Biology Reference
In-Depth Information
FIGURE 6.6
Thatch panels used in the stimulated Ýeld study at MERTU/G. Guatemala. Thatch panels constructed
of materials from the ChagasÔ diseaseÏendemic region of Olopa, Guatemala. CRUZIGARD droplets are visible
as black spots. [Adapted from Durvasula, R.V., Kroger, A., Goodwin, M., Panackal, A., Kruglov, O., Taneja, J.,
Gumbs, A., Richards, F.F., Beard, C.B., and Cordon-Rosales, C. (1999b).
Ann. Entomol. Soc. Am.
92:
937Ï943.]
were further tested by replica plating of gut contents. Between 89 and 96% of total CFUs of
R.
rhodnii
in these bugs were comprised of genetically altered bacteria.
This study demonstrated the efÝcacy of CRUZIGARD under conditions of microbial compe-
tition. Though genetically altered
R. rhodnii
were present in only 50% of the target bugs, they
comprised nearly 100% of the total CFUs in the bug, when infection was successful. Mixed
infections, involving transformed and wild-type
R. rhodnii
or other environmental microbes, were
not found. Previously, in experiments involving sequential infection of aposymbiotic Ýrst-instar
nymphs of
Rhodnius prolixus
with wild-type and recombinant bacteria, we determined that the
initial infection established as the predominant gut organism throughout maturation of the bug.
Data from the Guatemala trial suggest that in 50% of the target bugs, recombinant
Rhodococcus
rhodnii
were successfully established as the predominant gut bacteria. Perhaps a larger number of
Ýrst-instar larvae were exposed to the transgenic bacteria. This, however, cannot be conÝrmed with
this study. Nevertheless, this study provided evidence that the CRUZIGARD mechanism of gene
delivery could be used under conditions that simulated the Ýeld environment.
FOREIGN GENE SPREAD UNDER SPATIALLY
ACCURATE CONDITIONS
Initial studies with CRUZIGARD were conducted in small laboratory chambers that maximized
the likelihood of contact between emerging nymphs of
Rhodnius prolixus
and recombinant bacteria.
To test efÝcacy of CRUZIGARD under conditions that approximate the dimensions of a real hut,
we are currently conducting trials in a greenhouse on the Centers for Disease Control campus in
Chamblee, Georgia. An environmentally controlled greenhouse has been Ýtted for arthropod con-
tainment with multiple screen barriers
(Figure 6.7).
A plywood ÑhutÒ measuring 5 5 8 has
been constructed. The ÑthatchÒ roof of the hut has been treated with CRUZIGARD impregnated
with
Rhodococcus rhodnii
expressing the foreign gene c-galactosidase. Adult
Rhodnius prolixus
carrying wild-type
Rhodococcus rhodnii
have been introduced to the hut, and we are assaying F1
progeny for spread of recombinant
R. rhodnii
. These studies will provide valuable information
about the efÝcacy of CRUZIGARD under spatially accurate conditions, ultimate dosing of trans-
genic bacteria in the CRUZIGARD formulation, and the possible role of CRUZIGARD baited with
attractants in a gene-delivery system.
