Histochemical observations on the exoerythrocytic malaria parasite Plasmodium berghei in rat liver

Summary Enzyme histochemical methods were performed on sporozoite infected liver tissue of rats in order to gain insight into the nutrition and metabolism of exoerythrocytic forms of Plasmodium berghei. The following enzymes were demonstrated in the hepatocytic stages of the parasites, obtained 41 and 48 h after inoculation of sporozoites: acid phosphatase, cytochrome oxidase, NADH-tetrazolium reductase, succinate dehydrogenase, NAD⁺ and NADP⁺ dependent isocitrate dehydrogenase, NADP⁺-dependent malate dehydrogenase, lactate dehydrogenases, 6-phosphogluconate dehydrogenase and glucose-6-phosphate dehydrogenases and -glycerol-phosphate dehydrogenase. The results suggest that a conventional Embden-Meyerhoff pathway, pentose phosphate pathway and Krebs' citric acid cycle may in part be present in these exoerythrocytic parasites. Alkaline phosphatase, nucleoside polyphosphatase, 5nucleotidase. glucose-6-phosphatase, -glucan phosphorylase, NAD⁺ dependent malate dehydrogenase, amino-peptidase M and non-specific esterases were not detected by our techniques in the parasite. The enzyme distribution of this intrahepatocytic malaria parasite revealed by histochemistry is compared with the enzyme distribution in the other phases of the parasite's life cycle.This study was made possible by grants from the Jan Dekker Foundation for Biomedical Research and the Niels Stensen Foundation, The Netherlands, to the first author     

Histochemical observations on the exoerythrocytic malaria parasite Plasmodium berghei in rat liver

Enzyme histochemical methods were performed on sporozoite infected liver tissue of rats in order to gain insight into the nutrition and metabolism of exoerythrocytic forms of Plasmodium berghei. The following enzymes were demonstrated in the hepatocytic stages of the parasites, obtained 41 and 48 h after inoculation of sporozoites: acid phosphatase, cytochrome oxidase, NADH-tetrazolium reductase, succinate dehydrogenase, NAD+ and NADP+ dependent isocitrate dehydrogenase, NADP+-dependent malate dehydrogenase, lactate dehydrogenases, 6-phosphogluconate dehydrogenase and glucose-6-phosphate dehydrogenases and alpha-glycerol-phosphate dehydrogenase. The results suggest that a conventional Embden-Meyerhoff pathway, pentose phosphate pathway and Krebs' citric acid cycle may in part be present in these exoerythrocytic parasites. Alkaline phosphatase, nucleoside polyphosphatase, 5' nucleotidase, glucose-6-phosphatase, alpha-glucan phosphorylase, NAD+ dependent malate dehydrogenase, amino-peptidase M and non-specific esterases were not detected by our techniques in the parasite. The enzyme distribution of this intrahepatocytic malaria parasite revealed by histochemistry is compared with the enzyme distribution in the other phases of the parasite's life cycle.     

Iron deficiency influences the course of malaria in Plasmodium berghei infected mice

Iron deficiency accelerates suicidal erythrocyte death, which is evident from phosphatidylserine exposure. The present study explored whether iron deficiency compromises intraerythrocytic growth of Plasmodium and enhances death of infected erythrocytes thus influencing the course of malaria. As a result, phosphatidylserine exposure is increased in Plasmodium falciparum infected human erythrocytes, an effect significantly more marked in iron deficiency. Moreover, iron deficiency impairs in vitro intraerythrocytic growth and infection of erythrocytes. In mice, iron-deficient erythrocytes are more rapidly cleared from circulating blood, an effect increased by infection with Plasmodium berghei. Parasitemia in P. berghei infected mice was significantly decreased (from 54% to 33% of circulating erythrocytes 20 days after infection) and mouse survival significantly enhanced (from 0% to 20% 30 days after infection) in iron-deficient mice. In conclusion, iron deficiency favourably influences the course of malaria, an effect partially due to accelerated suicidal death and subsequent clearance of infected erythrocytes.     

Functional equivalence of structurally distinct ribosomes in the malaria parasite, Plasmodium berghei

Unlike most eukaryotes, many apicomplexan parasites contain only a few unlinked copies of ribosomal RNA (rRNA) genes. Based on stage-specific expression of these genes and structural differences among the rRNA molecules it has been suggested that Plasmodium spp. produce functionally different ribosomes in different developmental stages. This hypothesis was investigated through comparison of the structure of the large subunit rRNA molecules of the rodent malaria parasite, Plasmodium berghei, and by disruption of both of the rRNA gene units that are transcribed exclusively during development of this parasite in the mosquito (S-type rRNA gene units). In contrast to the human parasite, Plasmodium falciparum, we did not find evidence of structural differences in core regions of the distinct large subunit rRNAs which are known to be associated with catalytic activity including the GTPase site that varies in P. falciparum. Knockout P. berghei parasites lacking either of the S-type gene units were able to complete development in both the vertebrate and mosquito hosts. These results formally exclude the hypothesis that two functionally different ribosome types distinct from the predominantly blood stage-expressed A-type ribosomes, are required for development of all Plasmodium species in the mosquito. The maintenance of two functionally equivalent rRNA genes might now be explained as a gene dosage phenomenon.     

Plasmodium berghei: development of an irreversible experimental malaria model in Wistar rats

A protocol to infect five-week-old Wistar rats by Plasmodium berghei which resulted in 100% mortality was developed in this work. In order to accomplish this goal, the effect of the administration of 10(7) and 10(8) parasitized erythrocytes by i.v. and i.p. route was investigated. The animals inoculated with 10(7) parasitized red blood cells by i.p. and i.v. routes showed 25 and 50% mortality, respectively. Inoculation with 10(8) parasitized erythrocytes by both routes resulted in a 100% lethal infection. The i.v. inoculation showed less scattered results and it was preferred over the i.p. route. The suitability of the protocol developed was evaluated by treating infected Wistar rats with chloroquine (30 mg/kg/day). A decreased parasitemia after the treatment was observed until the complete eradication of the parasite, around 10 days post-inoculation. Parasitemia depression after chloroquine treatment demonstrates the utility of the model developed to test new antimalarial drugs.