PROGRESS IN PROTOZOOLOGY

2.32
I. CUNNINGHAM
Chemically defined media are essential for metabolic studies. In
1957, Dr. Trager devised a medium for the in vitro cultivation of
L. tarentolae, a parasite of lizards. Steiger and colleagues (1976,
1977) designed a medium which could support the growth of large
populations of L. donovani and L. braziliensis. Another defined medium
described by Be r ens and his co-workers (1978) was based on their
earlier HO-MEM mixture.
(2) Amastigotes
These are round to oval, intracellular organisms which occur in the
vertebrate host and can only be grown in vitro at host body temperature.
Until recently, amastigotes were difficult to collect or cultivate in
sufficient quantities for physiological, biochemical or immunological
investigations. Limited studies of morphology and 02 uptake during
transformation of amastigotes into promastigotes have been reported
by Simpson (1968) and Brun and Krassner (1976).
Mammalian cell cultures can support the growth of amastigote stages
with varying degrees of success. The culture system of Walton
et al. (1972) could provide enough amastigotes grown in monolayers of
VERO (African green monkey kidney) cells for the immunofluorescent
antibody test (IFAT), biochemical and metabolic studies. Cells derived
from dog sarcoma and hamster peritoneum and infected by promastigotes
of L. tropica, L. mexicana and L. donovani can be used in drug
assays (Mattock and Peters 1975). A more authentic system for
testing drug activity was described by Berman et al. (1979) in which
amastigotes became established and multiplied in human peripheral
macrophage cultures: the parasites reside in the macrophages in an
infected human host. Promastigotes of Leishmania isolated from patients
in Schneider's medium for less than a few weeks were infective for human
macrophage cells in vitro. Using this macrophage-parasite in vitro
system, it has been shown that patients with a clinical resistance to
antimony harbor parasites that are drug resistant in vitro. This method
was applied to visceral Leishmania from Kenya and cutaneous leishmaniasis
from Panama.
A technique has been devised for the routine in vitro transformation
of promastigotes into amastigote-like forms of L. braziliensis panamensis
and L. mexicana. Promastigotes which have entered the log phase of
growth in Schneider's medium at 24°C were collected by centrifugation
and transferred to medium TC 199 supplemented with 30°/o foetal bovine
serum and incubated at 37°C. Within 48-96 h approximately 95%
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