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The objectives of the present study were to investigate the risk of B. burgdorferi s.1. (Bb)-transmission by I. ricinus-nymphs to a host (i) after different periods of feeding, and (ii) with regard to the particular method of tick removal. On each of 72 Mongolian gerbils 3 tick nymphs taken from a highly infected batch were allowed to feed in a small capsule. Feeding ticks were removed 16.7, 28.9, 47.0, and 65.2 hrs post-attachment. In each of these 4 groups 3 sub-groups with 6 gerbils each were deticked by (a) pulling ticks out with forceps without any pretreatment, (b) pulling ticks out after 3 min of intensive squeezing, and (c) applying nail polish to ticks 1.1 hrs before removal. The infection status in each gerbil was subsequently determined by larval xenodiagnosis. All gerbils with ticks removed > or = 47 hrs post-attachment were found to be infected. After 16.7 hrs as well as after 28.9 hrs of tick feeding, approximately 50% of the gerbils had acquired a transmissible infection, thus Bb-transmission to a host may even occur in the early phases of I. ricinus feeding. There is no evidence from this study that the tick removal method used has any significant influence on a host’s Bb-infection risk.