Pleomorphism and Viability of the Lyme Disease Pathogen Borrelia burgdorferi Exposed to Physiological Stress Conditions: A Correlative Cryo-Fluorescence and Cryo-Scanning Electron Microscopy Study

Abstract

To understand the response of the Lyme disease spirochete Borrelia burgdorferi exposed to stress conditions and assess the viability of this spirochete, we used a correlative cryo-fluorescence and cryo-scanning microscopy approach. This approach enables simple exposition of bacteria to various experimental conditions that can be stopped at certain time intervals by cryo-immobilization, examination of cell viability without necessity to maintain suitable culture conditions during viability assays, and visualization of structures in their native state at high magnification. We focused on rare and transient events e.g., the formation of round bodies and the presence of membranous blebs in spirochetes exposed to culture medium, host sera either without or with the bacteriolytic effect and water. We described all crucial steps of the workflow, particularly the influence of freeze-etching and accelerating voltage on the visualization of topography. With the help of newly designed cryo-transport device, we achieved greater reproducibility.

Lyme Disease Frontiers: Reconciling Borrelia Biology and Clinical Conundrums

“…[311,312], and Borrelia spp. [270,[313][314][315]. Indeed, a report in the 1990s of atypical borrelial morphologies recovered from human specimens spurred interest in the phenotypic plasticity of this organism [316].…”

“…In light of these findings, it is speculated that pleomorphy plays a role in the pathology of LD and that these diverse morphological forms may confer, or contribute to, improved persistence within the human host [270,313,314,317,318]. There are many studies that demonstrate the in vitro pleomorphic capabilities of Borrelia burgdorferi [313][314][315][319][320][321], and a few reports that have identified similar forms in clinical samples [313,316,322]. Pleomorphism can encompass a variety of morphologies, so it is important to develop consistent terminology when referring to these forms.…”

Development of a sensitive molecular diagnostic assay for detectingBorrelia burgdorferiDNA from blood of Lyme disease patients by digital PCR

“…We did observe a gradual clearing of the signal as the antibiotic treatment continued in 2016 patient samples, as compared to 2018 patient samples. It is difficult to explain why this happened but in patient samples the integrity of the spirochetes is quite questionable: they can be fragmented, can form round bodies or blebs or even be hiding in biofilms [25,26,27]. This could lead to variability in extraction of spirochetal DNA from such patient samples using regular DNA extraction kits.…”

The need to unravel the twisted nature of the Borrelia burgdorferi sensu lato complex across Europe

“…The complement system is one of the major innate defence mechanisms B. burgdorferi has to overcome to establish a stable infection of the mammalian host. Different genospecies of B. burgdorferi differ in their sensitivity to human and animal sera [81,83]. A number of borrelial polymorphic proteins may be involved in host-to-host differences in complement evasion.…”