Biochemical analysis of human body fluids to produce reliable and timely data to enhance the accuracy of physician diagnoses and improve patient health outcomes.

Characterization of viral infection dynamics in marine cyanobacterial hosts and investigation of subsequent effects on bacterial chemotactic responses utilizing microfluidics, video microscopy, and automated image analysis in MATLAB.Viruses are ubiquitous in the marine environment and are causative agents of significant turnover of microbial populations yet their contribution to nutrient flux and carbon cycling has largely been overshadowed by physical and chemical forces in the field of oceanography. As a graduate student, I sought to elucidate the potential of marine viruses to influence interactions between microbial primary and secondary producers which ultimately may have consequences on carbon cycling in the oceans. Specifically, I worked with the virus-host pair of S-SSM5 and Synechococccus sp. WH8102. Synechococcus are a cyanobacterial genera with broad geographic range and large estimated contribution to global primary production. I used microfluidic assays and video microscopy to assess the chemotactic responses of the heterotrophic bacteria V. alginolyticus and P. haloplanktis to metabolic exudates collected from cultures of Synechococcus during different stages of infection. Additionally, chemotaxis of V. alginolyticus was characterized in the presence of actively infected Synechococcus cells. Timelapse images of bacterial chemotaxis were analyzed using custom code in MATLAB and revealed positive chemotactic responses of V. alginolyticus and P. haloplanktis to Synechococcus metabolic exudates and virus-infected cells. These results suggest a potential of marine viruses to influence microbial interactions which may ultimately have downstream implications on carbon cycling in the oceans.

Molecular engineering and characterization of live-attenuated multivalent vaccine strains targeting the enteric pathogens Enterotoxigenic E. coli (ETEC) and S. flexneri which overwhelmingly afflict developing regions. S. flexneri and ETEC are diarrheal pathogens that are significant sources of morbidity and mortality in children aged 0-5 years old in developing regions. Roadblocks to vaccine delivery in these regions include a lack of infrastructure (electricity, refrigeration, etc.) and difficulty in recruitment of individuals to undergo vaccination. Therefore, implementation of systems requiring the fewest immunizations may be preferable until these logistical challenges can be adequately addressed. One possible solution is to engineer multivalent vaccines that can target two or more pathogens with a single vaccination. To this end, we generated live-attenuated S. flexneri strains using the lambda red recombinase system to create guanine auxotrophic mutants. Immunogenic ETEC antigens were then cloned by PCR, ligated into the vector TN7, and transformed into DH5 alpha E. coli for initial screening and characterization. Promising constructs were electroporated into the attenuated S. flexneri and, via temperature sensitive homologous recombination, the sequences specifically encoding ETEC antigens were inserted into the chromosome of S. flexneri. Expression of ETEC antigens in S. flexneri was confirmed using western blot, slide agglutination, and transmission electron microscopy. Cell adherence and infection assays, animal vaccination and challenge studies, and ELISA were employed to further characterize the safety and immunogenicity of recombinant S. flexneri strains. We generated numerous promising recombinant strains with the potential to proceed to clinical trials.