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Whether malaria parasites can manipulate mosquito host choice in ways that enhance parasite transmission toward suitable hosts and/or reduce mosquito attraction to unsuitable hosts (i.e. specific manipulation) is unknown. To address this question, we experimentally infected three species of mosquito vectors with wild isolates of the human malaria parasite Plasmodium falciparum, and examined the effects of immature and mature infections on mosquito behavioural responses to combinations of calf odour, human odour and outdoor air using a dual-port olfactometer. Regardless of parasite developmental stage and mosquito species, P. falciparum infection did not alter mosquito activation rate or their choice for human odours. The overall expression pattern of host choice of all three mosquito species was consistent with a high degree of anthropophily, with infected and uninfected individuals showing higher attraction toward human odour over calf odour, human odour over outdoor air, and outdoor air over calf odour. Our results suggest that, in this system, the parasite may not be able to manipulate the early long-range behavioural steps involved in the mosquito host-feeding process. Future studies are required to test whether malaria parasites can modify their mosquito host choice at a shorter range to enhance transmission.

Plasmodium falciparum infections in malaria endemic areas often harbor multiple clones of parasites. However, the transmission success of the different genotypes within the mosquito vector has remained elusive so far. The genetic diversity of malaria parasites was measured by using microsatellite markers in gametocyte isolates from 125 asymptomatic carriers. For a subset of 49 carriers, the dynamics of co-infecting genotypes was followed until their development within salivary glands. Also, individual oocysts from midguts infected with blood from 9 donors were genotyped to assess mating patterns. Multiplicity of infection (MOI) was high both in gametocyte isolates and sporozoite populations, reaching up to 10 genotypes. Gametocyte isolates with multiple genotypes gave rise to lower infection prevalence and intensity. Fluctuations of genotype number occurred during the development within the mosquito and sub-patent genotypes, not detected in gametocyte isolates, were identified in the vector salivary glands. The inbreeding coefficient Fis was positively correlated to the oocyst loads, suggesting that P. falciparum parasites use different reproductive strategies according to the genotypes present in the gametocyte isolate. The number of parasite clones within an infection affects the transmission success and the mosquito has an important role in maintaining P. falciparum genetic diversity. Our results emphasize the crucial importance of discriminating between the different genotypes within an infection when studying the A. gambiae natural resistance to P. falciparum, and the need to monitor parasite diversity in areas where malaria control interventions are implemented.

Pathogens, which have recently colonized a new host species or new populations of the same host, are interesting models for understanding how populations may evolve in response to novel environments. During its colonization of South America from Africa, Plasmodium falciparum, the main agent of malaria, has been exposed to new conditions in distinctive new human populations (Amerindian and populations of mixed origins) that likely exerted new selective pressures on the parasite's genome. Among the genes that might have experienced strong selective pressures in response to these environmental changes, the eba genes (erythrocyte-binding antigens genes), which are involved in the invasion of the human red blood cells, constitute good candidates. In this study, we analysed, in South America, the polymorphism of three eba genes (eba-140, eba-175, eba-181) and compared it to the polymorphism observed in African populations. The aim was to determine whether these genes faced selective pressures in South America distinct from what they experienced in Africa. Patterns of genetic variability of these genes were compared to the patterns observed at two housekeeping genes (adsl and serca) and 272 SNPs to separate adaptive effects from demographic effects. We show that, conversely to Africa, eba-140 seemed to be under stronger diversifying selection in South America than eba-175. In contrast, eba-181 did not show any sign of departure from neutrality. These changes in the patterns of selection on the eba genes could be the consequence of changes in the host immune response, the host receptor polymorphisms and/or the ability of the parasite to silence or express differentially its invasion proteins., (© 2014 John Wiley & Sons Ltd.)

During the last four years, knowledge about the diversity of Plasmodium species in African great apes has considerably increased. Several new species were described in chimpanzees and gorillas, and some species that were previously considered as strictly of human interest were found to be infecting African apes. The description in gorillas of P. praefalciparum, the closest relative of P. falciparum which is the main malignant agent of human malaria, definitively changed the way we understand the evolution and origin of P. falciparum. This parasite is now considered to have appeared recently, following a cross-species transfer from gorillas to humans. However, the Plasmodium vector mosquito species that have served as bridge between these two host species remain unknown. In order to identify the vectors that ensure ape Plasmodium transmission and evaluate the risk of transfer of these parasites to humans, we carried out a field study in Gabon to capture Anopheles in areas where wild and semi-wild ape populations live. We collected 1070 Anopheles females belonging to 15 species, among which An. carnevalei, An. moucheti and An. marshallii were the most common species. Using mtDNA-based PCR tools, we discovered that An. moucheti, a major human malaria vector in Central Africa, could also ensure the natural transmission of P. praefalciparum among great apes. We also showed that, together with An. vinckei, An. moucheti was infected with P. vivax-like parasites. An. moucheti constitutes, therefore, a major candidate for the transfer of Plasmodium parasites from apes to humans.

The origin of Plasmodium falciparum in South America is controversial. Some studies suggest a recent introduction during the European colonizations and the transatlantic slave trade. Other evidence--archeological and genetic--suggests a much older origin. We collected and analyzed P. falciparum isolates from different regions of the world, encompassing the distribution range of the parasite, including populations from sub-Saharan Africa, the Middle East, Southeast Asia, and South America. Analyses of microsatellite and SNP polymorphisms show that the populations of P. falciparum in South America are subdivided in two main genetic clusters (northern and southern). Phylogenetic analyses, as well as Approximate Bayesian Computation methods suggest independent introductions of the two clusters from African sources. Our estimates of divergence time between the South American populations and their likely sources favor a likely introduction from Africa during the transatlantic slave trade.

Both Plasmodium falciparum and Anopheles gambiae show great diversity in Africa, in their own genetic makeup and population dynamics. The genetics of the individual mosquito and parasite are known to play a role in determining the outcome of infection in the vector, but whether differences in infection phenotype vary between populations remains to be investigated. Here we established two A. gambiae s.s. M molecular form colonies from Cameroon and Burkina Faso, representing a local and a foreign population for each of the geographical sites. Experimental infections of both colonies were conducted in Cameroon and Burkina Faso using local wild P. falciparum, giving a sympatric and allopatric vector-parasite combination in each site. Infection phenotype was determined in terms of oocyst prevalence and intensity for at least nine infections for each vector-parasite combination. Sympatric infections were found to produce 25% fewer oocysts per midgut than allopatric infections, while prevalence was not affected by local/foreign interactions. The reduction in oocyst numbers in sympatric couples may be the result of evolutionary processes where the mosquito populations have locally adapted to their parasite populations. Future research on vector-parasite interactions must take into account the geographic scale of adaptation revealed here by conducting experiments in natural sympatric populations to give epidemiologically meaningful results.

Malaria remains a devastating disease despite efforts at control and prevention. Extensive studies using mostly rodent infection models reveal that successful Plasmodium parasite transmission by the African mosquito vector Anopheles gambiae depends on finely tuned vector-parasite interactions. Here we investigate the transcriptional response of A. gambiae to geographically related Plasmodium falciparum populations at various infection intensities and different infection stages. These responses are compared with those of mosquitoes infected with the rodent parasite Plasmodium berghei. We demonstrate that mosquito responses are largely dependent on the intensity of infection. A major transcriptional suppression of genes involved in the regulation of midgut homeostasis is detected in low-intensity P. falciparum infections, the most common type of infection in Africa. Importantly, genes transcriptionally induced during these infections tend to be phylogenetically unique to A. gambiae. These data suggest that coadaptation between vectors and parasites may act to minimize the impact of infection on mosquito fitness by selectively suppressing specific functional classes of genes. RNA interference (RNAi)-mediated gene silencing provides initial evidence for important roles of the mosquito G protein-coupled receptors (GPCRs) in controlling infection intensity-dependent antiparasitic responses.

Many genes involved in the immune response of Anopheles gambiae, the main malaria vector in Africa, have been identified, but whether naturally occurring polymorphisms in these genes underlie variation in resistance to the human malaria parasite, Plasmodium falciparum, is currently unknown. Here we carried out a candidate gene association study to identify single nucleotide polymorphisms (SNPs) associated with natural resistance to P. falciparum. A. gambiae M form mosquitoes from Cameroon were experimentally challenged with three local wild P. falciparum isolates. Statistical associations were assessed between 157 SNPs selected from a set of 67 A. gambiae immune-related genes and the level of infection. Isolate-specific associations were accounted for by including the effect of the isolate in the analysis. Five SNPs were significantly associated to the infection phenotype, located within or upstream of AgMDL1, CEC1, Sp PPO activate, Sp SNAKElike, and TOLL6. Low overall and local linkage disequilibrium indicated high specificity in the loci found. Association between infection phenotype and two SNPs was isolate-specific, providing the first evidence of vector genotype by parasite isolate interactions at the molecular level. Four SNPs were associated to either oocyst presence or load, indicating that the genetic basis of infection prevalence and intensity may differ. The validity of the approach was verified by confirming the functional role of Sp SNAKElike in gene silencing assays. These results strongly support the role of genetic variation within or near these five A. gambiae immune genes, in concert with other genes, in natural resistance to P. falciparum. They emphasize the need to distinguish between infection prevalence and intensity and to account for the genetic specificity of vector-parasite interactions in dissecting the genetic basis of Anopheles resistance to human malaria.

Endemic malaria has been eradicated from France, but some falciparum malaria cases have been described in patients who have never travelled outside the country. Ms. V. 21 year-old and Mr. M. 23 year-old living together in Paris were on holiday in Saint Raphaël (French Riviera). They presented with fever, vertigo and nausea. A blood smear made to control thrombocytopaenia revealed intra-erythrocytic forms of Plasmodium falciparum. The parasitaemia level was 0.15% for Ms. V and 3.2% for Mr. M. This couple had no history of blood transfusion or intravenous drug use. They had never travelled outside metropolitan France, but had recently travelled around France: to Saint Mard (close to Paris Charles de Gaulle (CdG) airport), to Barneville plage (in Normandy) and finally to Saint Raphaël. The most probable hypothesis is an infection transmitted in Saint Mard by an imported anopheline mosquito at CdG airport. The DNA analysis of parasites from Ms. V.'s and Mr. M.'s blood revealed identical genotypes. Because it is unlikely that two different anopheline mosquitoes would be infected by exactly the same clones, the two infections must have been caused by the infective bites of the same infected mosquito.

Background: The time necessary for malaria parasite to re-appear in the blood following treatment (re-infection time) is an indirect method for evaluating the immune defences operating against pre-erythrocytic and early erythrocytic malaria stages. Few longitudinal data are available in populations in whom malaria transmission level had also been measured., Methods: One hundred and ten individuals from the village of Ndiop (Senegal), aged between one and 72 years, were cured of malaria by quinine (25 mg/day oral Quinimax in three equal daily doses, for seven days). Thereafter, thick blood films were examined to detect the reappearance of Plasmodium falciparum every week, for 11 weeks after treatment. Malaria transmission was simultaneously measured weekly by night collection of biting mosquitoes., Results: Malaria transmission was on average 15.3 infective bites per person during the 77 days follow up. The median reappearance time for the whole study population was 46.8 days, whereas individuals would have received an average one infective bite every 5 days. At the end of the follow-up, after 77 days, 103 of the 110 individuals (93.6%; CI 95% [89.0-98.2]) had been re-infected with P. falciparum. The median reappearance time ('re-positivation') was longer in subjects with patent parasitaemia at enrolment than in parasitologically-negative individuals (58 days vs. 45.9; p = 0.03) and in adults > 30 years than in younger subjects (58.6 days vs. 42.7; p = 0.0002). In a multivariate Cox PH model controlling for the sickle cell trait, G6PD deficiency and the type of habitat, the presence of parasitaemia at enrolment and age >/= 30 years were independently predictive of a reduced risk of re-infection (PH = 0.5 [95% CI: 0.3-0.9] and 0.4; [95% CI: 0.2-0.6] respectively)., Conclusion: Results indicate the existence of a substantial resistance to sporozoites inoculations, but which was ultimately overcome in almost every individual after 2 1/2 months of natural challenges. Such a study design and the results obtained suggest that, despite a small sample size, this approach can contribute to assess the impact of intervention methods, such as the efficacy vector-control measures or of malaria pre-erythrocytic stages vaccines.

Background: The adaptation of Anopheles gambiae to humans and its environment involves an ongoing speciation process that can be best demonstrated by the existence of various chromosomal forms adapted to different environments and of two molecular forms known as incipient taxonomic units., Methods: The aim of this study was to compare the epidemiologic role of Anopheles arabiens is and the molecular forms M and S of Anopheles gambiae in the transmission of Plasmodium in a rural areas of southern Senegal, Dielmo. The sampling of mosquitoes was carried out monthly between July and December 2004, during the rainy season, by human volunteers and pyrethrum spray catches., Results: Anopheles arabiensis, An. gambiae M and S forms coexisted during the rainy season with a predominance of the M form in September and the peak of density being observed in August for the S form. Similar parity rates were observed in An. arabiensis [70.9%] (n = 86), An. gambiae M form [68.7%] (n = 64) and An. gambiae S form [81.1%] (n = 156). The circumsporozoite protein (CSP) rates were 2.82% (n = 177), 3.17% (n = 315) and 3.45% (n = 405), with the mean anthropophilic rates being 71.4% (n = 14), 86.3% (n = 22) and 91.6% (n = 24) respectively for An. arabiensis and An. gambiae M and S forms. No significant difference was observed either in host preference or in Plasmodium falciparum infection rates between sympatric M and S populations., Conclusion: No difference was observed either in host preference or in Plasmodium falciparum infection rates between sympatric M and S populations, but they present different dynamics of population. These variations are probably attributable to different breeding conditions.

In much of sub-Saharan Africa, the mosquito Anopheles gambiae is the main vector of the major human malaria parasite, Plasmodium falciparum. Convenient laboratory studies have identified mosquito genes that affect positively or negatively the developmental cycle of the model rodent parasite, P. berghei. Here, we use transcription profiling and reverse genetics to explore whether five disparate mosquito gene regulators of P. berghei development are also pertinent to A. gambiae/P. falciparum interactions in semi-natural conditions, using field isolates of this parasite and geographically related mosquitoes. We detected broadly similar albeit not identical transcriptional responses of these genes to the two parasite species. Gene silencing established that two genes affect similarly both parasites: infections are hindered by the intracellular local activator of actin cytoskeleton dynamics, WASP, but promoted by the hemolymph lipid transporter, ApoII/I. Since P. berghei is not a natural parasite of A. gambiae, these data suggest that the effects of these genes have not been drastically altered by constant interaction and co-evolution of A. gambiae and P. falciparum; this conclusion allowed us to investigate further the mode of action of these two genes in the laboratory model system using a suite of genetic tools and infection assays. We showed that both genes act at the level of midgut invasion during the parasite's developmental transition from ookinete to oocyst. ApoII/I also affects the early stages of oocyst development. These are the first mosquito genes whose significant effects on P. falciparum field isolates have been established by direct experimentation. Importantly, they validate for semi-field human malaria transmission the concept of parasite antagonists and agonists.

We investigated patterns of genetic diversity of Plasmodium falciparum associated with its two main African vectors: Anopheles gambiae and Anopheles funestus. We dissected 10,296 wild-caught mosquitoes from three tropical sites, two in Cameroon (Simbock and Tibati, separated by 320 km) and one in Kenya (Rota, >2,000 km from the other two sites). We assayed seven microsatellite loci in 746 oocysts from 183 infected mosquito guts. Genetic polymorphism was very high in parasites isolated from both vector species. The expected heterozygosity (H(E)) was 0.79 in both species; the observed heterozygosities (H(O)) were 0.32 in A. funestus and 0.42 in A. gambiae, indicating considerable inbreeding within both vector species. Mean selfing (s) between genetically identical gametes was s = 0.33. Differences in the rate of inbreeding were statistically insignificant among sites and between the two vector species. As expected, because of the high rate of inbreeding, linkage disequilibrium was very high; it was significant for all 21 loci pairs in A. gambiae and for 15 of 21 pairs in A. funestus, although only two pairwise comparisons were between loci on the same chromosome. Overall, the genetic population structure of P. falciparum, as evaluated by F statistics, was predominantly clonal rather than panmictic, a population structure that facilitates the spread of antimalarial drug and vaccine resistance and thus may impair the effectiveness of malaria control efforts.

Parasites that cause malaria must complete a complex life cycle in Anopheles vector mosquitoes in order to be transmitted from human to human. Previous gene-silencing studies have shown the influence of mosquito immunity in controlling the development of Plasmodium. Thus, parasite survival to the oocyst stage increased when the parasite antagonist gene LRIM1 (leucine-rich repeat immune protein 1) of the mosquito was silenced, but decreased when the C-type lectin agonist gene CTL4 or CTLMA2 (CTL mannose binding 2) was silenced. However, such effects were shown for infections of the human mosquito vector Anopheles gambiae with the rodent parasite Plasmodium berghei. Here, we report the first results of A. gambiae gene silencing on infection by sympatric field isolates of the principal human pathogen P. falciparum. In contrast with the results obtained with the rodent parasite, silencing of the same three genes had no effect on human parasite development. These results highlight the importance of following up discoveries in laboratory model systems with studies on natural parasite-mosquito interactions.

Serpins are central to the modulation of various innate immune responses in insects and are suspected to influence the outcome of malaria parasite infection in mosquito vectors. Three Anopheles gambiae serpins (SRPN1, -2, and -3) were tested for their ability to inhibit the prophenoloxidase cascade, a key regulatory process in the melanization response. Recombinant SRPN1 and -2 can bind and inhibit a heterologous phenoloxidase-activating protease and inhibit phenoloxidase activation in vitro. Using a reverse genetics approach, we studied the effect of SRPN2 on melanization in An. gambiae adult females in vivo. Depletion of SRPN2 from the mosquito hemolymph increases melanin deposition on foreign surfaces such as negatively charged Sephadex beads. As reported, the knockdown of SRPN2 adversely affects the ability of the rodent malaria parasite Plasmodium berghei to invade the midgut epithelium and develop into oocysts. Importantly, we tested whether the absence of SRPN2 from the hemolymph influences Plasmodium falciparum development. RNAi silencing of SRPN2 in an An. gambiae strain originally established from local populations in Yaoundé, Cameroon, did not influence the development of autochthonous field isolates of P. falciparum. This study suggests immune evasion strategies of the human malaria parasite and emphasizes the need to study mosquito innate immune responses toward the pathogens they transmit in natural vector-parasite combinations.

Plasmodium falciparum expresses many antigens, which elicit various immune responses in exposed individuals, but no simple surrogate marker for protection has yet been developed. In this prospective survey, we looked for immune responses predictive of protection at various stages of progression from parasite inoculation to onset of disease. We studied 110 Senegalese volunteers from an area in which malaria is mesoendemic after they had received eradication therapy. We evaluated 4 protection-related outcomes (reappearance of parasitemia, duration of asymptomatic carriage, time to first clinical episode, and incidence of clinical episodes) in terms of levels of immunoglobulin G (IgG) against 3 crude parasite extracts and 5 conserved antigens during a 5-month period. Kaplan-Meier estimates and age-adjusted regression models showed these 4 outcomes to be associated with different patterns of IgG response to PfEMP3-cl5 (derived from P. falciparum erythrocyte membrane protein 3), PfEB200, MSP-1(19) (derived from merozoite surface protein-1), [NANP]10, infected red blood cell membrane, and merozoite and schizont extracts. It should, therefore, be possible to develop surrogate markers for each end point on the basis of IgG response to a limited number of conserved antigens.

Women may be more susceptible to malaria during late pregnancy and in the early postpartum period. Susceptibility was investigated in 38 pregnant women followed from preconception to one year after delivery in areas of high disease prevalence. The incidence of malaria was 2-3 times as high during the second and third trimester, and 4 times as high in the first two months postpartum, compared to preconception and the first trimester.

Background: The prevalence of Plasmodium falciparum and Plasmodium vivax malaria was very high in Corsica just before the Second World War. The last outbreak was in 1972 and the most recent indigenous case was in 2006. Results: Analysis of historical data shows that anopheline vectors were abundant. Recent surveys demonstrated that potential vectors are still present in Corsica, despite the likely disappearance of Anopheles sacharovi. Moreover, P. falciparum can develop experimentally into these mosquitoes, notably Anopheles labranchiae, which is locally abundant, and parasites are regularly introduced into the island. Discussion, Conclusions: The presence of vectors, the introduction of parasites and the conducive climate raise questions about the possibility of malaria re-emerging and becoming re-established in Corsica. Analysis of historic and current parasitological and entomological data shows that the current theoretical risk of indigenous cases or malaria foci is negligible, particularly since there is very little contact between humans and Anopheles mosquitoes, Plasmodium carriers are reliably treated and there is a widespread vector control on the island. [ABSTRACT FROM AUTHOR]

Summary: Entomological surveys were carried out in the town of Mbalmayo and in the nearby rural village of Olama, within the equatorial forest zone of Cameroon. Mosquitoes were captured when landing on human volunteers and by pyrethrum spray catches. Malaria vectors captured were Anopheles gambiae Giles (M and S forms) and A. moucheti Evans in both areas, together with A. funestus Giles in Mbalmayo. One A. marshallii (Theobald) specimen infected by Plasmodium falciparum was found in Olama. Anopheles moucheti was the most abundant anopheline species caught in Olama, while A. gambiae was the most abundant in Mbalmayo. All these vectors were highly anthropophilic as indicated by the fact that only 5 of 201 blood meals analysed had been taken from non-human hosts. Plasmodium falciparum was the only malaria parasite species found in Mbalmayo, while P. malariae was also found in Olama. The annual entomological inoculation rate was estimated at 129 infective bites/person/year in Mbalmayo and 322 in Olama. Comparison with data published in 1955 from Mbalmayo, before expansion of the town, showed the impact of urbanization on the composition of the vector system and malaria transmission dynamics. Such changes should be considered when implementing sustainable control measures. [Copyright &y& Elsevier]

An outbreak of Plasmodium falciparum malaria occurred in Tinzaouatine in southern Algeria in 2007. The likely vector, Anopheles gambiae mosquitoes, had not been detected in Algeria. Genes for resistance to chloroquine were detected in the parasite. The outbreak shows the potential for an increase in malaria vectors in Algeria. [ABSTRACT FROM AUTHOR]

We report 2 cases of Plasmodium falciparum malaria in southern France in a French woman and an American man of Togolese origin who reported no recent travel to malaria-endemic countries. Both infections occurred after a stay near Marseille, which raises the possibility of autochthonous transmission. Entomologic and genotypic investigations are described. [ABSTRACT FROM AUTHOR]