For the vast majority of vector borne parasites the ability to overcome the insect midgut defences is central to transmission. However, for many such diseases we know virtually nothing about the molecular mechanisms involved. For vectors such as tsetse flies and sand flies the prospects for rapidly improving our understanding of key interactions occurring in the midgut when challenged by parasites is bleak. This is because the ‘tool box’ required untangling the interactions is very unlikely to be rapidly developed. For example, there is no realistic prospect of producing transgenic technology for tsetse flies because eggs are inaccessible due to intrauterine development of larvae; maintenance of multiple lines of either sand or tsetse flies permitting genetic studies is impossible because of the cost and complexity of culturing colonies; bioinformatic resources are still in their infancy. In this application we suggest that under these circumstances a comparative approach, in which kinetoplastid interactions in Drosophila melanogaster are studied in the first instance, will permit us to make significant progress in understanding the more important cases of insect-parasite interactions (Trypanosome brucei spp in tsetse and Leishmania in sandflies). Herpetomonas ampelophilae is a natural kinetoplastid parasite of Drosophila melanogaster, which establishes infection in the midgut of the fruit fly and can go on to invade the salivary glands. We now have this protozoan in culture and intend, through a combination of genomics, cell biology and RNAi experiments to identify the gut-specific host genomic contingent involved in parasite challenge. In addition, we will study the interaction between the indigenous flora and the parasite and the role of the former in protecting the host from parasite infection. These studies will outline the major immune pathways and interactions by which insects and their gut microflora respond to kinetoplastid challenge in the midgut.