The presently disclosed subject matter provides methods, compositions, and kits for assessing the viability of bacteria from the Borrelia genus, assessing the antibiotic susceptibility of bacteria from the Borrelia genus, and identifying compounds with anti-persister activity for bacteria from the Borrelia genus. Methods for inhbiting the growth and/or survival of bacteria from the Borrelia genus and for treating Lyme disease in a subject are also are provided.