A paratransgenic strategy aimed at decreasing transmission of trypanasoma cruzi, the causative agent of Chagas disease, from reduviid vectors to man is being developed. The symbiotic bacterium, Rhodococcus rhodnii, that resides in the reduviid vector, Rhodnius prolixus, has been transformed to export peptides toxic to T. cruzi and single chain antibodies that may be engineered to target T. cruzi. In this proposal, a strategy to spread engineered symbiotic bacteria in field populations of the vector will be studied. The synthetic fecal paste, CRUZIGARD, that has been formulated to mimic naturally occurring coprophagic methods of symbiont transfer in Rhodnius prolixus will be evaluated as a gene spreading strategy.
In Aim I 'the fitness of engineered symbiotic bacteria to compete with wild-type organisms to establish infection in the host vector will be assessed. In co-infection studies of aposymbiotic R. prolixus (bugs that are specially reared to be free of gut-associated bacteria), effects of different gene constructs on fitness will be determined.
In Aim 2, the potential non-target gene spread will be evaluated. In closed cage-studies uptake and retention of transgenes by arthropods that do not transmit T. cruzi (ants, crickets, fleas and cockroaches) but-may be exposed to CRUZIGARD will be determined. Furthermore, transfer of genetic material from engineered bacteria to other environmental microbes will be characterized.
Aim 3 will involve simulated release of transgeuic microbes in life-size huts that are contained within a greenhouse facility. Huts will be constructed of standard materials and infested with reduviid bugs. Various applications of CRUZIGARD will be used to determine extent of gene spread in a hut and possible hut-to-hut transfer of genetic materials. From these studies, optimal methods for gene delivery via CRUZIGARD can be established.
