PROGRESS IN PROTOZOOLOGY

Malaria
IN VITRO CULTIVATION OF PARASITIC PROTOZOA 233
Dr. T r a g e r outlined the most important advances made in the
cultivation of Plasmodium spp. since the first report of continuous culture
of the erythrocytic stage of Plasmodium falciparum in human erythrocytes
maintained under appropriate conditions (T r a g e r and Jensen
1976). Subsequently the erythrocytic stages of four species of
malarial parasites of rhesus monkeys have been placed in continuous
culture using similar methods: P. knowlesi (Chen et al. 1980, W i cklam
et al. 1980) with a 24-hour cycle; P. fragile (Chin et al. 1979),
a falciparum-like organism; P. inui (N g u y e n - D i n h et al. 1978)
with a 72-hour cycle; and P. cynomolgi (N g u y e n - D i n h et al. 1980),
a vivax-like parasite. A culture of P. malariae mixed with P. falciparum
in human red cells has been reported by R a i C h o w d h u r i et al.
(1979). The second most important human malarial parasite, P. vivax
has been cultivated by Larrouy et al. (1981) who used methods
similar to those applied to P. falciparum but with enhanced glucose
levels and a change of medium three times daily instead of only once
a day. All the culture systems have involved the use of various modifications
of the petri dish method or the continuous flow technique
originally designed for P. falciparum. The most suitable medium seems
to be a mixture of RPMI 1640 and human serum which, for some
purposes, can be replaced by rabbit serum (Sax and R i e c k m a n
1980) or calf serum with protease peptone (I f e d i b a and V a n d e nberg
1980). Most studies are concentrated on P. falciparum and several
culture lines of this species have been established. There is no indication
of loss of infectivity resulting from prolonged cultivation in vitro.
A morphological change, however, was observed in some culture lines
after more than 18 months in continuous culture (L a n g r e t h et al.
1979). There was a tendency for the loss of the "knobs," — structures
which appear on the membranes of infected erythrocytes when the parasite
becomes a trophozoite.
A further advance was the establishment of cloned populations prepared
by the dilution method or by microscopic selection of samples
with a single organism. These clones seemed to differ with regard to
the appearance of knobs o i the membrane of the host cell containing
trophozoites or late stages (K i 1 e j i a n 1979). Preliminary studies using
clones of a line from Gambia indicated that the knobbed (K + ) clone was
infective and pathogenic in Aotus, whereas the knobless (K~) clone produced
a barely detectable infection. The K+ and K~~ clones differed also
in their in vitro response to chloroquine. It is most probable that the
culture method, combined with cloning offers good material for studies
on the occurrence and nature of drug-resistant strains.
http://rcin.org.pl