Pools containing varying numbers of black flies were prepared and the heads and bodies separated by freezing and agitation, as previously described [7] . A single O. volvulus L3 was added to each pool. Head and body pools were placed in a 1.5 ml microcentrifuge tube and purified using magnetic silica coated beads (Machery-Nagel GmbH & Co, Bethlehem, PA, USA) following the instructions provided by the manufacturer. In brief, the pools were homogenized in 200 ml of T1 buffer, 25 ml of proteinase K solution provided in the kit was added and the homogenates were incubated at 56uC for 30 minutes. The homogenates were subjected to centrifugation at 13,4006g for 5 minutes at room temperature. A total of 225 ml of the supernatant was transferred to a fresh tube containing 24 ml B-beads (Machery-Nagel) and 360 ml MB2 buffer (Machery-Nagel), and the tube shaken for 5 minutes at room temperature. The magnetic beads were isolated by placing the tubes in a six tube magnetic separator (Dynal MPC-S; Invitrogen). The supernatants were removed and discarded, and 600 ml of MB3 wash buffer was added to each sample. The bead/ DNA complexes were washed by shaking for 5 min at room temperature. The beads were collected in the magnetic separator