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The longevity and infectivity of isolated populations of Plasmodium falciparum gametocytes were studied. Following chloroquine treatment gametocyte numbers fell with a constant rate of loss over a period of 16–24days; the populations had a half-life of 2·4 days. The sex ratio stayed constant throughout at 4 female:1 male. The ability of the microgametocytes to exflagellate and the infectivity of the population to mosquitoes persisted for 3 weeks. Antibodies to the gametocytes were detected but not in every patient studied. It was concluded that the gametocytes of P. falciparum are both long-lived and show persistent infectivity to mosquitoes. They can stimulatae antibody production but the immune response appears to play no part in their elimination, which probably takes place in the spleen as a part of the normal process of removing old, damaged and malformed red cells.

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The ultrastructural organization of the mature gametocytes of Plasmo­dium falciparum isolated from the peripheral circulation of naturally in­fected Gambians is examined and compared with immature forms obtained from the peripheral circulation of a chloroquine treated patient. The latter are recognized as the stage 2 and 3 developmental forms (Hawking, Wilson & Gammage 1971 Trans. R. Soc. trop. Med. Hyg . 65, 549-559) observed by light microscopy and are distinguished in the electron microscope by three characters; (i) they do not fill the host cell, (ii) they contain few, if any, osmiophilic bodies, (iii) they possess an extensive subpellicular tubule system. Maturation (capacitation) of these immature parasites takes many days and is followed by an extended period of maturity during which the gametocytes will exflagellate. Mature macro- and microgametocytes have numerous characters in common with the gametocytes of avian and reptilian Plasmodiidae, namely a tripartite pellicle, cristate mitochondria, a comparatively high density of osmiophilic bodies in the macrogametocyte, cytostomal feeding, Golgi body, and persistent nucleolus in the female gametocyte. These similarities together with the unexpected nuclear changes detected in macrogametogenesis suggest that P. falciparum is best considered as pre-dating the ‘malariae’ and ‘vivax’ groups and not as having evolved from them. Light microscopy, scanning and transmission electron microscopy and videotape analyses of gamete formation were undertaken. Nuclei in the mature gametocytes are Feulgen negative but upon activation rapidly become Feulgen positive. The gametes also are Feulgen positive. The crescentic parasites swell to become large spherical cells and escape from the host cell by osmotic or enzymic activity. The microgametocyte undergoes three mitotic divisions during which the chromosomes are sequenti­ally reduced in number such that ca . 7 are incorporated into each gametic nucleus. The microtubule organizing centre (m. t. o. c.), which in the mature gametocyte is associated with the intranuclear body, is attached to the centriolar plaque of the first division spindle. There it differentiates into kinetosomes which act as foci for the polymerization of axonemes. The kinetosomes and axonemes remain attached to the centriolar plaques during division and are segregated synchronously with the genome at each division. Subsequently one axoneme enters each haploid gamete at exflagellation. Exflagellation is accompanied by a significant reduction in microgametocyte volume which is associated with an increase in density of the cytoplasm. The female gametocyte does not decrease in volume but undergoes nuclear changes in which a single pole of an intranuclear spindle is detected. Comparisons are made with macrogametogenesis in avian malarial parasites from which it is suggested that this spindle, if not half of a normal mitotic spindle, is an atavistic trait. The possibility of a meiotic gametic division is discussed but discounted. The activity pattern of the microgamete was found to be similar to that of other malarial parasites, with states of high and low activity or immobility. High activity, which results in rapid movement through the medium, is produced by long wavelength (12 μm), low amplitude (1.1μm) waves generated at ca . 12 waves per second; low activity, which results in contorted gyrating on the spot, is produced by long wavelength (14.1), high amplitude (2.3) waves produced at ca 1 wave per second. Following an initial period of continuous activity the gamete usually alter­nates between high and low activity states. Subsequent low activity and immobility is in turn followed by death. Microgamete activity was pro­foundly affected by the plasma of some patients, presumably as a result of the antigametocyte antibodies present. The microgamete contains a single axoneme, at one end of which lies the kinetosome with the juxtakinetosomal sphere and granule. It is this end which emerges first from the parental gametocyte. A single nucleus is centrally located in many microgametes although 23% are anucleate.

SUMMARYReproducible growth of gametocytes of Plasmodium falciparum in vitro was obtained from ring-stages taken directly from naturally infected patients and from the same material following storage in liquid nitrogen. Progressive sexual differentiation in vitro was examined for a finite period of 9 days in microcultures and was, for convenience, divided into 5 stages using established morphological criteria (Hawking, Wilson & Gammage, 1971). This microculture system was adapted as a bioassay for various anti-metabolites. Drug activity was measured by observing the inhibition of the established pattern of sequential development in experimental as compared to control cultures. Inhibitors used were directed against DNA, RNA and protein metabolism and microtubule assembly. As a result of these studies it is proposed that the sexual cell-cycle of P. falciparum is characterized by 4 phases. (1) A G1 period which lasts only a few hours. (2) The S phase, where DNA synthesis occurs, occupies the remainder of the first 2 days of development – both G1 and S are confined to stage I and II gametocytes. (3) G2, which is subdivided into 2 sections: G2A, characterized by stage II and III gametocytes, in which significant RNA and protein synthesis continue to occur; and G2B, where there is a progressive increase in transcription control resulting in the depression of both RNA and protein synthesis. Nonetheless, continued morphological differentiation occurs in the latter section transforming the parasites to stage IV and the morphologically and functionally mature stage V. The final M phase is marked by the brief and explosive events of gametogenesis, during which further protein synthesis occurs de novo. The proposed cell-cycle is examined as a model for studies on the activity of gametocytocidal compounds.