Date of Award

January 2025

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Biomedical Sciences

First Advisor

Catherine Brissette

Abstract

As climate change leads to global expansion of tick habitats, tick borne diseases are becoming more prevalent and widespread, and emerging tickborne pathogens Borrelia miyamotoi and Borrelia mayonii warrant increased study. Both B. miyamotoi and B. mayonii are vectored by the same tick species as B. burgdorferi, the causative agent of Lyme disease, with B. miyamotoi causing a relapsing fever-like disease known as Borrelia miyamotoi disease (BMD), and B. mayonii causing a Lyme disease-like illness with atypical clinical presentations. This dissertation has two aims. The first aim encompasses B. miyamotoi infection and growth, isolation of clonal populations, and characterization of differences between clonal isolates to determine potential novel virulence factors. The second aim investigates B. mayonii pathology, both typical and atypical – arthritis and carditis, and spirochetemia – in Lyme disease-susceptible mice. For Study 1, we used the North American tick isolate Borrelia miyamotoi LB-2001, a heterogenous strain that reliably infects immunocompromised mice. However, cultivating B. miyamotoi LB-2001 in vitro posed issues, with difficulty achieving reliable growth from freezer stocks. To mitigate this, cultures were continually grown in vitro until LB-2001 adopted a high-passage, culture-adapted phenotype. This high-passage B. miyamotoi grew to densities 10 times higher than original low-passage cultures and was readily revived from frozen stocks. However, high-passage cultures were no longer able to infect severe combined immunodeficiency disease mice (SCID). Culture adaptation has been known to alter physiological traits relevant to pathogenicity and host interaction. To elucidate the change that occurred in the high-passage cultures leading to loss of infectivity, clonal populations were necessary. Until this point, all laboratory strains of B. miyamotoi were heterogenous, as poor in vitro growth prevented the isolation of single cells to produce clonal populations. Our lab was able to isolate clonal populations from both high- and low-passage cultures, resulting in clonal populations that demonstrate varied infection phenotypes in SCID mice, known as LB-V, LB-A, and LB-S. Isolates are described based on differences in vivo and in vitro, including growth in culture and protein production; high levels of spirochetes in the blood (spirochetemia), hepatosplenomegaly, and anemia in SCID mice; and variants identified using DNA sequencing. The study concludes as the first to isolate clonal populations of B. miyamotoi LB-2001, describe loss of infectivity and resultant changes to in vitro growth, and offer insights to future B. miyamotoi investigators keen on identifying novel virulence determinants. For Study 2, we utilized Borrelia mayonii, a novel Lyme disease causative agent in the Upper Midwestern United States. Borrelia mayonii infection presents with some atypical clinical manifestations, of which unusually high spirochetemia is of particular interest. While high spirochetemia has been reported in 75% of infected patients, B. mayonii has been undetectable in the blood of immunocompetent laboratory mice, though no study has purposely investigated spirochetemia. Working toward elucidating this conundrum, spirochetemia was assessed by utilizing both immunocompetent C3H and immunodeficient SCID mice. However, this study revealed that B. mayonii spirochetemia observed in human patients is not recapitulated in mice. Further, little is known of the classic Lyme disease symptoms – arthritis and carditis – caused by B. mayonii infection. Utilizing the mouse model of acute Lyme disease, C3H mice, these symptoms were assessed by measuring tibiotarsal joints for swelling as well as histopathology of the tibiotarsal joints and heart. The results of this study demonstrate that B. mayonii infection coincides with mild carditis evidenced by infiltration and valvular endocarditis, however infected mice demonstrated no signs of Lyme arthritis by tibiotarsal joint swelling or histopathology. This demonstrates that B. mayonii causes different pathologies from B. burgdorferi and requires further studies to fully elucidate the host-pathogen interactions.

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