Establishing a Zebrafish Model for Borrelia burgdorferi Infection Using Immersion and Microinjection Methods.

Borrelia burgdorferi is the spirochetal bacterium that causes Lyme disease. Even though antimicrobial sensitivity of B. burgdorferi has been widely studied, there is still a need to develop an affordable, practical, high-throughput in vivo model which can be used to find effective antibiotic therapies, especially for the recently discovered persister and biofilm forms. Here, we describe the immersion and microinjection methods to introduce B. burgdorferi spirochetes into zebrafish larvae. The B. burgdorferi-zebrafish model can be produced by immersing 5-day post-fertilization (dpf) zebrafish in a B. burgdorferi culture, or by injecting B. burgdorferi into the hindbrain of zebrafish at 28 h post-fertilization (hpf). To demonstrate that B. burgdorferi indeed infect the fish, nested polymerase chain reaction (PCR), reverse transcription PCR (RT-PCR), live fluorescence imaging, histological staining, and wholemount immunohistochemical (IHC) methods can be used on B. burgdorferi-infected zebrafish.