Immune Evasion by Parasites Causing Tropical Diseases 
phages, where they reside as spherical amasti- 
gotes within acidic phagolysosomes. Thus the 
parasites evade the immune response by "hid- 
ing" inside macrophages — one of several cell 
types of the immune system that normally help to 
destroy foreign pathogens and substances. When 
a sandfly ingests amastigote-laden macrophages 
during a blood meal, the life cycle is completed. 
With a grant from the National Institutes of 
Health, we are studying how these organisms can 
survive in hostile environments as diverse as a 
sandfly midgut and an acidic macrophage 
phagolysosome. 
A major glycoprotein on the surface of both 
promastigotes and amastigotes is a metal lopro- 
tease of about 63 kDa (gp63) . This enzyme partic- 
ipates in the promastigotes' uptake by macro- 
phages and contributes to the amastigotes' 
survival within them. The amount of gp63 on the 
surface of promastigotes increases about 1 0-fold 
as the parasites develop into the highly in- 
fectious, virulent form during growth in culture. 
We have discovered that three different RNA spe- 
cies encoding gp63 occur during cultured pro- 
mastigote growth. 
One RNA species occurs in the promastigotes 
only during their early, logarithmic growth when 
they are less infectious and have a small amount 
of gp63 on their surface. This gp63 RNA is re- 
placed by another gp63 RNA species as the pro- 
mastigotes enter stationary phase and become 
more infectious. The presence of this second RNA 
correlates with the increased amount of gp63 
protein. The third gp63 RNA species is continu- 
ously present at a low level throughout cultured 
growth and encodes an altered, transmembrane 
form of the protein. 
The three RNA species are derived from a fam- 
ily of about 1 5 different genes, some of which are 
identical and all of which are continuously tran- 
scribed independent of the stage of cultured 
growth. The 3'-untranslated regions of the three 
RNA species have completely different se- 
quences, and we have shown that these se- 
quences contribute to the different steady-state 
amounts of the RNAs at the different growth 
stages. In addition, we are introducing recombi- 
nant DNAs into leishmania to amplify, alter, or 
delete individual gp63 genes so that we may de- 
termine the functions of each of the three differ- 
ent gp63 proteases during the promastigote and 
amastigote stages. One of our long-term goals is 
to alter genetically, or attenuate, the parasite so 
that it can be readily maintained under laboratory 
culture conditions but cannot survive in human 
macrophages. Such parasites may be useful in the 
development of a vaccine against leishmaniasis. 
Onchocerciasis 
Onchocerca volvulus is a filarial nematode 
that causes onchocerciasis, or river blindness, in 
Africa and Latin America. Female O. volvulus 
worms grow to 50 cm in length and reside 
throughout the body within nodules. In the nod- 
ules they produce thousands of microfilariae 
each day that migrate throughout the body and 
enter the eyes, where they produce lesions that 
can lead to blindness. It is not known how any of 
the developmental stages of the parasite evade 
the immune response of an infected person. 
The parasites are difficult to study in the labora- 
tory, because there is no good experimental ani- 
mal model; they infect only humans and 
chimpanzees. 
We have constructed cDNA expression librar- 
ies of the mRNAs from the infective L3 stage lar- 
vae of the parasite, and we are using specific 
cDNA clones to overproduce parasite antigens 
from this larval stage that are recognized by anti- 
sera from onchocerciasis patients. This approach 
has revealed several proteins that are unique to 
the parasite and may be valuable for improved 
diagnosis, treatment, and prevention of the 
disease. 
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