C-type lectin 4 regulates broad-spectrum melanization-based refractoriness to malaria parasites

Anopheles gambiae melanization-based refractoriness to the human malaria parasite Plasmodium falciparum has rarely been observed in either laboratory or natural conditions, in contrast to the rodent model malaria parasite Plasmodium berghei that can become completely melanized by a TEP1 complement-like system-dependent mechanism. Multiple studies have shown that the rodent parasite evades this defense by recruiting the C-type lectins CTL4 and CTLMA2, while permissiveness to the human malaria parasite was not affected by partial depletion of these factors by RNAi silencing. Using CRISPR/Cas9-based CTL4 knockout, we show that A. gambiae can mount melanization-based refractoriness to the human malaria parasite, which is independent of the TEP1 complement-like system and the major anti-Plasmodium immune pathway Imd. Our study indicates a hierarchical specificity in the control of Plasmodium melanization and proves CTL4 as an essential host factor for P. falciparum transmission and one of the most potent mosquito-encoded malaria transmission-blocking targets.

Sensitization of Chrysosplenetin against Artemisinin Resistant Plasmodium berghei K173 via Blocking Host ABC Transporters Potentially Mediated by NF-κBB p52 or PXR/CAR Signaling Pathway

This study investigated if artemisinin-chrysosplenetin combination (ART-CHR) improved ART antimalarial efficacy against resistant Plasmodium berghei K173 via depressing host ABC transporter and potential molecular mechanism. Parasitaemia% and inhibition% were calculated and gene/protein expressions of ABC transporters or PXR/CAR/NF-κB p52 were detected by Western-blot and RT-qPCR. In vitro transcription of PXR/CAR was studied by dual-luciferase reporter assay. Our data indicated that ART-CHR improved ART efficacy against resistant parasites. P-gp inhibitor verapamil and CHR showed a stronger effect in killing resistant parasites while vehicle and Bcrp inhibitor novobiocin did not. ART activated intestinal ABCB1/ABCG2 and CHR inhibited them. ART decreased Bcrp protein whereas CHR increased it. ART ascended ABCC1/ABCC4/ABCC5 mRNA but ART-CHR descended them. CHR as well as rifampin (RIF) or 5-fluorouracil (5-FU) increased transcription levels of PXR/CAR while showed a versatile regulation on in vivo hepatic and enternal PXR/CAR in Mdr1a+/+ (WT) or Mdr1a-/- (KO) mice infected with sensitive or resistant parasites. Oppositely, hepatic and enteric N-7κB p52 mRNA was conformably decreased in WT but increased in KO-resistant mice. NF-κB pathway should potentially involved in the mechanism of CHR on inhibiting ABC transporters and ART resistance while PXR/CAR play a more complicated role in this mechanism.

Euterpe oleracea fruit (Açai)-enriched diet suppresses the development of experimental cerebral malaria induced by Plasmodium berghei (ANKA) infection

Background Cerebral malaria is one of the most severe complications attributed to protozoal infection by Plasmodium falciparum, gaining prominence in children mortality rates in endemic areas. This condition has a complex pathogenesis associated with behavioral, cognitive and motor sequels in humans and current antimalarial therapies have shown little effect in those aspects. Natural products with antioxidant and anti-inflammatory properties have become a valuable alternative therapeutic option in the treatment of distinct conditions. In this context, this study investigated the neuroprotective effect of Euterpe oleracea (açai) enriched diet during the development of experimental cerebral malaria induced by the inoculation of Swiss albino mice with Plasmodium berghei ANKA strain. Methods After Plasmodium infection, animals were maintained on a feeding with Euterpe oleracea enriched ration and parameters such as survival curve, parasitemia and body weight were routinely monitored. The present study has also evaluated the effect of açai-enriched diet on the blood-brain barrier leakage, histological alterations and neurocognitive impairments in mice developing cerebral malaria. Results Our results demonstrate that between 7th–19th day post infection the survival rate of the group treated with açai enriched ration was higher when compared with Plasmodium-infected mice in which 100% of mice died until the 11th days post-infection, demonstrating that açai diet has a protective effect on the survival of infected treated animals. The same was observed in the brain vascular extravasation, where Evans blue dye assays showed significantly less dye extravasation in the brains of Plasmodium-infected mice treated with açai enriched ration, demonstrating more preserved blood-brain barrier integrity. Açai-enriched diet also attenuate the histopathological alterations elicited by Plasmodium berghei infection. We also showed a decrease of the neurological impairments arising from the exposure of cerebral parenchyma in the group treated with açai diet, ameliorating motor and neuropsychiatric changes, analyzed through the SHIRPA protocol. Conclusion With these results, we conclude that the treatment with açai enriched ration decreased the mortality of infected animals, as well as protected the blood-brain barrier and the neurocognitive deficits in Plasmodium-infected animals.

Antimalarial Activity of Ethanol Extract of Kelakai Leaves (Stenochlaena palustris) to Parasitemia and Splenomegaly in BALB/c Mice Infected with Plasmodium berghei ANKA

Introduction: Malaria is one of global health problems. Splenomegaly is one of malaria symptoms. Antimalarial drug resistance had been reported. Alternative treatment is by using traditional medicinal plants such as kelakai (Stenochlaena palustris). Kelakai contains alkaloid and flavonoid which had been reported to have antimalarial activity. The aim of this study was to discover antimalarial activity of ethanol extract of kelakai leaves to parasitemia and splenomegaly of Plasmodium berghei ANKA in infected BALB/c mice.Methods: This study was based on a modified Peter test using BALB/c mice infected with P. berghei ANKA treated with ethanol extract of kelakai leaves, with chloroquine diphosphate as a positive control. The negative control was P. berghei ANKA infected mice without any additional treatment. Administration of ethanol extract of kelakai leaves was performed for 4 days with a serial doses of 100, 10, and 1 mg/kg body weight. The positive control was given chloroquine diphosphate 20 mg/kg body weight. Parasitemia was observed daily prior to the calculation of the percentage of parasite growth and parasite growth inhibition. At the end of the test, the mice were sacrificed and spleens were isolated to measure their sizes. Probit analysis was performed to obtain ED50 to find the effect of extract in parasite killing by 50%. Spearman test was performed to analyze the correlation of doses of extract and splenomegaly.Results: Parasitemia growth inhibition was directly proportional to the dose. Higher parasitemia inhibition was obtained at higher doses and vice versa. Result of probit analysis showed an ED50 was 77.05 mg/kg body weight. Statistical analysis resulted in insignificant correlation between doses and splenomegaly p = 1.0 (significancy < 0.05).Conclusion: Ethanol extract of kelakai leaves possessed good antimalarial activity and there was no correlation between extract doses and splenomegaly in Plasmodium berghei ANKA-infected mice.

Histological and Weight Changes in the Testes of Plasmodium Berghei- Infected Swiss Mice Treated with Chloroquine

Chloroquine has gained great emphasis in the treatment of malaria. This study sought experimentally to determine the histomorphological and weight changes in the testes of male mice infected with Plasmodium berghei and treated with chloroquine. The study used 30 Swiss mice divided into five groups. Group 1 is Control that was not infected with Plasmodium berghei and not treated with chloroquine as control, Group 2 is Plasmodium (Plasmodiul berghei) Infected animals but not treated, Group 3 is Plasmodium Infected animals + Chloroquine (5mg/kg), Group 4 is Plasmodium Infected animals + Chloroquine (10mg/kg) and Group 5 is Plasmodium Infected animals + Chloroquine (15mg/kg). The mice were treated for 7 days after parasitaemia was confirmed and the Group 2-5 testes studied with reference to Group 1. The results showed that the Group 2 showed a little distortion, difference in spermatogenic activities and increased cellular activities; Group 3 showed large, convoluted tubules, moderate number of spermatids and large interstitial spaces, Group 4 showed Large seminiferous tubules, large spermatids, increased distortion and group 5 showed shrinking of seminiferous tubules, degeneration of interstitial cells of Leydig cells and Sertolic cells with spermatids. Groups 1-5 showed no significant effect in in body weights and testes weights of Swiss mice. Plasmodium berghei (malaria) and chloroquine have effects on histomorphological structures of Swiss mice testes but not on their teste’s weights. The testicular section from Swiss mice infected with malaria and treated at various doses when compared with the Control (Group 1) showed some moderate distortion in some structures like seminiferous tubules, connective tissues between the tubules, lumen and interstitial spaces. It can be deducted that Plasmodium berghei which caused parasitaemia in mice could cause a little tissue effect on mice if not treated.

Histopathological Effects of Seed Oil of Moringa oleifera Lam. on Albino Mice Infected with Plasmodium berghei (NK65)

The study assessed the histopathological effects of seed oil of Moringa oleifera on albino mice infected with Plasmodium berghei. This work included a good idea in the treatment of a causing agent of malaria with Moringa seed oil as bio-natural treatment. Thirty-five mice were divided equally and grouped into five. The mice were acclimatised for seven days and thereafter infected with 0.2 mL Plasmodium berghei (NK65) parasite. The parasites were allowed to establish in the mice for five days before commencement of treatment. Group A - negative control (untreated), group B - positive control (10mg/kg chloroquine treated), group C, D and E were respectively treated with 800, 400, 200 mg/kg seed oil of Moringa oleifera. By oral administration of 0.2 mL of treatment dose, treatment was carried out in four consecutive days and the mice were sacrificed five days thereafter. The liver and kidney extracted from the mice were processed for histological studies. Findings revealed group A had the least packed cell volume (PCV) of 22.23±1.98% and group B had the most PCV of 48.31±1.55% after treatment. The PCV in groups C, D and E were 45.34±1.11%, 41.40±1.00% and 39.19±1.82% respectively after treatment. Coagulative necrosis and inflammation characterised the liver and kidney of mice in groups C and D. Lesions were observed in all the liver of mice treated with the seed oil of M. oleifera and chloroquine. Overall, it can be inferred that the higher the PCV of mice after treatment, the higher the performance of chemotherapeutic agents against parasitaemia. Thus, at 800, 400 and 200 mg/kg dosage, the seed oil of Moringa oleifera could possibly treat malaria. However, administration of a higher dose of the oil and chloroquine should be with caution as both drugs may pose adverse effects on the kidney and liver.

Structural basis of malaria transmission blockade by a monoclonal antibody to gamete fusogen HAP2

HAP2 is a transmembrane gamete fusogen found in multiple eukaryotic kingdoms and is structurally homologous to viral class II fusogens. Studies in Plasmodium have suggested that HAP2 is an attractive target for vaccines that block transmission of malaria. HAP2 has three extracellular domains, arranged in the order D2, D1, and D3. Here, we report monoclonal antibodies against the D3 fragment of Plasmodium berghei HAP2 and crystal structures of D3 in complex with Fab fragments of two of these antibodies, one of which blocks fertilization of Plasmodium berghei in vitro and transmission of malaria in mosquitoes. We also show how this Fab binds the complete HAP2 ectodomain with electron microscopy. The two antibodies cross-react with HAP2 among multiple plasmodial species. Our characterization of the Plasmodium D3 structure, HAP2 ectodomain architecture, and mechanism of inhibition provide insights for the development of a vaccine to block malaria transmission.

Repurposing Dihydroartemisinin-Piperaquine-Doxycycline as an Antimalarial Drug: A Study in Plasmodium berghei-Infected Mice

Artemisinin-based combination (ACT) therapy is the mainstay for malaria treatment. However, Plasmodium parasite with decreased susceptibility to ACT has emerged. Hence, it is imperative to discover new drugs or explore new drug combinations that can decrease Plasmodium parasite resistance. This study assessed the antiplasmodial activity of dihydroartemisinin-piperaquine- doxycycline (D-P-DX) on mice infected with Plasmodium berghei. Swiss albino mice (25-30g) of both sexes inoculated with 1x107 Plasmodium berghei intraperitoneally were used. The mice were randomly grouped and orally treated with DX (2.2 mg/kg), D-P (1.71/13.7 mg/kg) and D-P-DX daily in curative, suppressive and prophylactic studies. The negative and the positive controls were treated daily with normal saline (0.2mL) and chloroquine (CQ) (10mg/kg), respectively. After treatment, blood samples were assessed for percentage parasitemia, hematological and lipid parameters. Also, the mice were observed for mean survival time. D-P, DX, and D-P-DX produced significant decreases in percentage parasitemia at p<0.05, p<0.01 and p<0.001, respectively when compared to negative control. In the curative study, D-P, DX, and D-P-DX produced 64.9%, 71.1%, and 93.6% parasitemia inhibitions when compared to 70.0% inhibition produced by CQ. Plasmodium berghei -induced alterations in packed cell volume, white blood cells, red blood cells, hemoglobin, high-density lipoprotein cholesterol, total cholesterol, low-density lipoprotein cholesterol, and triglyceride levels were significantly restored by DX (p<0.05) and D-P (p<0.01) and D-P-DX (p<0.001) when compared to the negative control. D-P-DX showed significant antiplasmodial activity against Plasmodium berghei- infected mice. It may be clinically useful for the treatment of malaria.