Leveraging <i>Borrelia burgdorferi</i> peptidoglycan chemistry to understand Lyme disease symptom progression

Abstract

With an estimated 476,000 cases each year, Lyme disease (LD) is fast becoming an epidemic in the Northeastern and Midwestern United States. Caused by the spirochete Borrelia burgdorferi, LD is characterized by a wide variety of disease manifestations such as the bull's eye rash or erythema migrans despite the lack of classical virulence factors like lipopolysaccharide, toxins, or secretion systems to transport them. More debilitating late-stage symptoms include Lyme arthritis (LA), acrodermatitis chronica atrophicans, and neuroborreliosis, each strongly associated with particular species of Borrelia. The bacterial peptidoglycan component of the cell wall has been recognized as an antigen in numerous other studies of human pathogens. Indeed, previous studies have shown that the peptidoglycan component of the cell wall alone is sufficient to induce severe LA in a murine model in a matter of days. Furthermore, it has been demonstrated that B. burgdorferi PG persists in host tissue and in the synovial fluid of LA patients, and the unique structure of B. burgdorferi PG may play a key role in that. As B. burgdorferi lacks the machinery to recycle shed PG fragments, the bacterium sheds nearly 50% of its cell wall every generation. All this taken as a whole, investigations into how PG mediates its diverse manifestations are warranted. In this dissertation, a D-Alanyl-D-Alanine carboxypeptidase in an infectious model of B. burgdorferi was characterized and disrupted, causing an alteration of PG chemistry. Using a murine model, we demonstrate that this subsequently results in the near total attenuation of LA, which we confirmed through visual tracking of LA progression and histopathology, and changes in tissue tropism. In addition, this change in PG chemistry causes a change in the binding of peptidoglycan associated proteins (PAPs), particularly P83/100. P83/100 was previously shown to drastically affect bacterial burden in a murine model, and using immunofluorescence, we established that our mutant strain exhibits low abundance of this protein. With this as impetus, we were interested as to whether different pathogenic Borrelia species have distinct PG structural motifs that may explain their different disease manifestations. We employed an -omics approach to analyzing liquid chromatography–mass spectrometry (LCMS) data and have observed that despite conserved amino acids, the PG varies significantly. With currently approved LD diagnostics leaving much to be desired, especially in the detection of early-stage Lyme disease, we sought to leverage the unique PG chemistry of B. burgdorferi and have screened a panel of high affinity and specific antibodies against it which may be the first step in the development of reliable LD diagnostics. We have made an endeavor to leverage Borrelia PG and have provided unique insights into how it may possibly mediate disease progression. Further understanding these mechanisms may potentially reveal novel targets for the specific treatment of LD.