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Updated: May 12, 2026
Detecting the Lyme Disease Spirochete, Borrelia Burgdorferi, in Ticks Using Nested PCR
Published on: February 4, 2018
Isolation and Genetic Characterization of the Human Relapsing Fever Spirochete Borrelia persica from a Dog with
Dor Shwartz1, Yaarit Nachum-Biala1, Bar Ben-Shitrit1
1Koret School of Veterinary Medicine, Hebrew University of Jerusalem, P.O. Box 12, Rehovot 7610001, Israel.
Abstract:
Borrelia persica is the causative agent of tick-borne relapsing fever in Israel and is prevalent in the Eastern Mediterranean basin and parts of Asia. Infection with B. persica causes severe illness and potentially life-threatening complications in humans and companion animals. Isolation and in vitro cultivation of B. persica in culture medium is difficult and there are only two previous isolates of this spirochete. Here, we describe the first cultivation of B. persica from an ill dog. Isolation from the dog's blood was performed with the Pettenkofer-LMU Bp medium and spirochetes with vibrant motility and cell density of 2 × 106/ml were observed in culture by dark-field microscopy 3 days after sampling. The isolate was passaged every 3-4 days with cell densities reaching up to 3 × 107/ml achieved over 30 passages. Cryopreservation was made at -80°C without any cryoprotectant additive, and successful growth from thawed culture samples was observed up to 3 months after freezing with repeated freeze and thaw cycles. Generation time during the exponential growth phase was 14.6 hr. Genetic characterization by polymerase chain reaction amplification and DNA sequencing of the flagellin, glycerophosphodiester phosphodiesterase, and 16S rRNA genes indicated that the isolate is a B. persica genotype I strain associated with human relapsing fever. The new canine isolate would be helpful for research on the pathogenesis of relapsing fever and the new modifications in cultivation and preservation methods may assist in future studies of relapsing fever by simplifying previously reported in vitro methods.
Insights
This study reports the first successful cultivation of Borrelia persica, the cause of tick-borne relapsing fever, from an infected dog. New methods simplify its in vitro study, aiding research into this severe illness.
Area of Science:
- Medical Microbiology
- Parasitology
- Veterinary Infectious Diseases
Background:
- Borrelia persica causes severe tick-borne relapsing fever in humans and animals, prevalent in the Eastern Mediterranean.
- Cultivating Borrelia persica in vitro is challenging, with only two prior isolates available for research.
- Infection poses risks of severe illness and potentially fatal complications.
Purpose of the Study:
- To describe the first successful isolation and in vitro cultivation of Borrelia persica from a naturally infected dog.
- To optimize cultivation and cryopreservation techniques for Borrelia persica.
- To genetically characterize the canine isolate and confirm its genotype.
Main Methods:
- Isolation of Borrelia persica from canine blood using Pettenkofer-LMU Bp medium.
- Serial passaging, dark-field microscopy for monitoring growth and motility.
- Cryopreservation at -80°C without cryoprotectants and subsequent viability assessment.
- Genetic analysis via PCR amplification and sequencing of key genes (flagellin, glycerophosphodiester phosphodiesterase, 16S rRNA).
Main Results:
- Successful cultivation of Borrelia persica from an ill dog, achieving high cell densities (up to 3×10^7/ml) over 30 passages.
- Demonstrated viability of the isolate after 3 months of cryopreservation with repeated freeze-thaw cycles.
- Genetic sequencing confirmed the isolate as Borrelia persica genotype I, associated with human relapsing fever.
- Calculated generation time during exponential growth was 14.6 hours.
Conclusions:
- The first canine isolate of Borrelia persica provides a valuable resource for relapsing fever research.
- Optimized cultivation and preservation methods simplify in vitro studies of Borrelia persica.
- This research facilitates further investigation into the pathogenesis and treatment of tick-borne relapsing fever.
