Genetic manipulation of Anopheles stephensi immunity to increase plasmodium falciparum salivary gland sporozoite infection levels

Human malaria is responsible for over 700,000 deaths a year. To stay abreast of the threat posed by the parasite, a constant stream of new drugs and vector control methods are required. This study focuses on a vaccine that has the potential to protect against parasite infection, but has been hindered by developmental challenges. In malaria prevention, live, attenuated, aseptic, Plasmodium falciparum sporozoites (PfSPZ) can be administered as a highly protective vaccine. PfSPZ are produced using adult female Anopheles stephensi mosquitoes as bioreactors. Production volume and cost of a PfSPZ vaccine for malaria are expected to be directly correlated with Plasmodium falciparum infection intensity in the salivary glands. The sporogonic development of Plasmodium falciparum in A. stephensi to fully infected salivary gland stage sporozoites is dictated by the activities of several known components of the mosquito’s innate immune system. Here I report on the use of genetic technologies that have been rarely, if ever, used in Anopheles stephensi Sda500 to increase the yield of sporozoites per mosquito and enhance vaccine production. By combining the Gal4/UAS bipartite system with in vivo expression of shRNA gene silencing, activity of the IMD signaling pathway downstream effector LRIM1, an antagonist to Plasmodium development, was reduced in the midgut, fat body, and salivary glands of A. stephensi. In infection studies using P. berghei and P. falciparum these transgenic mosquitoes consistently produced significantly more salivary gland stage sporozoites than wildtype controls, with increases in P. falciparum ranging from 2.5 to 10 fold. Using Plasmodium infection assays and qRT-PCR, two novel findings were identified. First, it was shown that 14 days post Plasmodium infection, transcript abundance of the IMD immune effector genes LRIM1, TEP1 and APL1c are elevated, in the salivary glands of A. stephensi, suggesting the salivary glands may play a role in post midgut defense against the parasite. Second, a non-pathogenic IMD signaling pathway response was observed which could suggest an alternative pathway for IMD activation. The information gained from these studies has significantly increased our knowledge of Plasmodium defense in A. stephensi and moreover could significantly improve vaccine production.

Regenerative Aquaculture: Designing for Resilience of the Chesapeake Tidewater

The Chesapeake Bay is out of balance. As the effects of decades of overharvesting, overdevelopment, and pollution have taken their toll, tidewater communities are confronted with a loss of culture, livelihood, and the environment in which they live. This thesis seeks to reframe the problem of regeneration of community and environment, through the use of resilient design. Resilient design is the process of designing for an uncertain at risk future. Through resilient design, architecture and aquaculture can be combined with food culture to foster stewardship of place. This thesis will explore interconnectedness of tidewater food culture, the waterman culture, aquaculture, and regenerative design in an effort to generate a holistic solution. The final product will consist of a methodology of planning for resilience at a framework scale, and will also propose an architectural solution that combines educational facilities with commercial aquaculture, to foster stewardship and regeneration in the Chesapeake Tidewater.

The Influence of Place Attachment, Aspirations, and Rapidly Changing Environments on Resettlement Decisions

Resettlement associated with development projects results in a variety of negative impacts. This dissertation uses the resettlement context to frame the dynamic relationships formed between peoples and places experiencing development. Two case studies contribute: (a) the border zone of Mozambique’s Limpopo National Park where residents contend with changes to land access and use; and (b) Bairro Chipanga in Moatize, Mozambique where a resettled population struggles to form place attachment and transform the post-resettlement site into a “good” place. Through analysis of data collected at these sites between 2009 and 2015, this dissertation investigates how changing environments impact person-place relationships before and after resettlement occurs. Changing environments create conditions leading to disemplacement—feeling like one no longer belongs—that reduces the environment’s ability to foster place attachment. Research findings indicate that responses taken by individuals living in the changing environment depend heavily upon whether resettlement has already occurred. In a pre-resettlement context, residents adjust their daily lives to diminish the effects of a changing environment and re-create the conditions to which they initially formed an attachment. They accept impoverishing conditions, including a narrowing of the spaces in which they live their daily lives, because it is preferred to the anxiety that accompanies being forced to resettle. In a post-resettlement context, resettlement disrupts the formation of place attachment and resettled peoples become a placeless population. When the resettlement has not resulted in anticipated outcomes, the aspiration for social justice—seeking conditions residents had reason to expect—negatively influences residents’ perspectives about the place. The post-resettlement site becomes a bad place with a future unchanged from the present. At best, this results in a population in which more members are willing to move away from the post-resettlement site, and, at worse, complete disengagement of other members from trying to improve the community. Resettlement thus has the potential to launch a cycle of movement- displacement-movement that prevents an entire generation from establishing place attachment and realizing its benefits. At the very least, resettlement impedes the formation of place attachment to new places. Thus, this dissertation draws attention to the unseen and uncompensated losses of resettlement.