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A technique for inoculating and removing substances via the anus of vector ticks was devised to define features of vector competence precisely. Calibrated inocula (greater than 5 nanoliter) containing aqueous dye and polystyrene beads as well as infectious agents were infused into the rectal sacs of ticks using glass microcapillary pipettes placed within the expanded anal orifice. The guts of preadult and adult ticks, Ixodes dammini Spielman, Clifford, Piesman & Corwin, Dermacentor variabilis (Say), Hyalomma impeltatum Schulze & Schlottke, and Amblyomma americanum (L.), were thereby infused with these inocula. Distribution of inocula was determined by examining hemolymph and sectioned ticks and confirmed that material placed in the rectal sac spread throughout the midgut diverticula. Ticks survived for greater than 6 mo after this procedure and were able to feed, molt to the next stage, or oviposit. In contrast, fewer ticks survived after intracelomic inoculation. The course of infection in ticks receiving anal infusions of Borrelia burgdorferi (the
Lyme disease spirochete) was assessed. Such infections appear to differ from those established by feeding on infected hosts. Contents of the tick gut can be sampled nondestructively by anal perfusion to diagnose infection by this spirochete.