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Updated: Dec 5, 2025
Cultivation Methods of Spirochetes from Borrelia burgdorferi Sensu Lato Complex and Relapsing Fever Borrelia
Published on: November 25, 2022
Borrelia miyamotoi strain LB-2001 retains plasmids and infectious phenotype throughout continuous culture passages as
Robert D Gilmore1, Sierra Mikula1, Emma K Harris1
1Bacterial Diseases Branch, Division of Vector-Borne Diseases, National Center for Emerging and Zoonotic Infectious Diseases, Centers for Disease Control and Prevention, Fort Collins, CO, USA.
Abstract:
Borrelia miyamotoi is a tick-borne spirochete of the relapsing fever borrelia group and an emerging pathogen of public health significance. The genomes of relapsing fever borreliae and Lyme disease borreliae consist of multiple linear and circular plasmids in addition to the chromosome. Previous work with B. burgdorferi sensu lato found diminished infectivity upon continuous in vitro culture passage that was attributable to plasmid loss. The effect of long-term culture passage on B. miyamotoi is not known. We generated a series of plasmid-specific primer sets and developed a multiplex PCR assay to detect the 14 known plasmids of B. miyamotoi North American strains LB-2001 and CT13-2396. We assessed the plasmid content of B. miyamotoi LB-2001 over 64 culture passages spanning 15 months and determined that strain LB-2001 retained all plasmids upon prolonged in vitro cultivation and remained infectious in mice. We also found that strain LB-2001 lacks plasmid lp20-1 which is present in strain CT13-2396. These results suggest that B. miyamotoi remains genetically stable when cultured and passaged in vitro.
Insights
Borrelia miyamotoi, an emerging tick-borne pathogen, maintains all its plasmids and infectivity after extensive laboratory cultivation. This indicates genetic stability during in vitro passage, crucial for studying this public health concern.
Area of Science:
- Microbiology
- Infectious Diseases
- Genetics
Background:
- Borrelia miyamotoi is an emerging tick-borne spirochete with public health significance.
- Relapsing fever borreliae, including B. miyamotoi, possess complex genomes with multiple plasmids.
- Previous studies on B. burgdorferi showed reduced infectivity after in vitro culture due to plasmid loss.
Purpose of the Study:
- To investigate the effect of long-term in vitro culture passage on the genetic stability and infectivity of Borrelia miyamotoi.
- To determine if B. miyamotoi undergoes plasmid loss during prolonged cultivation, similar to other Borrelia species.
Main Methods:
- Developed plasmid-specific primer sets for B. miyamotoi.
- Created a multiplex PCR assay to detect 14 known plasmids in North American strains.
- Monitored plasmid content and infectivity of B. miyamotoi strain LB-2001 over 64 culture passages (15 months) and in mouse models.
Main Results:
- Borrelia miyamotoi strain LB-2001 retained all its plasmids throughout 64 passages in vitro.
- The strain remained infectious in mice after prolonged laboratory cultivation.
- Strain LB-2001 was found to lack plasmid lp20-1, which is present in strain CT13-2396.
Conclusions:
- Borrelia miyamotoi demonstrates significant genetic stability during extended in vitro culture and passage.
- Prolonged cultivation does not lead to plasmid loss or diminished infectivity in B. miyamotoi.
- These findings are important for understanding B. miyamotoi pathogenesis and for developing reliable in vitro models for research.
