BackgroundThe biology of adult tsetse (Glossina spp), vectors of trypanosomiasis in Africa, has been extensively studied – but little is known about larviposition in the field.Methodology/Principal FindingsIn September-November 1998, in the hot-dry season in Zimbabwe’s Zambezi Valley, we used artificial warthog burrows to capture adult females as they deposited larvae. Females were subjected to ovarian dissection and were defined as perinatal flies, assumed to have entered burrows to larviposit, if oocyte sizes indicated >95% pregnancy completion. Perinatal flies were defined as full-term pregnant if there was a late third instar larva in utero, or postpartum if the uterus was empty. All other females were defined as pre-full-term pregnant (pre-FT). Of 845 G. m. morsitans captured, 91% (765) were female and 295/724 (41%) of females dissected were perinatal flies. By contrast, of 2805 G. pallidipes captured only 71% (2003) were female and only 33% (596/1825) of females were perinatal. Among all perinatal females 67% (596/891) were G. pallidipes. Conversely, in burrows not fitted with traps – such that flies were free to come and go – 1834 (59%) of pupae deposited were G. m. morsitans and only 1297 (41%) were G. pallidipes. Thus, while more full-term pregnant G. pallidipes enter burrows, greater proportions of G. m. morsitans larviposit in them, reflecting a greater discrimination among G. pallidipes in choosing larviposition sites. Catches of males and pre-FT females increased strongly with temperatures above 32°C, indicating that these flies used burrows as refuges from high ambient temperatures. Conversely, catches of perinatal females changed little with maximum temperature but declined from late September through November: females may anticipate that burrows will be inundated during the forthcoming wet season. Ovarian age distributions of perinatal and pre-FT females were similar, consistent with all ages of females larvipositing in burrows with similar probability.Conclusions/SignificanceArtificial warthog burrows provide a novel method for collecting tsetse pupae, studying tsetse behaviour at larviposition, assessing the physiological status of female tsetse and their larvae, and of improving understanding of the physiological dynamics of terminal pregnancy, and population dynamics generally, with a view to improving methods of trypanosomiasis control.