Asymptomatic Plasmodium vivax malaria in the Brazilian Amazon: Submicroscopic parasitemic blood infects Nyssorhynchus darlingi

Individuals with asymptomatic infection due to Plasmodium vivax are posited to be important reservoirs of malaria transmission in endemic regions. Here we studied a cohort of P. vivax malaria patients in a suburban area in the Brazilian Amazon. Overall 1,120 individuals were screened for P. vivax infection and 108 (9.6%) had parasitemia detected by qPCR but not by microscopy. Asymptomatic individuals had higher levels of antibodies against P. vivax and similar hematological and biochemical parameters compared to uninfected controls. Blood from asymptomatic individuals with very low parasitemia transmitted P. vivax to the main local vector, Nyssorhynchus darlingi. Lower mosquito infectivity rates were observed when blood from asymptomatic individuals was used in the membrane feeding assay. While blood from symptomatic patients infected 43.4% (199/458) of the mosquitoes, blood from asymptomatic infected 2.5% (43/1,719). However, several asymptomatic individuals maintained parasitemia for several weeks indicating their potential role as an infectious reservoir. These results suggest that asymptomatic individuals are an important source of malaria parasites and Science and Technology for Vaccines granted by Conselho Nacional de may contribute to the transmission of P. vivax in low-endemicity areas of malaria.

Evaluation of two Plasmodium vivax sexual stage antigens as transmission-blocking vaccine candidates

Background Plasmodium vivax transmission-blocking vaccines (TBVs) are receiving increasing attention. Based on excellent transmission-blocking activities of the PbPH (PBANKA_0417200) and PbSOP26 (PBANKA_1457700) antigens in Plasmodium berghei, their orthologs in P. vivax, PVX_098655 (PvPH) and PVX_101120 (PvSOP26), were selected for the evaluation of their potential as TBVs. Methods Fragments of PvPH (amino acids 22–304) and PvSOP26 (amino acids 30–272) were expressed in the yeast expression system. The recombinant proteins were used to immunize mice to obtain antisera. The transmission-reducing activities of these antisera were evaluated using the direct membrane feeding assay (DMFA) using Anopheles dirus mosquitoes and P. vivax clinical isolates. Results The recombinant proteins PvPH and PvSOP26 induced robust antibody responses in mice. The DMFA showed that the anti-PvSOP26 sera significantly reduced oocyst densities by 92.0 and 84.1% in two parasite isolates, respectively, whereas the anti-PvPH sera did not show evident transmission-reducing activity. The variation in the DMFA results was unlikely due to the genetic polymorphisms of the two genes since their respective sequences were identical in the clinical P. vivax isolates. Conclusion PvSOP26 could be a promising TBV candidate for P. vivax, which warrants further evaluation. Graphical Abstract

Effect of seasonal malaria chemoprevention plus azithromycin on Plasmodium falciparum transmission: gametocyte infectivity and mosquito fitness

Background Seasonal malaria chemoprevention (SMC) consists of administration of sulfadoxine-pyrimethamine (SP) + amodiaquine (AQ) at monthly intervals to children during the malaria transmission period. Whether the addition of azithromycin (AZ) to SMC could potentiate the benefit of the intervention was tested through a double-blind, randomized, placebo-controlled trial. The effect of SMC and the addition of AZ, on malaria transmission and on the life history traits of Anopheles gambiae mosquitoes have been investigated. Methods The study included 438 children randomly selected from among participants in the SMC + AZ trial and 198 children from the same area who did not receive chemoprevention. For each participant in the SMC + AZ trial, blood was collected 14 to 21 days post treatment, examined for the presence of malaria sexual and asexual stages and provided as a blood meal to An. gambiae females using a direct membrane-feeding assay. Results The SMC treatment, with or without AZ, significantly reduced the prevalence of asexual Plasmodium falciparum (LRT X22 = 69, P < 0.0001) and the gametocyte prevalence (LRT X22 = 54, P < 0.0001). In addition, the proportion of infectious feeds (LRT X22 = 61, P < 0.0001) and the prevalence of oocysts among exposed mosquitoes (LRT X22 = 22.8, P < 0.001) was reduced when mosquitoes were fed on blood from treated children compared to untreated controls. The addition of AZ to SPAQ was associated with an increased proportion of infectious feeds (LRT X21 = 5.2, P = 0.02), suggesting a significant effect of AZ on gametocyte infectivity. There was a slight negative effect of SPAQ and SPAQ + AZ on mosquito survival compared to mosquitoes fed with blood from control children (LRTX22 = 330, P < 0.0001). Conclusion This study demonstrates that SMC may contribute to a reduction in human to mosquito transmission of P. falciparum, and the reduced mosquito longevity observed for females fed on treated blood may increase the benefit of this intervention in control of malaria. The addition of AZ to SPAQ in SMC appeared to enhance the infectivity of gametocytes providing further evidence that this combination is not an appropriate intervention.

Evaluation of two Plasmodium vivax sexual-stage antigens as transmission-blocking vaccine candidates

Background: Plasmodium vivax transmission-blocking vaccines (TBVs) have received high attention. PVX_098655 (PvPH) and PVX_101120 (PvSOP26) were predicted to be potential TBV antigens based on the studies of their orthologs in Plasmodium berghei. Methods: Fragments of PvPH (amino acids 22–304) and PvSOP26 (amino acids 30–272) were expressed in the yeast expression system. The recombinant proteins were used to immunize mice to obtain the antisera. The transmission-reducing activities of these antisera were evaluated using the standard membrane feeding assay (SMFA) using Anopheles dirus mosquitoes and P. vivax clinical isolates. Results: The recombinant proteins PvPH and PvSOP26 induced robust antibody responses in mice. With SMFA, the anti-PvSOP26 sera significantly reduced oocyst densities by 92.0% and 84.1% in two parasite isolates, while the anti-PvPH sera did not show evident transmission-reducing activity. Both PvPH and PvSOP26 showed limited gene polymorphisms in the clinical P. vivax isolates. Conclusion: PvSOP26 could be a promising TBV candidate for P. vivax.

Experimental Transmission of Plasmodium malariae to Anopheles gambiae

Our current knowledge of the clinical burden, biology, and transmission of Plasmodium malariae is extremely scarce. To start addressing some of those questions, we experimentally infected Anopheles gambiae mosquitoes with fresh P. malariae isolates obtained from asymptomatic individuals in Lambaréné, Gabon. The proportion of mosquitoes infected via direct membrane feeding assay with either P. malariae monoinfections (16% [19 of 121]) or coinfections (28% [31 of 112]) was higher after serum replacement than in parallel groups without serum replacement (4% [4 of 102] and 4% [2 of 45], respectively; P &lt; .01). Our results show that isolates from asymptomatic carriers can be used for experimental studies of P. malariae transmission.

Experimental Study: The Relationship between Plasmodium falciparum Gametocyte Carriage and Mosquitoes Infectiousness in Two Sympatric Ethnic Groups in Burkina Faso

Aims: The lower susceptibility of the Fulani to malaria compared to Mossi was previously described in Burkina Faso in West Africa. The mature gametocyte stage of Plasmodium falciparum is known to be the only stage capable of infecting the mosquito though this process is disrupted by the action of immunity and other factors as well. Our study aims to assess the ability of two sympatric ethnic groups known to have different susceptibility to Plasmodium falciparum malaria, to infect mosquitoes through an experimental membrane feeding assay. Methodology: Study participants were gametocyte carriers aged from 2 to 12 years recruited in the village of Barkoundouba where Fulani and Mossi are living in sympatric. A venous blood was obtained from each participant for direct membrane feeding assay of insectary reared mosquitoes. Blood fed mosquitoes were stored for 7 days with sugar water as the only food source, then dissected for the microscopic detection for oocysts. Results: A total of 1050 mosquitoes were used for the experimental infections. Eight day after feeding, a total of 897 mosquitoes were dissected, 275 from the Fulani and 622 from the Mossi group. With an average of 43 stomachs examined by experimentation, the mosquito infestation rate was 10.5% in Fulani and 13.2% in Mossi group (p=0.569). The fed mosquito rate was 95 % and 95.6% in Fulani and Mossi ethnic group respectively (p=0.241). The rate of survival mosquitoes after the feeding was 96.5% and 87.5% in Fulani and Mossi ethnic group respectively (p=0.088). The proportion of dissected mosquitoes was 100% and 99.2% in Fulani and Mossi ethnic group respectively (p=0.138) leading to an average oocystic load of 249 in Fulani and 21 in Mossi group. The success rate of DMFA in both groups combined was 57.14%. Indeed, this rate was 33.33% and 66.67% in Fulani and Mossi group respectively. Conclusion: Our study showed that there is no significant difference found between the two ethnic group with the fed, survival, dissected and the infested mosquitoes rate. However, the average of oocystic load was higher in Fulani than the Mossi group despite the low infection in Fulani group. There is a need to explore the mechanism underlying such difference between the two ethnic groups.