Molecular testing of Lyme disease

Abstract

Lyme disease is a tick-vectored multisystemic illness caused by the bacteria Borrelia burgdorferi sensu lato contracted throughout North America and Eurasia. Although Lyme disease is the most common tick vectored illness in North America, many questions about the disease such as pathophysiology and organ involvement remained unresolved. Additionally, persistence of the Borrelia bacteria after antibiotic treatment is controversial and the role of antibiotic tolerant bacteria in causing chronic Lyme disease symptoms is even less well understood. The reliance on serological testing in Lyme disease leaves many questions unanswered and accurate diagnosis of Lyme disease in certain situations is challenging. To address some of these outstanding questions, direct detection of Borrelia from biopsy and autopsy tissue and culturing of Borrelia from patients showing persistent Lyme disease symptoms can be valuable. In this study, I investigated autopsy tissues using the DNA-based detection methods of nested PCR and Fluorescent in situ hybridization (FISH) and the protein-based technique of immunohistology from seropositive, seronegative, and serologically untested individuals. This study showed that Borrelia was widely distributed in different organs in both seropositive and seronegative participants. These techniques were then extended to biopsy tissue samples. My findings showed that B. burgdorferi is pleiomorphic, with FISH and immunohistology showing predominantly round body and biofilm forms with fewer in the spirochetal form. Finally, I showed that culture of viable Borrelia is possible from different body fluids from participants with persistent Lyme disease symptoms. This finding strengthens the evidence that chronic Lyme disease is associated with an active and long-standing Borrelia infection. Findings from this study could be extended to investigation of other tick-vectored diseases and reinforce the potential for the use of direct molecular methods in disease diagnosis.