Language Pathology and Audiology / MA / Unrestricted

Exposure to violence and dysfunction in childhood is a major public health concern. The Adverse Childhood Experiences (ACEs) study demonstrated that childhood maltreatment and family dysfunction impact adult health, contributing to risk behaviors, infectious and chronic disease, and premature death. South Africa (SA) has one of the highest rates of violence and family dysfunction globally, and those living in townships are suspected to be disproportionately affected. Munsieville, the oldest undeveloped township in SA, has reported high rates of violence in the community. This study aimed evaluate the association between a history of childhood sexual abuse and various forms of risky sexual behavior. Data were collected by a team of researchers from the College of Public Health as part of a pilot study of ACEs in Munsieville. Self-report of sexual abuse before age 18 was used to compute the independent variable, which ranged from 0-1, with 0 implying the absence of any type of childhood sexual abuse and 1 implying one or more forms of childhood sexual abuse. Age and gender were deemed potential confounders. Two binary l outcomes representing forms of risky sexual behavior were considered, self-report of transactional sex and substance use before sexual activity. Descriptive analysis examined the frequency of childhood sexual abuse by each category of the study outcomes. Two multiple logistic regression models were individually constructed to examine the association between childhood sexual abuse and transactional sex, and substance use before sexual activity. Odds ratios and corresponding 95% confidence intervals were reported. Findings of the descriptive analysis indicated that 8.83% (43) of the sample reported participating in transactional sex, 22.4% (101) reported using either drugs or alcohol before sex. Moreover, 21.6% (103) reported experiencing one or more forms of childhood sexual abuse. A positive statistically significant association between self-reported childhood sexual abuse and transactional sex was identified (OR: 3.45; 95% CI: 1.71 – 6.98), illustrating that those who experienced any type of child sexual abuse had a 3.5 times as likely to report transactional sex compared to those who did not experience sexual abuse during childhood. Childhood sexual abuse was also significantly associated with substance use before sexual activity (OR: 1.93; 95% CI: 1.11 – 3.34). The study findings suggests a need for further research to understand the long term effects of child sexual abuse. Further, future public health interventions aimed at reducing sexual abuse and violence inflicted on South African children should be employed as means to improve their wellbeing in adulthood.

Host metabolism is profoundly altered during bacterial infection, both as a consequence of immune activation and secondary to virulence strategies of invading pathogens. As a result, the metabolic pathways that regulate nutrient acquisition, energy storage, and resource allocation in host cells must adapt to pathogen stress in order to meet the physiological demands of the host during infection. In this work, we uncover that the synthesis of the monounsaturated fatty acid (MUFA) oleate is necessary for the pathogen-mediated induction of immune defense genes. Accordingly, C. elegans deficient in oleate production are hypersusceptible to infection with diverse human pathogens, which can be rescued by the addition of exogenous oleate. However, oleate is not sufficient to drive protective immune activation. Oleate is also important for proper lipid storage and abundance. We found that exposure to pathogenic bacteria drives rapid somatic depletion of lipid stores in C. elegans. Activating the p38/MAPK immune signaling pathway in the absence of pathogens was also sufficient to drive loss of somatic fat. In addition, we found that transcriptional suppression of MUFA synthesis occurs during P. aeruginosa infection, in a manner dependent on pathogen virulence. Finally, we showed that the host compensates for the pathogen-induced depletion of fatty acids by promoting the redistribution of oleate from non-intestinal tissues to support immune function in the intestine. Together, these data add to the known health-promoting effects of MUFAs, and suggest an ancient link between nutrient stores, metabolism, and host responses to bacterial infection.

Study Objective: To compare the rates and severity of hospital-acquired Clostridium difficile infection (CDI) among patients taking proton pump inhibitors (PPIs) versus those not taking PPIs. Design: Retrospective, single-center, cohort study. Setting: Tertiary community hospital with a teaching service. Patients: A total of 41,663 patients with CDI who were hospitalized between January 2013 and May 2014; of those, 17,471 patients (41.9%) had received at least one dose of a PPI (PPI group), and 24,192 patients (58.1%) had no PPI exposure (control group). Measurements and Main Results: A total of 348 patients had CDI during the study period, with 269 cases present on admission. Hospital-acquired CDI was defined as CDI diagnosis occurring on or after the third calendar day of admission. After excluding those patients with CDI on admission, 65 (0.38%) of 17,302 patients later developed CDI in the hospital in the PPI group compared with only 14 (0.058%) of 24,092 patients in the control group. Of these patients, 36 patients (0.21%) in the PPI group met the definition of severe CDI compared with 8 (0.03%) in the control group. This demonstrated an unadjusted relative risk (RR) of 6.46 (95% confidence interval [CI] 3.63–11.51, p<0.0001) of developing hospital-acquired CDI and an unadjusted RR of 6.27 (95% CI 2.91–13.48, p<0.0001) of developing severe CDI while taking a PPI. When evaluating only patients who developed severe-complicated CDI, there were 22 cases in the PPI group and 2 cases in the control group, demonstrating an unadjusted RR of 15.3 (95% CI 3.6–65.13, p=0.0002) of developing severe-complicated CDI. Confounding variables were similar between groups. Conclusion: PPI use was associated with an increase in both the rate and severity of hospital-acquired CDI.

To develop a scale to assess infectious disease-specific health literacy (IDSHL) in China and test its initial psychometric properties. Methods: Item pooling, reduction and assessment of psychometric properties were conducted. The scale was divided into 2 subscales; subscale 1 assessed an individual's skills to prevent/treat infectious diseases and subscale 2 assessed cognitive ability. In 2014, 9000 people aged 15-69 years were randomly sampled from 3 provinces and asked to complete the IDSHL questionnaire. Cronbach's á was calculated to assess reliability. Exploratory factor analysis, t-test, correlations, receiver operating characteristic (ROC) curve and logistic regression were used to examine validity. Results: Each of the 22 items in subscale 1 had a content validity index >0.8. In total, 8858 people completed the scale. The principal components factor analysis suggested a 5-factor solution. All factor loadings were >0.40 ( p<0.05). The IDSHL score was 22.07±7.91 (mean±SD; total score=38.62). Significant differences were observed across age (r=.0.276), sex (males: 21.65±8.03; females: 22.47±7.78), education (14.16±8.19 to 26.55±6.26), 2-week morbidity (present: 20.62±8.17, absent: 22.35±7.83; p<0.001) and health literacy of the highest and lowest 27% score groups (all p<0.05). The ROC curve indicated that 76.2% of respondents were adequate in IDSHL. Binary logistic regression analysis revealed 12 predictors of IDSHL adequacy (p<0.05). Among the 22 remaining items, Corrected Item-Total Correlation ranged from 0.316 to 0.504 and Cronbach's á values ranged from 0.754 to 0.810 if the items were deleted. The overall á value was 0.839 and the difficulty coefficient ranged from 1.19 to 4.08. For subscale 2, there were statistically significant differences between the mean scores of those with a correct/incorrect answer (all p<0.001). Conclusions: The newly developed 28-item scale provides an efficient, psychometrically sound and userfriendly measure of IDSHL in the Chinese population.

Background Vancomycin dosing strategies targeting trough concentrations of 15–20 mg/L are no longer supported due to lack of efficacy evidence and increased risk of nephrotoxicity. Area-under-the-curve (AUC24) nomograms have demonstrated adequate attainment of AUC24 goals ≥ 400 mg h/L with more conservative troughs (10–15 mg/L). Objective The purpose of this study is to clinically validate a vancomycin AUC24 dosing nomogram compared to conventional dosing methods with regards to therapeutic failure and rates of acute kidney injury. Setting This study was conducted at a tertiary, community, teaching hospital in the United States. Method This retrospective, cohort study compared the rates of therapeutic failures between AUC24-extrapolated dosing and conventional dosing methods. Main outcome measure Primary outcome was treatment failure, defined as all-cause mortality within 30 days, persistent positive methicillin-resistant Staphylococcus aureus blood culture, or clinical failure. Rates of acute kidney injury in non-dialysis patients was a secondary endpoint. Results There were 96 participants in the extrapolated-AUC24 cohort and 60 participants in the conventional cohort. Baseline characteristics were similar between cohorts. Failure rates were 11.5% (11/96) in the extrapolated-AUC24 group compared to 18.3% (11/60) in the conventional group (p = 0.245). Reasons for failure were 6 deaths and 5 clinical failures in the extrapolated-AUC24 cohort and 10 deaths and 1 clinical failure in the conventional group. Acute kidney injury rates were 2.7% (2/73) and 16.4% (9/55) in the extrapolated-AUC24 and conventional cohorts, respectively (p = 0.009). Conclusion Extrapolated-AUC24 dosing was associated with less nephrotoxicity without an increase in treatment failures for bloodstream infections compared to conventional dosing. Further investigation is warranted to determine the relationship between extrapolated-AUC24 dosing and clinical failures.

Chitin is an abundant polysaccharide found in fungal cell walls, crustacean shells, and insect exoskeletons. The immunological properties of both chitin and its deacetylated derivative chitosan are of relevance due to frequent natural exposure and their increasing use in translational applications. Depending on the preparation studied and the endpoint measured, these compounds have been reported to induce allergic responses, inflammatory responses, or no response at all. Highly purified chitosan and chitin were prepared and the capacity of these glycans to stimulate the release of the inflammasomeassociated cytokine IL-1β was examined. Chitosan was shown to be a potent inflammasome activator in mouse bone marrow macrophages, macrophages polarized towards a M1 or M2 phenotype, dendritic cells, peritoneal cells, and human PBMCs. Acetylation of the chitosan to chitin resulted in a near total loss of IL-1β activity in all cell types tested. The size of the chitosan particles played an important role, with small particles eliciting the greatest activity. An inverse relationship between size and stimulatory activity was demonstrated using chitosan passed through size exclusion filters as well as with chitosan-coated beads of defined size. Partial digestion of chitosan with pepsin resulted in a larger fraction of small phagocytosable particles and more potent inflammasome activity. Inhibition of phagocytosis with cytochalasin D abolished the IL- 1β stimulatory activity of chitosan, offering an explanation for why the largest particles were nearly devoid of activity. Thus, the deacetylated polysaccharide chitosan potently activates the NLRP3 inflammasome in a phagocytosis-dependent manner. The reason for chitin’s inability to elicit IL-1β is unknown, but it does not appear to be due to active inhibition of the inflammasome and while chitin appears to be more readily digested by macrophage cell lysates, it does not occur at a rate which would likely impact inflammasome activation. There are three proposed mechanisms for NLRP3 inflammasome activation: K+ efflux, ROS, and lysosomal destabilization. The contributions of these mechanisms were tested and it was revealed that each of these pathways participated in optimal NLRP3 inflammasome activation by chitosan. Finally, the laminin receptor was evaluated as a potential chitin receptor. These studies provide insight into the activating properties of chitin and chitosan and highlight the importance of matching particle size and degree of acetylation to the level of activity desired for translational applications.