Leishmania tropica: studies on the mechanisms of attachment to and entry into macrophages

The in vitro attachment and entry of promastigotes of Leishmania tropica into mouse peritoneal macrophages were studied under defined experimental conditions. The attachment of promastigotes to macrophages was found to be dependent on temperature, macrophage intracellular metabolism, and the normal function of microfilaments and microtubules. Parasite binding was abolished by fixation of macrophages but not of the parasites. Modification of the surface of macrophages and parasites by various surface reactive agents revealed that surface carbohydrates of parasites and macrophages were important in parasite binding. Periodate and neuraminidase affected both parasites and macrophages in a complementary manner. Treatment of macrophages with trypsin and concanavalin-A also prevented parasite attachment. In the extracellular medium, parasite binding required the presence of magnesium, glucose and a macromolecular component of foetal calf serum and was inhibited by 2-deoxyglucose. The requirement for extracellular glucose could be replaced by mannose, suggesting that the requirement for glucose is structural rather than for purposes of energy metabolism. The active fraction of serum was nondialyzable, heat labile and precipitable by ammonium sulfate. The extracellular and surface structural requirements of parasite binding suggest the involvement of a biochemical process which is probably mediated by an enzymatic reaction. Scanning electron microscopic examination of macrophages infected with parasites revealed that motile promastigotes attached to host cells by their flagellar tips to which macrophages responded by producing closely fitting lamellar sheaths and progressively enveloping first the flagellum and then the body of the parasite. Lamellar advance during engulfment was rapid in the first lo minutes but much slower later on. Fully engulfed parasites could be seen after one hour, but most parasites associated with host cells remained extracellular even after four hours. On the other hand, parasites immobilized by fixation or heat shock adhered by either their flagellar or somatic ends. Engulfment proceeded at a steady rate and was complete by two hours. No clear distinction between the attachment and engulfment stages of parasite uptake could be established since both were inhibited by low-temperature, cytochalasin and mild fixation of macrophages. The rheological features of the host cells' response to parasite attachment indicate that invasion by parasites is through phagocytosis rather than penetration. surface structural requirements of parasite binding suggest the involvement of a biochemical process which is probably mediated by an enzymatic reaction. Scanning electron microscopic examination of macrophages infected with parasites revealed that motile promastigotes attached to host cells by their flagellar tips to which macrophages responded by producing closely fitting lamellar sheaths and progressively enveloping first the flagellum and then the body of the parasite. Lamellar advance during engulfment was rapid in the first 10 minutes but much slower later on. Fully engulfed parasites could be seen after one hour, but most parasites associated with host cells remained extracellular even after four hours. On the other hand, parasites immobilized by fixation or heat shock adhered by either their flagellar or somatic ends. Engulfment proceeded at a steady rate and was complete by two hours. No clear distinction between the attachment and engulfment stages of parasite uptake could be established since both were inhibited by low-temperature, cytochalasin and mild fixation of macrophages. The rheological features of the host cells' response to parasite attachment indicate that invasion by parasites is through phagocytosis rather than penetration.