On the other hand, the higher persister frequencies for B. burgdorferi than E. coli could indicate that B. burgdorferi may form persisters more readily or reflect differences in the speed of growth of the organisms, the age of culture when Eticlopride hydrochloride antibiotic is added, and the dilution factor which affects the number of persisters carried over during the subculture. In addition, we found that the persister frequencies vary according to antibiotic exposure, with the more effective antibiotic ceftriaxone having a lower persister frequency than amoxicillin, a finding that is consistent with previous studies. It remains to be determined if there are differences in persistence of B. burgdorferi strains and if the high persister frequencies in B. burgdorferi strains are associated with recalcitrance to antibiotic therapies. In conclusion, we found there is a hierarchy of B. burgdorferi persisters with increasing antibiotic tolerance as the culture ages from log phase to stationary phase with morphological changes from spirochetal form to round body and microcolony forms. Importantly, we identified drug combinations that have high activity against B. burgdorferi persisters with daptomycin- containing combinations achieving the best activity. The most effective drug combination used daptomycin, cefoperazone and doxycycline which appeared to render resistant microcolony forms of B. burgdorferi unable to resuscitate viability upon subculture, a feature not previously described using any other antibiotic singly or in combinations. While important to state that the role of any persister organisms in human disease is far from elucidated, these findings may have implications for the ESI 09 treatment of certain Lyme disease patients with slow to resolveor antibiotic-refractory arthritis or possibly stubborn ongoing symptoms. Direct extrapolation of these in vitro findings to human treatment would be unwise and premature. Future studies are needed to confirm whether such combination drug therapy yields benefit in animal models and possibly then in clinical studies. While monitoring the phosphorylation status at both S716 and S814 on non-phosphorylatable Kif23-iso1 mutants, we could uncover an interplay between these two sites. Namely, mutation of S716 drastically reduced phosphorylation of the distant S814, but not the opposite. This would imply that phosphorylations at S716 and S814 obey to a hierarchical order in Kif23-iso1, whereby S716 phosphorylation would occur first and be necessary for efficient phosphorylation at S814. We note that this cross-talk between these two phosphorylation events was not observed in our in vitro NDR/LATS kinase assay using a 20 kDa Kif23 fragment.