Master of Science, The Ohio State University, 2023, Anatomy

One of the most common eukaryotic infections across the globe is cutaneous leishmaniasis, caused by various species of the parasite leishmania. These infections are characterized by ulcerations of the skin where the parasite has entered through the bite of a sandfly. Even though the presence of leishmania in the skin causes immune cell infiltration and inflammation, patients often report having little or no pain associated with the lesion. This could mean the leishmania parasites are interacting with peripheral nerves and nerve endings in some way. How leishmania influences the cellular mechanisms of a pain response has not been studied in much detail, especially in new world species like Leishmania Mexicana. Some of the most significant channels in promoting a pain response are found in the Transient Receptor Potential Vanilloid (TRPV) family of calcium channels, whose activation sensitizes sensory neurons to alert the body of noxious stimuli among other factors. These channels could be manipulated by a cutaneous leishmaniasis infection, and over time sensory neurons may change their expression of these receptors. The inflammation caused by L. Mexicana infection occurs in the dermis and hypodermis, where there are not only nerve endings but cutaneous nerves before they innervate the skin. Other species of leishmania have been found to interact with both fibroblasts and Schwann cells, but the impacts of L. Mexicana on either cell type have not been studied. After leaving the periphery, sensory neurons synapse in the dorsal horn of the spinal cord where there are various glial cells for support. These glial cells are highly sensitive to the input from sensory neurons, and often replicate in response to various pain conditions. The influence of L. Mexicana infection on glial cells is also not known and could reveal how the parasite is able to cause painless lesions. 16 mice were infected with L. Mexicana and sacked at various intervals throughout the infection to (open full item for complete abstract)

Committee: Melissa Quinn (Advisor); Chistopher Pierson (Committee Chair); James Cray (Committee Chair); Abhay Satoskar (Advisor) Subjects: Anatomy and Physiology

Doctor of Philosophy, The Ohio State University, 2022, Microbiology

Leishmaniasis is a neglected protozoan disease affecting over 12 million people globally. Cutaneous leishmaniasis (CL) is the most common form, characterized by chronic skin lesions. Currently, there are no approved vaccines for human use. We have generated centrin knock out Leishmania (L.) mexicana (LmexCen-/-) mutants using CRISPR/Cas9. Centrin is a cytoskeletal protein required only for intracellular amastigote replication in Leishmania. Here, we investigated the safety, immunogenicity, and efficacy of LmexCen-/- parasites in vitro and in vivo. Our data shows that LmexCen-/- amastigotes present a growth defect, which results in significantly lower parasitic burdens and increased protective cytokine production in infected macrophages and dendritic cells, compared to LmexWT. Furthermore, LmexCen-/- parasites are safe in susceptible mouse models and efficacious against challenge with LmexWT in genetically different BALB/c and C57BL/6 mice. Vaccinated mice did not develop cutaneous lesions, displayed protective immunity, and showed significantly lower parasitic burdens compared to the controls. Overall, we demonstrate that LmexCen-/- parasites are a promising candidate vaccine against CL in pre-clinical models. Next, we explored the metabolic drivers of these vaccine-mediated immunological profiles. Metabolomics are emerging as a useful tool to uncover unknown networks that govern immune regulation and determine functional specialization. We analyzed the metabolic changes occurring after immunization with LmexCen-/- and compared them with LmexWT infection. Our results show enriched aspartate metabolism and pentose phosphate pathway (PPP) in ears immunized with LmexCen-/- parasites. These pathways are both known to promote M1 polarization in macrophages, and PPP in particular induces nitric oxide production in macrophages cultured with LmexCen-/-, suggesting a shift to a pro-inflammatory phenotype following immunization. Furthermore, immunized mice showed enriched t (open full item for complete abstract)

Committee: Abhay Satoskar (Advisor); Pravin Kaumaya (Committee Member); Steve Oghumu (Committee Member); Jesse Kwiek (Committee Member) Subjects: Immunology; Microbiology; Neurosciences; Parasitology