The Plasmodium berghei serine protease PbSUB1 plays an important role in male gamete egress.

The Plasmodium subtilisin-like serine protease SUB1 is expressed in hepatic and both asexual and sexual blood parasite stages. SUB1 is required for egress of invasive forms of the parasite from both erythrocytes and hepatocytes, but its subcellular localisation, function, and potential substrates in the sexual stages are unknown. Here, we have characterised the expression profile and subcellular localisation of SUB1 in Plasmodium berghei sexual stages. We show that the protease is selectively expressed in mature male gametocytes and localises to secretory organelles known to be involved in gamete egress, called male osmiophilic bodies. We have investigated PbSUB1 function in the sexual stages by generating P. berghei transgenic lines deficient in PbSUB1 expression or enzyme activity in gametocytes. Our results demonstrate that PbSUB1 plays a role in male gamete egress. We also show for the first time that the PbSUB1 substrate PbSERA3 is expressed in gametocytes and processed by PbSUB1 upon gametocyte activation. Taken together, our results strongly suggest that PbSUB1 is not only a promising drug target for asexual stages but could also be an attractive malaria transmission-blocking target.

Predicting transmission blocking potential of anti-malarial compounds in the Mosquito Feeding Assay using Plasmodium falciparum Male Gamete Inhibition Assay.

Plasmodium falciparum Standard Membrane Feeding Assay (PfSMFA) is the current gold standard mosquito based confirmatory transmission blocking (TrB) assay for human malaria. However, owing to its complexity only selected gametocytocidal molecules are progressed into SMFA. Predictive tools for evaluation of TrB behavior of compounds in SMFA would be extremely beneficial, but lack of substantially large data sets from many mosquito feeds preempts the ability to perform correlations between outcomes from in vitro assays and SMFA. Here, a total of 44 different anti-malarial compounds were screened for inhibitory effect on male gamete formation in exflagellation inhibition assay (EIA) and the same drug-treated parasites were fed to mosquitoes in SMFA. Regression analysis was performed between outcomes of the two assays and regression models were applied to a randomly selected validation set of four compounds indicating no overfitting and good predictive power. In addition, the pIC50 for 11 different compounds obtained in the EIA was also correlated with pIC50's in SMFA. Resulting regression models provided pIC50 predictions in SMFA with reasonably good accuracy thereby demonstrating the use of a simple in vitro assay to predict TrB of molecules in a complex mosquito based assay.

Genome analysis and polar tube firing dynamics of mosquito-infecting microsporidia.

Microsporidia are highly divergent fungi that are obligate intracellular pathogens of a wide range of host organisms. Here we review recent findings from the genome sequences of mosquito-infecting microsporidian species Edhazardia aedis and Vavraia culicis, which show large differences in genome size, although similar numbers of predicted genes. We also show a video of E. aedis polar tube firing, which is the dramatic mechanism used by microsporidia to deliver the germ cell (sporoplasm) into the host cell to initiate intracellular infection.

The flagellum in malarial parasites.

The malarial parasites assemble flagella exclusively during the formation of the male gamete in the midgut of the female mosquito vector. The observation of gamete formation ex vivo reported by Laveran (Laveran MA: De la nature parasitaire des accidents de l'impaludisme. Comptes Rendues De La Societe de Biologie. Paris 1881, 93:627-630) was seminal to the discovery of the parasite itself. Following ingestion of malaria-infected blood by the mosquito, microgamete formation from the terminally arrested gametocytes is exceptionally rapid, completing three mitotic divisions in just a few minutes, and is precisely regulated. This review attempts to draw together the diverse original observations with subsequent electron microscopic studies, and recent work on the signalling pathways regulating sexual development, together with transcriptomic and proteomic studies that are paving the way to new understandings of the molecular mechanisms involved and the potential they offer for effective interventions to block the transmission of the parasites in natural communities.

The course of infections and pathology in immunomodulated NOD/LtSz-SCID mice inoculated with Plasmodium falciparum laboratory lines and clinical isolates.

Human chimeras are potentially invaluable models for hemoprotozoan parasites such as Plasmodium falciparum. The work presented assesses the susceptibility of immunomodulated NOD/LtSz-SCID mice to genetically distinct P. falciparum parasites. To this end, mice grafted with human erythrocytes were inoculated with two P. falciparum laboratory lines, 3D7 and Dd2 and four clinical isolates, ISCIII-230, ISCIII-231, ISCIII-381 and ISCIII-399. The results showed that, without a previous period of parasite adaptation, 100% of the inoculated mice developed an infection, generally self-limited, though some mice died. The parasitemias ranged from 0.05 to 8% and lasted an average of 19 days (15-26 days) depending on the line or isolate studied. Sexual forms of different maturity, stage II-IV and mature gametocytes were observed in the peripheral blood of mice in 22, 50, 25, 72 and 80% of the mice infected with Dd2, ISCIII-399, ISCIII-230, ISCIII-231 and ISCIII-381 isolates, respectively. The study of the clinical symptoms, the haematological parameters and the histopathological changes in the infected mice showed that most of the malaria features were present in the infected mice except that the sequestration of infected erythrocytes was absent or at most a minor phenomenon, as also indicated by the presence of mature forms of the parasites in the peripheral blood. This study shows that the human chimeras allow the complete asexual and sexual erythrocytic cycle of different P. falciparum lines and clinical isolates to be observed in vivo. It opens a new way to investigate any parasite population in terms of infectivity, transmission, and drug resistance.

Parasitic castration of Eurhomalea lenticularis (Bivalvia: Veneridae) by a digenetic trematode: quantitative histological analysis.

The clam Eurhomalea lenticularis may be parasitized by digenean trematodes of the family Plagiorchidae, specifically in the gonads (parasitic castration). A quantitative histological analysis of the parasitized gonads demonstrated a significant decrease in gonadal area, in the size of individual acini, and in the numbers of differentiated germ cells compared to unparasitized clams. Castration may be caused by mechanical compression due to trematode sporocyst growth. However, the uniform loss of germ cells in areas without sporocysts suggests that a more generalized mechanism is responsible. We suggest that parasitic castration has a primary effect on the host's neuroendocrine and gametogenic systems that regulate gamete production.