Plasmodium falciparum is associated with perturbations in B cell sub-set homeostasis, including expansion atypical memory B cells. However, B cell perturbations immediately following acute malaria infection have been poorly characterized, especially with regard to their relationship with immunity to malaria.Repeated exposure to To better understand the kinetics of B cell sub-sets following malaria, the proportions of six B cell sub-sets were assessed at five time points following acute malaria in four to 5 years old children living in a high transmission region of Uganda. B cell sub-set kinetics were compared with measures of clinical immunity to malaria—lower parasite density at the time of malaria diagnosis and recent asymptomatic parasitaemia.Atypical memory B cell and transitional B cell proportions increased following malaria. In contrast, plasmablast proportions were highest at the time of malaria diagnosis and rapidly declined following treatment. Increased proportions of atypical memory B cells were associated with greater immunity to malaria, whereas increased proportions of transitional B cells were associated with evidence of less immunity to malaria.These findings highlight the dynamic changes in multiple B cell sub-sets following acute, uncomplicated malaria, and how these sub-sets are associated with developing immunity to malaria. Plasmodium falciparum continues to cause over a half million deaths each year, with children being disproportionately affected [Malaria caused by affected . Childreaffected , 3. One affected –6. Antibaffected –10.Plasmodium infection generally develop only after years of repeated exposure, likely due to immune immaturity of the host and antigenic variation of parasites [Plasmodium infection may interfere with B cell development and maintenance of memory responses [Plasmodium alters the distribution of these B cell sub-sets, and has been associated with an expansion of ‘atypical’ MBCs in individuals living in malaria-endemic areas [P. falciparum exposure [Effective B cell and antibody responses to arasites –12. Anotesponses –17. Afteesponses ; these cic areas –15, 19. ic areas , 20, 21.ic areas , 21. On exposure , 22–24. Plasmodium chabaudi have found that newly differentiated plasmablasts only circulate in the blood for a short time following primary or secondary infection while other sub-sets such as transitional, naïve B cells and MBCs fluctuate greatly but remain readily detectable in the peripheral blood [B cell sub-sets generated during malaria episodes may indicate which B cells are associated with developing immunity. Various studies have described multiple B cell sub-sets in people exposed to varying levels of malaria , 25, 26,P. falciparum parasites by thick blood smear. Febrile participants that had any detectable parasites by thick blood smear were diagnosed with symptomatic malaria and treated with an artemisinin-based combination therapy. Participants also had monthly blood smears to assess parasitaemia regardless of symptoms. For this study, 38 consecutive children were enrolled that presented with symptomatic malaria, with parasite densities greater than 2000 parasites/µl. Asymptomatic parasitaemia was defined by parasitaemia in afebrile participants occurring at least 14 days after and five days before a malaria episode.Samples were obtained from participants between 4.6 and 5.0 years of age enrolled in the Tororo Child Cohort (TCC) study in Tororo, Uganda, an area of intense malaria transmission  . Cohort All study participants or their parents or guardians provided individual written informed consent. Approval was granted by the Uganda National Council of Science and Technology, the Makerere University Research and Ethics Committee, the University of California, San Francisco Committee on Human Research,A half ml of whole blood was collected through finger prick or phlebotomy. Whole blood was washed with PBS and then labelled for 30 min with antibodies against CD3 (clone UCHT1), CD14 (clone M5E2), CD19 (clone HIB19), CD10 (clone HI10a), CD38 (clone HIT2), CD27 (clone O323), IgG (clone G18-145) , and IgD (clone IA6-2) (BD Biosciences)  , and on days 3, 7, 14, and 28 following treatment. Flow cytometry was used to measure the relative proportions of: transitional B cells (CD19+CD10+), naïve B cells (CD19+CD10−IgD+CD27−), innate-like MBCs (CD19+CD10−IgD+CD27+), classical MBCs (CD19+CD10−IgD−CD27+), atypical MBCs (CD19+CD10−IgD−CD27−), and plasmablasts/plasma cells  cells and naïve B cells decreased following malaria. Mean PC proportion was highest at 6.1 % on day 0 and then rapidly declined to 3.3 % by day 7 (p = 0.002). Participants with a second malaria episode during the 28-day study had increased PC proportions following second episode diagnosis  at the time of malaria, participants with lower parasite density had higher proportions of atypical MBCs during the 28 days following malaria , but were not significantly associated with measures of immunity to malaria. There were no significant associations between measures of immunity to malaria and the proportions of classical MBCs, innate-like MBCs or naïve B cells.P. falciparum infection.This study investigated the kinetics of different B cell sub-sets at the time of and following symptomatic malaria and then evaluated whether proportions of these sub-sets were associated with measures of immunity to malaria. The proportions of atypical MBCs and transitional B cells both increased during the 28 days following symptomatic malaria, whereas naïve B cell proportions declined slowly and circulating PCs proportions declined rapidly following treatment of malaria, increasing again in participants who had a second episode of malaria. Notably, children with evidence of greater immunity to malaria had higher proportions of atypical MBCs and lower proportions of transitional B cells following malaria, and these associations remained stable over the 28 days following malaria. These findings suggest that children with higher proportions of these cells during and following malaria have improved immune responses to Plasmodium exposure [P. falciparum exposure, as results were unchanged in a multivariate analysis including estimates of exposure. One possibility is that the ability to sustain asymptomatic infections, which tend to be of longer duration than treated symptomatic malaria infections, leads to higher proportions of atypical MBCs due to chronic antigen exposure. Alternatively, it may be that higher parasite densities seen in less immune individuals drive down the proportion of atypical MBCs, possibly as a result of apoptosis or homing of atypical MBCs to tissues. It is also possible that atypical MBCs contribute, in an as-yet undefined manner, to anti-malarial immunity. Further studies are necessary to determine if atypical MBCs are causally associated with immunity to malaria.People living in malaria-endemic regions have increased proportions of atypical MBCs, with these proportions increasing with age and with cumulative exposure , 13, 14.exposure , 21. Desexposure , other sexposure , 21. Witexposure , 24. ThiP. falciparum induced, non-specific, polyclonal B cell activation [Increases in transitional B cell proportions following malaria have been previously noted, and this expansion was hypothesized to be due to disruption of B cell homeostasis . This sttivation  and apoptivation  in indivtivation , similartivation . Transittivation , 43, 44.P. chabaudi [Outside of vaccine trials, plasmablasts/plasma cells can be difficult to characterize due to their paucity in the peripheral blood and highly synchronous migration to the bone marrow. Development and migration of PCs has been characterized in experimental infection of mice with chabaudi , 45, 46,P. falciparum, an important consideration when evaluating immunity.Some unique strengths of this study are the detailed longitudinal follow up of participants, allowing characterization of relevant clinical outcomes and the measurement of numerous B cell sub-type proportions at multiple timepoints during and following an episode of symptomatic malaria. By evaluating proportions of these sub-types at the time of malaria and following treatment, the potential confounding effects of the duration of time since malaria, which can affect the proportions of different cell types, was limited. Similarly, by evaluating children in a very narrow age range, the potential confounding effect of age, which is associated with immunity to malaria and proportions of B cell sub-types, was reduced , 13. TheP. chabaudi that memory B cell responses circulating in the blood reflected the general composition of B cells in peripheral tissues [A limitation of this study is the inability to determine whether associations between clinical phenotypes and proportions of B cell sub-sets represent a direct effect of these cell types. In addition, evaluation was limited to B cell sub-sets trafficking through peripheral blood which, while clearly an important compartment for interaction with a blood stage pathogen, may not represent proportions in other important compartments such as secondary lymphoid organs or peripheral tissues. There is evidence using experimental mouse infection with  tissues . Since p tissues , 22, 47,P. falciparum greatly affects B cell homeostasis, but further research is necessary to understand the full implications of these changes. Since most vaccines are designed to induce a protective B cell response that can be sustained over time, it is vital to understand consequences of P. falciparum infection on the humoral immune system, memory formation and maintenance of memory.The findings of this study highlight the importance of investigating the dynamics of multiple B cell sub-sets, including consideration of the timing of these measurements with respect to malaria, in order to understand the contribution of the humoral immune system in developing immunity to malaria. Infection by


Anaemia is an increasingly recognized health problem in Africa, particularly in infants and pregnant women. Although malaria is known to be the main risk factor of anaemia in both groups, the consequences of maternal factors, particularly malaria in pregnancy (MiP), on infant haemoglobin (Hb) concentrations during the first months of life are still unclear.st clinical antenatal visit (ANC), 2nd ANC and delivery were used to assess malaria during pregnancy. Infant Hb concentrations were measured at birth, 6, 9 and 12 months of age. A mixed multi-level model was used to assess the association between MiP and infant Hb variations during the first 12 months of life.We followed-up a cohort of 1005 Beninese pregnant women from the beginning of pregnancy until delivery. A subsample composed of the first 400 offspring of these women were selected at birth and followed until the first year of life. Placental histology and blood smear at 1Placental malaria  and maternal peripheral parasitaemia at delivery  were the main maternal factors significantly associated with infant Hb concentrations during the first year of life. Poor maternal nutritional status and malaria infection during infancy were also significantly associated with a decrease in infant Hb.Antimalarial control and nutritional interventions before and during pregnancy should be reinforced to reduce specifically the incidence of infant anaemia, particularly in Sub-Saharan countries. The World Health Organization (WHO) has estimated that over 50% of children less than 4 years are anaemic in developing countries . Iron deIn Africa, very few studies have focused on the effects of maternal factors during pregnancy on postnatal child’s health. Although the risk factors of anaemia in childhood are fairly well known –7, the iOn the occasion of a multi-center trial of Intermittent Preventive Treatment in Pregnancy (IPTp) comparing sulfadoxine-pyrimethamine and mefloquine  , we had st ANC) to delivery in the framework of the MiPPAD clinical trial. The first 400 infants to be delivered from these women, were enrolled from January 2010 until June 2011 and followed throughout the first year of life.Prospective cohort study, set-up between January 2010 and May 2012 to follow pregnant women from the first antenatal clinical visit .The study was conducted in two maternity clinics (Attogon and Sékou) in the district of Allada, a semi-rural area located 50 km north of Cotonou and the economic capital of Benin. Allada district is characterized by a subtropical climate and malaria is hyperendemic with an average of 20.5 infected anopheles bites/person/year . Plasmodst ANC to be taken at home (100 mg twice day for 3 days) according to the guidelines of the Beninese Ministry of Health. In addition, supplements of oral ferrous sulfate (200 mg/day) and folic acid (5 mg/day) were given to the pregnant women to be taken at home. Pregnant women with a Hb concentration below 110g/L were treated according to the severity of anaemia  and referred to the district tertiary hospital in case of severe anaemia (Hb < 70 g/L)). Mild or moderate infant anaemia was treated with 10 mg/kg/day of iron syrup for two months. Children with severe anaemia were also referred to the district tertiary hospital for blood transfusion. Infant with positive stool test were treated with mebendazole syrup (100 mg twice day for 3 days). All the medications prescribed to the women and the children during their participation in the study were free of charge.The study population was composed of HIV—negative pregnant women and their children residing in the district of Allada. Twin pregnancies, pregnancies complicated by stillbirth or fetal abnormalities have been excluded. IPTp was administered according to the clinical trial protocol and the details are presented elsewhere . Brieflyst ANC, women were examined and gestational age, middle upper arm circumference (MUAC), weight and height were recorded. This information, except for height, was also collected at 2nd ANC and delivery. Gestational age was determined from fundal height measurement by bimanual palpation and following McDonald's rules , and eij is the residual variation [eij—N]. Fixed effects parameters were estimated using the maximum likelihood method, and variance components were estimated using the restricted maximum likelihood method. To take into account the numerous physiological changes and the non linear evolution of Hb during the first 3 months of life, we excluded the first Hb measurement (at birth) from the hierarchical mixed model. Also, we adjusted our longitudinal analysis on the hematological parameters of the newborn such as iron or folic acid deficiencies at birth that could modify the levels of Hb during the first year of life. All variables with P values below 0.2 in univariate analysis and which were not in the causal pathway between placental malaria and infant Hb, were included in multivariate analyses. Manual backward selection procedure was performed and statistical significance was set at P < 0.05. For variables with more than two categories, P value of the global test is given.Then, we assessed the effect of MiP on infant Hb at birth using multivariable linear regression. Afterwards, we used a longitudinal approach to take into account Hb variations over time and to estimate the impact of MiP on infant Hb throughout the first year of life. Assuming that successive Hb measurements in the same infant were correlated, the data presented a hierarchical two-level structure, where Hb measurements (level 1) were clustered within infants (level 2) . TherefoThis study was approved by the Ethics Committee of the Health Sciences Faculty of Cotonou in Benin. Before each inclusion, all participants involved in our study provided their written informed consent to participate in this study. The study was also explained in the local language to the participant, and her voluntary consent was obtained. In case the woman could not read, an impartial witness was involved in the process. Mothers were free to interrupt their participation at any time in the study.Between January 2010 and June 2011, 400 mother-infant pairs were enrolled in the study. The proportion of lost to follow-up during the whole study period was 1.2% . Hb was st ANC to delivery (72.5% to 50%) and 36.9% (145/392) of women were anaemic during all scheduled visits of pregnancy. Helminth infestations decreased progressively from the 1st ANC (9.1%) to delivery (5.1%). The proportion of women with a positive blood smear decreased after IPTp administration (from 16% at 1st ANC to 4.1% at 2nd ANC), and increased again at delivery (10.8%). Eighty seven pregnant women (22.1%) had a placental malaria infection at histology evaluation. Infant families’ socio-economic level was low for 49.3% of the study population.P < 0.01). The Hb level decreased from birth to 6 months and remained almost constant until 12 months (P < 0.01). Over a third of the children presented with ID after 6 months. Malaria infection was common at all stages of follow-up, around 12%.2 months . Anaemiast ANC, 2nd ANC and delivery were not statistically associated with newborn Hb at birth  = −1.1 g/L, 95% CI , P = 0.30). After adjustment, newborn ferritin level was significantly associated with a decrease in newborns' Hb concentrations while first pregnancy, Aïzo ethnic group, newborn’s folic acid and vitamin B12 concentrations were significantly associated with an increase in Hb concentrations at birth . Consideat birth .The relation between the number of maternal anaemia episodes during pregnancy and newborns’ Hb concentrations was not significant even if a lower Hb concentrations were observed in infants born to mothers with at least two episodes of anaemia during gestation  .Newborns of mothers with more than 3 ANC visits tended to present lower Hb concentrations than those of mothers with less than 3 ANC visits, but the association was not statistically significant .st and 2nd ANC were not associated with infant Hb modification , 9.2% (95% CI = 7.6–10.7%), 7.7% (95% CI = 6.3–9.1%), 29.8% (95% CI = 27.3–32.3%) of infant anaemia in this population were attributable to maternal peripheral parasitaemia at delivery, placental malaria infection, maternal underweight and malaria infection during infancy, respectively.This study has showed an important reduction of Hb concentrations in Beninese infants born to mothers with peripheral parasitaemia at delivery or placental malaria infection compared to infants from uninfected mothers. Our study also found a high prevalence of anaemia throughout infancy in the study population.To our knowledge, this is one of the few studies to evaluate the effect of MiP on infant Hb variations through a one-year follow-up of the newborns in Sub-Saharan Africa (SSA). As Hb varies physiologically during the first year of life, an important strength of our study was to use a multilevel linear regression, best adapted to repeated measures to take into account the effect of time on infants’ Hb levels. Our analyses suggested that 65% of the total variance could be explained by the mixed model. Therefore, this model seems to be well suitable to the analyses of data. We also recorded few missing data (1.2%) reflecting the good quality of the follow-up. Furthermore, we were able to take into account potential confounding factors like estimated pre-pregnancy BMI, infants’ malaria infections and nutritional status  at different times during the follow-up period.We found that infant’s anaemia was very prevalent in southern Benin, particularly in the second half of the first year of life (> 65%). These high proportions are consistent with data from the Beninese Departmental Health and Demographic survey . In our st to 2nd ANC probably because of IPTp efficacy in parasite clearing [nd IPTp dose and delivery had frequently exceeded two months. Consequently the pregnant women remained insufficiently protected for the last weeks of pregnancy, after the second IPTp dose. A similar result had previously been reported on Beninese pregnant women, clearly demonstrating that women who had an early intake of last IPTp dose were more at risk of LBW [When we considered the effect of MiP timing, only maternal peripheral parasitaemia at delivery was significantly associated with decreasing of infant Hb. In our study, malaria infection decreased from 1clearing , therefok of LBW .Maternal peripheral parasitaemia at delivery and placental malaria infection were the main maternal factors that contributed to reduce infant Hb concentrations during the first year of life. Placental malaria is one of the major features of malaria during pregnancy and it has been widely used as a standard indicator to characterize malaria infection in epidemiological investigations ,26. Moreet al. showed that first malaria infections in early childhood could be attributed independently to both placental malaria and high levels of exposure to infected anopheles [et al. showed that placental malaria infection was strongly associated with an increased risk of non-malaria fever episodes as well as gastrointestinal and respiratory febrile syndromes [in-utero exposure of the fetus to a variety of malaria antigens that may induce immunological tolerance [The biological explanation of the effect of MiP on infant anaemia in this study is complex to establish. Infant anaemia may be indirectly due to a higher susceptibility to malaria, a well-known risk factor for anaemia in itself. Two potential mechanisms can be evoked. First, environmental factors are likely to play a role, as women infected by malaria at delivery may have been more exposed to anopheles bites during pregnancy than uninfected women. Because mothers infected by malaria parasites and their children live in the same area, the latter may also be more exposed to the risk of malaria, one of the most important causes of infants' anaemia. Secondly, an immune tolerance phenomenon can also be involved, a very likely mechanism, as in the analysis of our dataset the relationship between MiP and decrease of infant's Hb persisted after adjusting on infant malaria infection throughout the first year of life. In a recent study on a similar population in Benin, Le Port nopheles . In the yndromes . It can olerance  and thusolerance . Such imolerance ,32 potenet al. (1994) in Malawi had assumed that placental malaria infection might interfere with the hematologic status of the infant independently of birth weight and prematurity [et al. (1998) had also reported that placental malaria infection was the strongest risk factor for anaemia in six-month-old children in southern Cameroon [Earlier reports from Malawi and Cameroon had previously suggested a relation between placental malaria and infant Hb status. Redd maturity . Cornet Cameroon . These sCameroon .Our study suggests that children born to mothers with a very low BMI at the beginning of pregnancy (< 16 kg/m²) had a lower Hb concentration than children born to mothers with a normal BMI. It is also the first time that this relation is described. The low BMI reflects the maternal under-nutrition and could be a proxy of future infant's nutrition. Indeed, children born to underweight mothers may have a poor diet and therefore may be at higher risk of anaemia. Furthermore, underweight and stunted women are at high risk of delivering premature or LBW infants, who in turn, are at high risk of poor growth and anaemia in childhood and adolescence . Indeed et al., this mechanism also depends on the intensity of maternal iron depletion . In our study, we have only considered the number of anaemia episodes detected at IPTp administrations and this may not necessarily reflect the history of maternal anaemia during all pregnancy duration. Indeed, by studying repeated measures of Hb on a larger sample size (1005 mother-infant pairs) and by using a path analysis, Ouédraogo et al. found a direct, though border-line, association of maternal anaemia in the third trimester and newborn's hemoglobin level at birth .The number of maternal anaemia episodes during pregnancy was not associated with infant Hb concentration at birth and during the first year of life. It may seem surprising but similar results have been reported in Malawi , which mMalaria infection, fever and inflammatory syndrome in children at the time of Hb measurement were significantly associated with a decrease in infant Hb concentrations probably due to hemolytic anaemia. The association of anaemia with fever or inflammatory syndrome underlines the important contribution of infections in the occurrence of infant anaemia during the first months of life and the potential causes of inflammation in pregnancy as well as in infancy hence deserve further studies.This study has some limitations that should be considered. First, the absence of entomological data in our study such as the level of anopheles transmission did not allow the determination of the part of malaria risk attributable to a common exposure of the mother and her child to malaria vectors, even if the results rather suggest an effect of placental parasite sequestration on the immune tolerance of child. Second, the maternal nutritional status was assessed by BMI and weight was calculated approximately on the assumption of a 1kg/month gain from the end of the first trimester, which may not reflect correctly the nutritional status of women at the beginning of pregnancy because of the difference of weight gain from underweight to obese . FurtherAnaemia in infancy is very common in Benin. In the present study, MiP and malaria infection during infancy were independently associated with a substantial reduction in infant Hb during the first year of life. Low maternal BMI at the beginning of pregnancy was also related to a decrease in infants' Hb. Strategies to prevent anaemia in infants should as well integrate measures during pregnancy such as the prevention of MiP and the improvement of maternal nutritional status. IPTp and the use of bed nets by both mothers and infants should be reinforced. Micronutrient supplementation should also be implemented in women of childbearing age and pregnant women but with a lot of caution in a context of malaria-iron interactions where the protection of iron deficiency against malaria is still a controversial issue ,41.S1 FigDuring the first months of life, haemoglobin level declines from very high level at birth to its lowest level at 2–3 months of age. This decrease is known as the “physiologic anaemia of the newborn”. After Hb has reached its lowest level at ~2 months, it slowly increases again and becomes more or less stable between 6 and 9 months.(TIF)Click here for additional data file.


Plasmodium and helminth infection. Careful broad summarization of the existing literature suggests that Schistosoma mansoni and hookworm infections may increase the risk of clinical malaria and associated morbidities, but Trichuris trichiura infection is not associated with the occurrence of clinical malaria and related outcomes. However, findings about effect of Ascaris lumbricoides and Schistosoma haematobium infection on clinical malaria are contradictory. Furthermore, the nature of relationship of helminth infection with severe malaria has also not been determined with certainty. This review summarizes the findings of epidemiological studies of Plasmodium and helminth coinfection, placing greater emphasis on the impact of the coinfection on malaria. Possible reasons for the heterogeneity of the findings on malaria and helminth coinfections are also discussed.Understanding the impact of helminth infections on clinical malaria is useful for designing effective malaria control strategies. Plenty of epidemiological studies have been conducted to unravel the nature of interactions between Plasmodium and helminths when they coexist in a host [ Plasmodium infection may in turn impact the upcoming helminth infections and related morbidities [Although the nature of interaction remains uncertain, studies showed that an apparently true biological association exists betweenn a host , 2. Hencbidities , 2. As a Plasmodium [ Plasmodium to reproduce. Moreover, anaemia caused by helminth infection can lead to cardiovascular compensation, hyperventilation of CO2, and increased lactates which can make the host more attractive to mosquitoes [Despite such a bidirectional nature of interactions between the two groups of parasites , studiesasmodium . Helmintsquitoes . Plasmodium”, “Ascaris and malaria or Plasmodium”, “Schistosoma and malaria or Plasmodium”, “hookworm and malaria or Plasmodium”, “Trichuris and malaria or Plasmodium”, and “Strongyloides and malaria or Plasmodium” were used to search relevant references. Studies on Plasmodium and helminth coinfections in animals were excluded. In addition, human studies on the immunological evidences of Plasmodium and helminth, Ascaris, Schistosoma, hookworm, or Trichuris interactions were excluded. More than 7560 published articles were identified following the literature search. After exclusion of duplicates and screening of the titles and abstracts, a total of 107 articles were found that fulfill the inclusion criteria. About 50 articles were further excluded after reading the full-text. The characteristics of the studies on epidemiology of Plasmodium and helminth coinfection are provided in Table 1 in Supplementary Material available online at http://dx.doi.org/10.1155/2016/3083568.This paper provides a brief review of epidemiological studies on helminths and malaria interactions with emphasis on the impacts of the coinfection on malaria. Possible reasons for the inconsistency of the findings on malaria and helminth coinfections are discussed. Scholar search in PubMed, Embase, and Google was performed without restricting to language, publication date, study design, and nature of the study participants . Combinations of key words such as “helminths and malaria or Schistosoma and Plasmodium. More than 20 of these studies were conducted under natural conditions in humans. Of these studies, five looked at immunology, thus excluded from the current epidemiology focused review.Many studies have been conducted to evaluate the nature of the interactions between P. falciparum. However, findings about malaria prevalence or incidence, density of the parasite, and associated morbidities during S. haematobium infection were heterogeneous. While some studies reported decreased malaria prevalence [ Plasmodium density [ Plasmodium infection and density of the parasite [ S. haematobium. Still, other studies reported similar level of prevalence of Plasmodium infection [ S. haematobium.Most of the epidemiological studies were conducted among children infected withevalence , incidenevalence , Plasmod density , disease density , and ass density , others parasite –10 or loparasite  and enlaparasite  among innfection , 14 or dnfection , 15, 16  P. falciparum infection and related anaemia [ S. mansoni alone [ S. mansoni and S. haematobium [ S. mansoni infection [ Schistosoma and malaria seems to vary with the age of the individuals [ Schistosoma [On the other hand, prevalence , 18 or i anaemia  or hepat anaemia  increaseni alone , 21, 23 matobium , 22. Hownfection , 24. Theividuals  and inteistosoma , 25 infe P. falciparum incidence and density of the parasite after treatment of children for A. lumbricoides infection [ P. vivax and A. lumbricoides compared to children infected with only P. vivax [ A. lumbricoides infection with a low prevalence [ Plasmodium infection [ A. lumbricoides infection was also negatively correlated with the occurrence of cerebral malaria and body temperature among patients in Thailand [Randomized controlled trials in Madagascar and Comoros Islands reported increasednfection –28. SimiP. vivax . In addievalence  or incidevalence  and densnfection . A. lumbThailand , 34. A. lumbricoides infection with increased incidence of malaria [ A. lumbricoides infection and prevalence of malaria was observed among patients in Ethiopia and pregnant women in Ghana [ A. lumbricoides [ A. lumbricoides infection and prevalence [ Plasmodium density [ A. lumbricoides infection on clinical or severe malaria are inconsistent and it is difficult to make a clear conclusion about the nature of relation between the occurrence of A. lumbricoides infection and the risk of malaria based on the existing evidences.On the other hand, in a longitudinal study among pregnant women in Gabon and a case control study in Thailand, patients reported association of malaria . In addiin Ghana , 36. Sevricoides . On the evalence , 38–41 oevalence  of malar density . Overall Plasmodium infection and associated morbidities [Hookworm is widely distributed in most tropical regions where malaria is endemic . As a rebidities . Plasmodium density [ Plasmodium density [Cross-sectional studies among pregnant women in Thailand, Ghana, and Uganda reported increased malaria prevalence during hookworm coinfection , 36, 45. density , 32 amon density , 31, 48  density  of malar density . Yet, th T. trichiura infection and malaria. Perhaps this could be due to the restricted distribution of T. trichiura infections in the equatorial regions of Africa which may have resulted in a decreased risk of coinfection with Plasmodium [ T. trichiura and Plasmodium coinfection showed lack of association between the two groups of parasites. Whilst two studies observed association of T. trichiura infection with high prevalence of P. falciparum infection among patients in Thailand and Ethiopia [ T. trichiura infection and the occurrence of malaria or related outcomes. Other studies also reported lack of association between T. trichiura infection and malaria incidence [ Plasmodium density [ T. trichiura infection may not affect the occurrence of clinical malaria or associated morbidities. The two studies which showed positive association between T. trichiura infection and malaria were conducted among individuals visiting the outpatient clinics. Patients may have other infections or health problems that might have confounded/distorted the direction of relationship between these two groups of parasites.Unlike other helminth species, there are few epidemiological studies examining the relationship betweenasmodium . MajoritEthiopia , 49, thrEthiopia , 39, 45,Ethiopia , and anoEthiopia  did not ncidence  or Plasm density , 32 amon Plasmodium and helminths rather than considering intestinal helminth species independently. Although variation may exist in their pathogenicity, different helminth species have similar immunopathological impact on hosts. Most gastrointestinal helminths and Plasmodium affect host nutrition in a similar manner. Hence, it seems plausible to consider different gastrointestinal helminth species together while assessing the impact of helminth coinfection on malaria.Some studies considered pooled data of different helminth species as one variable while evaluating the nature of interactions between Plasmodium [ Plasmodium infection [While some studies reported increased prevalence , 50–52 oasmodium , 60. Stiasmodium , 38, 61 asmodium , 42, 62 nfection .Factors related to methodology, environment, host, and parasite may explain the lack of uniformity of the findings about the nature of relationship between helminth and malaria. Plasmodium and helminth infection, sample size, and the degree to which they control the effect of confounders. They were either cross-sectional, case control, or retrospective analysis of data collected previously for other purposes. Hence, it is difficult to confirm whether helminth infection occurred before the occurrence of malaria. Additionally, sample sizes in some previous studies were small. This might have reduced the power of the studies to detect differences in the occurrence of malaria and associated morbidities between helminth-infected and uninfected individuals. Variation also existed among previous studies on how the study subjects were selected and diagnosed for parasitic infection. Single Kato-Katz technique is less sensitive for examining light intensity helminth infection [ Plasmodium infections relies on blood film quality and experience of the examiner [ Plasmodium species may not be correctly identified when the examiner is less experienced.Most previous studies on malaria and helminth coinfection were not uniform in terms of design and methods used for assessingnfection . In addiexaminer . Thus, iWhile intensity and species of helminths are central determinants of infection-related impact on clinical and immune functions of malaria , 2, stud A. lumbricoides infection. However, this relationship was not maintained when data was analyzed after pooling the different helminth species in one group. Another study also showed negative correlation between S. haematobium and Plasmodium density, but this relationship was not maintained when analysis was done after pooling different helminth species together [The nature of relationship between helminth and malaria could vary based on the type of helminth species , 2. Henctogether .Contradictions in findings about the nature of relationship between malaria and helminth could also be due to variation among different geographic locations in their degree of endemicity for malaria and helminth. In areas where there is frequent exposure to malaria, strong immunity will be developed that can be affected during helminth coinfection. In contrast, malaria immunity will be less developed; therefore, the effect of helminth coinfection will be minimal when transmission intensity is low. Indeed, a study in Uganda, where malaria transmission is low, failed to show any association between helminth and malaria even when data were analyzed after stratifying by the type of helminth species and intensity of infection .In addition to methodological and environmental variations, previous studies vary in the nature of the study population. While most studies were conducted in children, some studies were conducted in adults. Some studies also involved pregnant women or immunocompromised individuals who were malnourished or had other viral, bacterial, parasitic, or chronic infections. Moreover, the genetic makeup of individuals, which may affect susceptibility of individuals to malaria, was likely different among the study population in previous studies. These factors may confound the nature of relationship between these two groups of parasites. Plasmodium were not similar among studies examining malaria and helminth interactions. Although different helminth species affect the immune system in similar manner, variation may exist in their degree of potency [ S. mansoni is known for its strong influence on the immune system [ Plasmodium cannot replicate [Intensity and species of helminths and potency . For exae system . In addieplicate . Intensieplicate . Plasmodium species could also be indispensable in evaluating the nature of relationships between helminths and malaria. P. falciparum and P. vivax vary in their pre-erythrocyte immunity profile [ P. falciparum malaria but failed to confirm the association among individuals infected with P. vivax [Similarly, the types of profile , suggestP. vivax . S. mansoni and hookworm infections may increase the risk of clinical malaria and associated morbidities, but T. trichiura infection is neither associated with the occurrence of clinical malaria nor associated with the morbidities. However, findings about association of A. lumbricoides and S. haematobium infection with clinical malaria are contradictory. Findings about the nature of relationship of helminth infection with severe malaria are also heterogeneous. It is indicated that most previous studies had limitations in methodology and design and there is a possibility that different socioeconomic and environmental conditions could confound the nature of interaction between helminth and malaria. Thus, well designed randomized controlled clinical trials involving periodic treatment with anthelmintic treatment from well-characterized populations are indispensable to make firm conclusion about the effect of helminth infection upon clinical or severe malaria and related morbidities. This will help to design effective disease management program. To make the conclusion more robust, studies should also focus on immunological analysis of the interaction considering different helminth species independently. Additionally, future studies should evaluate the effect of age, transmission intensity, and nutrition status on the nature of interaction between the two parasites.Studies have shown that helminth infection may affect the epidemiology of malaria. Careful broad summarization of the existing evidence suggests thatPlasmodium and helminth co-infection included in this review. For each included study information on the study area, reference number, age group, sample size, study design, helminth species, Plasmodium species and main finding about the nature of association of helminth and Plasmodium infection is summarized. Majority of the studies were conducted among children in Africa. P. falciparum was the most dominant species investigated in majority of the studies.Table 1 summarizes key characteristics and findings of studies on


Plasmodium falciparum and helminth infections have long been recognized as major contributors to anaemia in endemic countries. Several studies have explored the influence of helminth infections on the course of malaria in humans but how these parasites interact within co-infected individuals remains controversial.Malaria and helminthiases frequently co-infect the same individuals in endemic zones. In a community-based longitudinal study from March 2011 to February 2012, the clinical and malaria parasitaemia status of a cohort of 357 children aged 6 months to 10 years living in Mutengene, south-western region of Cameroon, was monitored. Following the determination of baseline malaria/helminths status and haemoglobin levels, the incidence of malaria and anaemia status was determined in a 12 months longitudinal study by both active and passive case detection.Plasmodium alone as the reference, children infected with helminths alone were associated with protection from anaemia . The mean haemoglobin level (g/dl) of participants co-infected with Plasmodium and helminths was higher (p = 0.006) compared to participants infected with Plasmodium or helminths alone.Among all the children who completed the study, 32.5 % (116/357) of them had at least one malaria episode. The mean (±SEM) number of malaria attacks per year was 1.44 ± 0.062 (range: 1–4 episodes) with the highest incidence of episodes occuring during the rainy season months of March–October. Children <5 years old were exposed to more malaria attacks  and were also more susceptible to anaemia  compared to older children (5–10 years old). Likewise children with malaria episodes  as well as those with asymptomatic parasitaemia  were susceptible to anaemia compared to their malaria parasitaemia negative counterparts. Considering children infected with Children below 5 years of age were more susceptible to malaria and anaemia. The high prevalence of anaemia in this community was largely due to malaria parasitaemia. Malaria and helminths co-infection was protective against anaemia.The online version of this article (doi:10.1186/s12936-016-1111-2) contains supplementary material, which is available to authorized users. Ascaris lumbricoides, Trichuris trichiura, and the hookworms Necator americanus and Ancylostoma duodenale  less susceptible to having a malaria episode compared to children in the 5–10 years old age group in bivariate analysis after controlling for haemoglobin genotype. However, the study also showed that children <5 years old were significantly  exposed to more malaria attacks compared to children in the 5–10 years old age group  to 60.1 % at 6 months and 43.5 % at 12 months  co-infection (2.9 %), Plasmodium alone (P) (13 %) and helminths alone (HL) (13.2 %). Considering children infected with Plasmodium alone as the reference and controlling for age, children infected with helminths alone were significantly associated with protection from anaemia  while those who were co-infected with both parasites were not protected from anaemia though the obvious association . However, the mean ± SEM haemoglobin level (g/dl) of participants co-infected with Plasmodium and helminths (10.53 ± 0.62) was significantly (p = 0.006) higher compared to participants infected with Plasmodium (9.61 ± 0.19) or helminths (10.45 ± 0.17) alone  proportion of children infected with Plasmodium alone (17.2 %) or helminths alone (13.7 %) at baseline , p = 0.013] compared to children in the 5–10 years age group after controlling for haemoglobin genotype, malaria parasitaemia status and helminths status in multivariate analysis , Trichuris density  and the density of other intestinal parasites  though correlation with the latter group was insignificant. Haemoglobin levels (g/dl) correlated positively with Ascaris density  and negatively with Trichuris density  as well as hookworm density  though insignificantly. However, haemoglobin levels (g/dl) correlated negatively  with the density of other intestinal parasites.Malaria parasitaemia density/µl was found to correlate negatively with haemoglobin levels , hookworm density  and positively with Plasmodium and intestinal helminths) and anaemia status and evaluated the relationship between malaria parasitaemia, malaria episodes, anaemia and helminths status. The malaria parasitaemia prevalence observed in this study was lower compared to that reported by Nkuo-Akenji et al. [Plasmodium infection, children less than 5 years old were exposed to more malaria episodes during the 1 year study period. This observation is in accordance with longstanding published reports that children below 5 years old are more susceptible to malaria. Malaria parasitaemia was predominantly associated with clinical manifestation as children who had malaria episodes had significantly higher malaria parasitaemia densities compared to those who were malaria parasitaemia positive without clinical symptoms. The highest incidence of malaria was observed in the month of March, which corresponds to the transition from the dry season to the rainy season (moderate rains), the incidence reduced slightly in the month of June (peak rainy season) and this coincided with the highest malaria prevalence recorded during the study period.There is published data on malaria and helminths co-infection in the south west region of Cameroon , 11, 23 i et al.  althoughP. falciparum and helminth infections, have long been recognized as major contributors to anaemia in endemic countries. A very high prevalence of anaemia was recorded at baseline (71.5 %) in contrast to the lower prevalences of 30.8, 11.9 and 10.9 % reported by Nkuo-Akenji et al. [Although the aetiology of anaemia is complex and multi-factorial, parasitic diseases, including i et al.  and Alemi et al. , respecti et al. . ChildreAscaris lumbricoides, Trichuris trichuria and the hookworms. This prevalence was comparable to that observed by Achidi et al. [Plasmodium and helminths were anaemic compared to a higher proportion of children infected with Plasmodium alone or helminths alone and this agrees with what was observed by Nkuo-Akenji et al. [Plasmodium or helminths alone. Co-infection with both parasites was protective against anaemia whereas children infected with Plasmodium alone were more susceptible to anaemia. This is probably because the outcome of Plasmodium–helminth interactions is beneficial to the host in that the individual negative effect of both infections on the host cancel out. The successful resolution of Plasmodium infection requires a coordinated succession from a T-helper cell type 1 (Th1) to a Th2 type response, and anything that upsets the timing or balance of this process can lead to chronic or severe infection [The baseline prevalence of intestinal helminths decreased from 19.8 to 10.2 % 6 months later probably due to the general deworming of all participants at enrolment and emphasis on the need for regular deworming. However it is worth mentioning that single stool specimen examination was employed in this case thus the possibility of reduced sensitivity. The helminths detected were almost entirely composed of the soil transmitted helminths i et al.  but lowei et al.  and Alemi et al. . Co-infei et al. , 8. Amoni et al. . A smalli et al. , but is nfection . Infectinfection  and moduPlasmodium seemed to interact negatively. Actually no co-infection was recorded involving hookworm infection in contrast to Naing et al. [Ascaris, Trichuris and other intestinal parasites seem to imply that these two groups of parasites interact positively. However, the malaria parasitaemia densities of children positive for Trichuris was higher compared to children positive for Ascaris (data not shown, Additional file Trichuris infection was associated with increased malaria parasitaemia. Ascaris infection did not appear to have any effect on haemoglobin levels while both Trichuris and hookworm infections seemed to have a negative effect and infection with the other intestinal parasites clearly had a negative effect on haemoglobin levels. In view of the variation in routes of entry to the host and the different clinical outcomes, the interaction of these helminths with malaria parasites is expected to be different from one another [The negative correlation of malaria parasitaemia with haemoglobin levels and hookworm consolidates the fact that malaria parasitaemia was an important contributor of anemia while hookworms and g et al.  who repo another .P. falciparum in adults in Thailand, whereas the second study on mothers and children in Zaire described a positive association between infection with Ascaris and the occurrence of P. falciparum as reviewed by Hartgers and Yazdanbakhsh, [Plasmodium infections, but had a far less effect on lethal Plasmodium infections and even tended to delay death in cerebral malaria models [No significant association of susceptibility to malaria during the study was observed between children positive or negative for helminths. Previous association of helminth infection with an increased risk for malaria incidence has been confirmed in two other studies. The first study reported a positive association between intestinal helminths and infection with nbakhsh, . Howevernbakhsh, . In a renbakhsh,  reports nbakhsh, . A recena models .Children less than 5 years old were more susceptible to malaria and anaemia. The prevalence of anaemia in this community was quite high and was largely due to malaria parasitaemia and to a smaller extent to helminth infection. In addition, nutritional factors probably also contributed especially to the anaemia observed at baseline. Malaria and helminths co-infection was rather beneficial to the children as this was associated with protection from anaemia. Studies assessing the effect of helminth infections on the course of malaria are very important in target malaria vaccine trial communities. Well-structured longitudinal studies investigating the underlying immune mechanisms involved in malaria helminths co-infection are very important in this regard. Our results also indicate that employing the proper intervention in a combined malaria–helminth approach will yield timely results in endemic communities (Additional file


Plasmodium falciparum or clinical malaria remains a major hurdle. In this study, an immuno-epidemiological survey was carried out between two populations naturally exposed to P. falciparum malaria to determine the immune correlates of protection.Malaria still represents a major cause of morbidity and mortality predominantly in several developing countries, and remains a priority in many public health programmes. Despite the enormous gains made in control and prevention the development of an effective vaccine represents a persisting challenge. Although several parasite antigens including pre-erythrocytic antigens and blood stage antigens have been thoroughly investigated, the identification of solid immune correlates of protection against infection by P. falciparum lysate was investigated. Purified IgG from these samples were furthermore tested in an invasion inhibition assay for their antiparasitic activity.Plasma samples of immune adults from two countries (Ghana and Madagascar) were tested for their reactivity against the merozoite surface proteins MSP1-19, MSP3 and AMA1 by ELISA. The antigens had been selected on the basis of cumulative evidence of their role in anti-malarial immunity. Additionally, reactivity against crude Significant intra- and inter- population variation of the reactivity of the samples to the tested antigens were found, as well as a significant positive correlation between MSP1-19 reactivity and invasion inhibition (p < 0.05). Interestingly, male donors showed a significantly higher antibody response to all tested antigens than their female counterparts. In vitro invasion inhibition assays comparing the purified antibodies from the donors from Ghana and Madagascar did not show any statistically significant difference. Although in vitro invasion inhibition increased with breadth of antibody response, the increase was not statistically significant.The findings support the fact that the development of semi-immunity to malaria is probably contingent on the development of antibodies to not only one, but a range of antigens and that invasion inhibition in immune adults may be a function of antibodies to various antigens. This supports strategies of vaccination including multicomponent vaccines as well as passive vaccination strategies with antibody cocktails. Anopheles mosquito, the vector. It is a leading cause of morbidity and mortality, particularly in children living in endemic regions, causing 124–283 million infections and approximately 584,000 deaths per annum with no signs of a significant decline [Malaria is a tropical disease found in most African countries including Ghana and Madagascar, because the high temperature and humidity combined with the presence of stagnant water provide ideal breeding conditions for the female Plasmodium falciparum genome sequence, which makes malaria one of the few remaining severe infectious childhood diseases without any efficient vaccine. This is caused by a combination of factors, including the multistage lifecycle of the parasite (each with stage-specific antigens), its genetic diversity, and an incomplete understanding of its immunopathology, resulting in a lack of immunological markers correlating with immunity.Effective malaria vaccines remain an elusive goal despite the availability of the Antigens expressed on the surface of asexual blood-stage malaria parasites are major targets for antibodies elicited by infection. These IgG antibodies prevent merozoite invasion of red blood cells, as well as opsonize parasitized red blood cells, and prevent cytoadherence. Thus, they form a major component of the defense against asexual blood-stage parasites and are therefore prime targets for vaccine development. Susceptibility to infection and episodes of disease decline in frequency and severity over time, but it is unclear which asexual blood-stage antigens are targets for this naturally acquired immunity. The most likely marker candidates include merozoite surface protein 1 (MSP1) and its C-terminal product, MSP1–19), apical membrane antigen 1 (AMA1) and merozoite surface protein 3 (MSP3), reflecting cumulative evidence of their role in naturally-acquired immunity to malaria based on epidemiological studies in countries such as Myanmar , Ghana –7, Kenya, apical MSP1 is a large protein which is proteolytically processed into the subunits MSP1-83, MSP1-30, MSP1-38 and MSP1-42 –13. The AMA1 appears on the surface of merozoites when released from the micronemes and undergoes processing from an 83-kDa precursor into a 66-kDa mature protein that is also known to play an essential role in erythrocyte invasion, forming the tight junction with the protein Ron2L . During Many individuals with naturally acquired immunity to malaria produce anti-MSP1-19 and anti-AMA1-66 antibodies that play a critical role in their immunity by inhibiting erythrocyte entry. There is a strong correlation between these antibody titers and the levels of protection against malaria in endemic regions [MSP3 is a 48-kDa protein found on the surface of merozoites, which unlike the other candidates, was identified by studying the monocyte-dependent parasite-inhibition effect observed following the passive transfer of IgG from immune African adults into infected Thai children . EpidemiThe present study profiled the immune response to MSP1-19, AMA1 and MSP3 within and between two diverse populations, in the malaria-endemic regions of Ghana and Madagascar, focusing on the ability of plasma from such individuals to inhibit erythrocyte invasion.Blood samples from a voluntary study population, randomly recruited in the local communities residing in Kumasi (Ghana) and Mahajanga (Madagascar) were taken in February and March 2010 (for Ghana) and January 2010 (for Madagascar). These time points correspond to the beginning of the rainy season in Ghana and the peak of the rainy season in Madagascar. Malaria transmission in Ghana is stable throughout the year and seasonal in Madagascar, with peak transmission during the rainy season , 25. A tP. falciparum 3D7, PlasmoDB, #Pf3D7_090300, amino acids 1608-1702) was produced in HEK293T in a construct containing a murine Igκ signal peptide for protein secretion and a C-terminal His6-tag for protein purification by immobilized metal affinity chromatography (IMAC), basically as described before [P. falciparum 3D7, PlasmoDB #Pf3D7_1035400, amino acids 25-354) was produced in tobacco plants (Nicotiana benthamiana) using a construct containing a murine IgG heavy chain signal peptide for protein secretion, a C-terminal His6-tag for protein purification by IMAC and C-terminal KDEL for retention in the ER as described before [P. falciparum 3D7 cultures were prepared as described before [Four different kinds of antigen preparations were used during the study. For measurement of reactivity against the AMA1 antigen, a mixture of the diversity covering variants (DiCo1-3) were used . MSP1-19d before . Two N-Pd before . Four poP. falciparum lysate, similar to the methods described before with minor modifications [The plasma samples from the African blood donors, as well as a pool of two plasma samples from European blood donors, were analysed by enzyme-linked immunosorbent assay (ELISA) to detect IgGs specific for the recombinant malaria proteins AMA1, MSP1-19 and MSP3, as well as general reactivity against the ications . Microtip-nitrophenyl phosphate (pNPP). Absorbance of the samples was recorded at 405 nm after 15 min.The plates were washed three times with PBST and bound IgG was detected with alkaline phosphatase conjugated goat anti-human IgG  for 1 h at 37 °C. After five washing steps in PBST, the signal was developed using the substrate vs the obtained absorbance. Values given are relative activity whereby the background reactivity of the non-immune European donor is equal to one. Positive reactivity was defined as reactivity superior to the European sample plus twice its standard deviation (SD). To account for the inter-plate variances, a pool of positive samples was used on each plate.Quantitative evaluation of the ELISA results were performed by calculation of the slopes of the linear range of the graphs obtained by the dilution g for 15 min in MicroSep columns  and the concentrates were sterile filtered and subsequently used in the erythrocyte invasion inhibition assay after concentration determination by Bradford analysis and confirmation of integrity by SDS-PAGE.To prevent nonspecific inhibition from heparin and other plasma components, the human antibodies were purified by Protein A affinity chromatography and tested for efficacy in invasion inhibition assays. Each 300 μl plasma sample was diluted such as to obtain 1 ml in PBS (pH 8.0) and passed over an equilibrated Protein A column by gravity flow. The flow-through was discarded and the column was washed with 35 column volumes of wash buffer. The antibodies were eluted in five column volumes of 0.1 M glycine (pH 2.5) and collected into 200 μl of 1 M Tris–HCl (pH 8.0) to neutralize the eluate and prevent denaturation. The eluates were concentrated to obtain the original antibody concentration by centrifugation at 300Plasmodium falciparum 3D7 parasites were cultured in standardized parasite culture medium , sodium bicarbonate 1 g/l, glucose 2 g/l, hypoxanthine 0.028 g/l, gentamycin 0.2 g/l, pH 7.2) and synchronized using the sorbitol method twice at ring stage during two consecutive cycles, the last one 24 h before starting the assay, as described before [P. falciparum schizonts  in 96-well round bottom plates. Cultures were kept at 37 °C for 96 h (two complete erythrocyte cycles). After this incubation time, parasite cultures were stained for 10 min using 0.1 % ethidium bromide, and subsequently analysed by flow cytometry using a BDFACSCanto II .d before . Protein2 (Chi squared) trend analysis. Spearman’s Rank Correlation Coefficient (rho) was used to test for the degree of association between test parameters. A p value of < 0.05 was considered to be statistically significant. P-values in figures are represented by asterisks  All statistical analyses were carried out using GraphPad Prism v6.07 for Windows .The results are presented as mean ± SD for normally distributed data and as means and 95 % Confidence Intervals (95 % CI) for non-normally distributed data. Comparisons were performed using the Kruskal–Wallis ANOVA tests and Dunn’s post-tests for selected multiple comparisons. In cases of comparison of two populations, Mann–Whitney U rank test was performed. Categorical data were analysed using Fisher’s exact test or χAmong the 136 participants selected in this study, 78 (57.4 %) were from Ghana and 58 (42.6 %) were from Madagascar. Approximately half (46.3 %) of the studied population were males, with a mean age of 31.9 ± 6.6 years, which was lower than that of the total study population (33.6 ± 6.8 years). The mean age of the females in the study was 35.1 ± 6.8 years. The participants from Ghana were younger (p < 0.0001) compared to their Malagasy counterparts. A similar trend (p = 0.007) was observed when the male study subjects were compared to their female counterparts.The mean reactivity of the samples was highest against the 3D7 lysate followed by AMA1 and then MSP3, and lowest in MSP1-19. The same trend was observed when the study population was stratified by country and by gender   (p < 0.0001). The mean reactivity of the male subjects was also significantly higher than that of the female subjects for MSP1-19 1.3 (95 % CI: 0.8–1.7) vs 0.6 (95 % CI: 0.4–0.7) (p = 0.0442), MSP3 1.6 (95 % CI: 1.2–2.0) vs 0.8 (95 % CI: 0.6–1.1) (p = 0.0063) and 3D7 lysate 4.4 (95 % CI: 3.5–5.3) vs 2.9 (95 % CI: 2.0–3.7) (p = 0.0006).Figure As shown in Fig. The invasion inhibition was estimated in vitro. The inhibitory effect of a positive control  was 84.8 ± 2 %. In the human study samples, despite the clear differences in antibody prevalence and reactivity, there was no significant difference in the efficacy of erythrocyte invasion inhibition when the study population was stratified by country and gender Fig. . The medFigure A Kruskal–Wallis test with a Dunn’s post test showed a gradual increase in the efficacy of erythrocyte invasion inhibition as the breadth of antibody response broadened Fig.  althoughP. falciparum 3D7 lysate within and between two diverse malaria endemic populations in Ghana and Madagascar with focus on the ability of plasma from such individuals to inhibit erythrocyte invasion. In this study, immune correlates of protection in two endemic regions were compared, in order to provide the basis for the distribution of the most antigen-specific immune responses in these areas. These study objectives were achieved by using ELISAs to measure total IgG antibody responses to MSP1-19, AMA1, MSP3 and the P. falciparum 3D7 lysate in the stratified study population. The prevalence of the antibodies was determined, and correlations between antibody reactivity, prevalence and erythrocyte invasion inhibition efficacy were investigated.This population-based cross-sectional immuno-epidemiological study profiled the immune response to MSP1-19, AMA1, MSP3 and the The relationship between the mean reactivity to MSP1-19, MSP3 and the 3D7 lysate antigens and age has been established in several studies –33. HoweThe reactivity of the samples to the three antigens  and one antigen mixture (3D7 lysate) considered in this study was related to the gender and ethnicity of the participants. This appears to be the first time in which all these antigens were examined in a single study to define the immune response in adults from malaria-endemic areas. Whereas Ghanaian men showed higher reactivity to all antigens in comparison to the Malagasy counterparts, Ghanaian women showed higher levels of reactivity against AMA1, MSP1-19 and 3D7 lysate compared to the Malagasy female counterparts  represents a holoendemic region with year-round malaria transmission, whereas Mahajanga  is meso- to hyperendemic with a seasonal transmission pattern , 37. TheA significant positive correlation between reactivity to MSP1-19 and invasion inhibition could be observed in the presented study, as well as a positive relationship between AMA1 and MSP3 reactivity and invasion inhibition albeit below the threshold of statistical significance. The clinical relevance for the specific in vitro invasion inhibition effect of antibodies which are specific for MSP1-19 and their protective effect from malaria infection has been reported before for a cohort from a highly endemic area in Kenya . In thisThe mechanism of how AMA1 antibodies protect from malaria are regarded as similar to the protection offered by MSP1-19 antibodies. Both antibody types are capable of inhibiting the merozoite invasion directly by steric hindrance –42.MSP3-specific antibodies are also described as being capable of inhibiting the invasion of merozoites into the erythrocytes directly , 43. GenThe selection of the study population was based on the absence of a clinical malaria infection for at least 2 years. Immunity to malaria is thought to be short-lived, nevertheless, continuous exposure steadily boosts the immune response . One expDespite the differences we found in polyclonal plasma IgG reactivity against AMA1, MSP1-19 and MSP3, a gender-related difference in the efficacy of invasion inhibition in vitro was not found in this study. The main aspect to consider here is that inhibition is likely brought about by a vast spectrum of different immunoglobulins—some of whose targets are yet to be identified—and not exclusively by AMA1-, MSP1-19- or MSP3-specific antibodies. Possibly, in women the protective antibody spectrum does differ as compared to men, but still unfolds the same inhibitory efficacy, at least in the in vitro assays.3 by the purification of the immunoglobulins by protein A chromatography. This might become especially important, when measuring the ADCI using monocytes, as here, IgG3 seems to play a crucial role [A technical aspect of the measurement of the invasion inhibition is the potential loss of some subclass of IgGial role .Studies such as the one presented are the essential basis for the characterization of different populations in order to define the best qualified groups of individual donors for the isolation of human antibodies , 30. FurThere is inter-population and inter-individual variation in the response of plasma samples to the malaria antigens tested. This is the first report of tests that (1) involves these selected antigens for the characterization of antibody levels in adults in two malaria endemic regions, and that (2) stratifies the study subjects on the basis of gender and geography; these may have implications on the design and distribution of a malaria vaccine.


Persistent neurocognitive impairments occur in a fifth of children hospitalized with severe falciparum malaria. There is little data on the association between different neurological phenotypes of severe malaria  and impairments in executive function.Executive functioning of children exposed to severe malaria with different neurological phenotypes (N = 58) and in those unexposed (N = 56) was examined using neuropsychological tests such as vigilance test, test for everyday attention test for children (TEA-Ch), contingency naming test (CNT) and self-ordered pointing test (SOPT). Linear regression was used to determine the association between neurological phenotypes of severe malaria and executive function performance scores, accounting for potential confounders.vs. 5.84 (4.71-6.42), P = 0.030) and SOPT (mean errors (standard deviation) = 29.50 (8.82) vs. 24.80 (6.50), P = 0.029) tests, but no differences were observed in TEA-Ch and CNT tests. Performance scores for other neurological phenotypes of severe malaria were similar with those of unexposed controls. After accounting for potential confounders, such as child’s age, sex, schooling; maternal age, schooling and economic activity; perinatal factors and history of seizures, complex seizures remained associated with efficiency scores in the vigilance test (beta coefficient (β) ) = -0.40 , P = 0.006) and everyday attention scores of the TEA-Ch test (β (95% CI) = -0.57 , P = 0.019); the association with SOPT error scores was weak (β (95% CI) = 4.57 (-0.73-9.89), P = 0.089). Combined neurological phenotypes were not significantly associated with executive function performance scores.Children with complex seizures in severe malaria performed more poorly than unexposed controls in the vigilance (median efficiency scores (interquartile range) = 4.84 (1.28-5.68) Executive function impairment in children with severe malaria is associated with specific neurological phenotypes, particularly complex seizures. Effective prophylaxis and management of malaria-associated acute seizures may improve executive functioning performance scores of children. Plasmodium falciparum malaria is the most common parasitic infection for the central nervous system in endemic areas, and seizures and impaired consciousness  are the main neurological manifestations. Compared with the unexposed group, the errors of omission (sustained attention) scores were different for complex seizures , but not for impaired consciousness (P = 0.457), malaria prostration (P = 0.562) and seizures with fever (P = 0.678). However in an adjusted linear regression model, errors of omission were not associated with specific neurological phenotypes of severe malaria. The overall average time to correct response differed between the exposed and unexposed group for complex seizures , but not for impaired consciousness (P = 0.324), prostration (P = 0.976) and seizures with fever (P = 0.552). However in an adjusted linear regression model average time to correct response was not associated with specific neurological phenotypes of severe malaria.The errors of commission scores in the The scores for errors in TEA-Ch were not different between the exposed and unexposed group for complex seizures group , P = 0.127)  Figure , but othThere were no differences in the average errors committed in the four CNT trials between the exposed and unexposed for all clinical sub-groups: complex seizures P = 0.993), impaired consciousness (P = 0.801), prostration (P = 0.590) and seizures with fever (P = 0.884). Average errors were not associated with neurological phenotypes in a linear regression model adjusted for potential confounders. Again there were no differences in the average efficiency in the fours CNT trials for all clinical sub-groups compared to controls: complex seizures (P = 0.828), impaired consciousness (P = 0.196), prostration (P = 0.246) and seizures with fever (P = 0.279). In a linear regression model adjusted for potential confounders, average efficiency in the fours CNT trials were not associated with neurological phenotypes of severe malaria  and unexposed group were consistently documented with a number of tests. The significant independent association between malaria seizures and efficiency scores of the vigilance test, which assessed errors of omission (sustained attention) and commission, suggests that seizures in malaria affect sustained attention and impulsivity in children. This is further supported by the independent association between errors of commission and complex malarial seizures in this study. Other previous studies have associated seizures and attention/impulsivity problems, althougWorking memory may be affected by complex seizures in malaria because of the more errors observed in this group compared to unexposed children in the SOPT test. This conclusion was further supported by the trend towards significance in the adjusted analysis. A previous study, using aThe mechanisms for the executive function impairment following seizures in malaria is not fully understand. Seizures may cause direct neuronal damage, compounded by the pathophysiology of impaired consciousness that may be related to the seizures and/or malaria. Therefore, the lack of an association between impaired consciousness alone and executive function scores in this study may imply that the impairments documented in the previous studies of cerebral malaria may be rThe strength of this study lies on multifaceted neurocognitive assessment tools that are likely to tap different components of executive functioning at the same time. Single and simple executive tests have been faulted because of their low predictive value of how one performs in another different test or in the real life situation. This apExecutive performance in different neurological phenotypes of malaria is provided and their different pathogenetic mechanisms can easily be related to the observed psychopathologies. The multivariate analysis is accounted for several potential confounders to ensure that the differences measured reflect as much as possible the effect of exposure to different phenotypes of severe malaria. The tests used were validated in the local population and found to possess good psychometric properties. HoweverIn conclusion, this study confirms that complex seizures in severe falciparum malaria are associated with impairments in executive functioning, suggesting possible frontal lobe involvement. Future larger studies are required to confirm these findings, accounting for potential confounders. The impairments should be assessed with more than one test since many specific cognitive components are involved, particularly impulsivity, working memory and sustained attention.


HBB, ABO and G6PD, by exploring the whole human genome in genome-wide association (GWA) studies. Molecular pathways controlling phenotypes representing effectiveness of host immunity, notably parasitemia and IgG levels, are of particular interest given the current lack of an efficacious vaccine and the need for new treatment options.Malaria remains a major worldwide public health problem with ~207 million cases and ~627,000 deaths per year, mainly affecting children under five years of age in Africa. Recent efforts at elaborating a genetic architecture of malaria have focused on severe malaria, leading to the identification of two new genes and confirmation of previously known variants in Plasmodium spp. to the development of the infection through liver and blood-stage multiplication cycles (parasitemia as a quantitative trait), to clinical malaria attack, and finally to severe malaria. Genetic polymorphism may control any of these stages, such that preceding stages act as mediators of subsequent stages. A biomarker of humoral immunity, IgG levels, can also be integrated into the framework, potentially mediating the impact of polymorphism by limiting parasitemia levels. Current knowledge of the genetic basis of parasitemia levels and IgG levels is reviewed through key examples including the hemoglobinopathies, showing that the protective effect of HBB variants on malaria clinical phenotypes may partially be mediated through parasitemia and cytophilic IgG levels. Another example is the IgG receptor FcγRIIa, encoded by FCGR2A, such that H131 homozygotes displayed higher IgG2 levels and were protective against high parasitemia and onset of malaria symptoms as shown in a causal diagram.We propose a global causal framework of malaria phenotypes implicating progression from the initial infection with We thus underline the value of parasitemia and IgG levels as phenotypes in the understanding of the human genetic architecture of malaria, and the need for applying GWA approaches to these phenotypes. Malaria in humans is caused by infection with the protozoan Plasmodium spp., falciparum, vivax, ovale, malariae or knowlesi. Plasmodium falciparum is responsible for the most severe form of malaria. No efficacious malaria vaccine exists, and current treatment options are threatened by the spread of drug resistant parasite strains. The malaria parasite has a complex lifecycle, initiated when an infected female Anopheles mosquito takes a bloodmeal, during which sporozoites are injected into the bloodstream and then invade liver cells. After asexual replication in the liver, merozoites are released into the blood and further asexual multiplication occurs in red blood cells. Merozoites mature into trophozoites, and a small proportion develop into female and male gametocytes, infectious to the mosquito, thereby completing the cycle. In endemic regions, it is expected that most individuals will become infected at some time, and a small fraction go on to develop life-threatening severe malaria with complications such as cerebral malaria, respiratory distress, or anemia. Detectable parasitemia is used to diagnose clinical malaria and high parasitemia to diagnose severe malaria (http://www.who.int/en/). The search for human genetic factors that influence parasitemia levels and other markers of anti-malarial host immunity could lead to novel molecular strategies for vaccine and treatment development, and malaria control. We therefore explore the potential of quantitative measures of malaria infection, including parasitemia and the biomarker of humoral host immunity, IgG levels, in the search for human genetic factors impacting on malaria, in this review.Malaria remains a major worldwide public health problem in spite of numerous international control efforts with ~207 million cases in 2012 and ~627,000 deaths per year, mainly affecting children under five years of age in Africa , antigen recognition (HLA-B), antibody response (IL4) and the proinflammatory response (NOS2A)  study approach. Two signals near well-known malaria protective variants in 25 cases , while aP. falciparum parasitemia. Mean parasitemia levels were found to differ between ethnicities based on studies conducted in Burkina Faso, Mali and Nepal for communities living in the same geographic area with similar levels of exposure to P. falciparum , with each measure defined as a density per uL of whole blood or as a percentage of red blood cells positive for trophozoites. Studies have generally focused on iewed in ). With riewed in . Siblingiewed in . Segregaiewed in .P. falciparum and P. vivax malaria, mean and maximum trophozoite densities during clinical episodes displayed similar heritabilities to those obtained in Senegal for both P. falciparum (0.16 and 0.16) and P. vivax (0.13 and 0.15) [P. falciparum gametocyte prevalence (0.15 - 0.57) for asymptomatic infections, but not during clinical attacks [Heritability provides a measure of the human genetic contribution (proportion of variance) to a phenotype. Several estimates have been made addressing parasitemia during clinical malaria attacks and during an asymptomatic infection. In two communities from Senegal, one holoendemic for malaria and another with seasonal transmission (mesoendemic), a heritability of 0.22 was obtained for mean parasitemia during clinical attacks in both communities. A significant heritability of 0.13 for maximum parasitemia was obtained for mesoendemic transmission only. A maximum asymptomatic parasitemia heritability estimate of 0.22 was obtained for both communities, and a significant heritability of 0.33 for mean asymptomatic parasitemia was obtained for the endemic community . In a hyparum 0.1 and 0.16Genome-wide linkage studies have been performed on various study populations leading to the identification of loci controlling parasitemia. Chromosomal region 5q31-q33, containing several cytokine genes involved in Th1/Th2 balance, was linked to parasitemia among families in Cameroon  and in tPlasmodium spp. to the development of the infection through liver and blood-stage multiplication cycles (parasitemia as a quantitative trait), to clinical malaria attack, and to severe malaria. The initial infection stimulates the immune system, elicits protective IgG antibodies that limit parasitemia levels and represent the humoral aspect of anti-malarial host immunity. The arrows imply a causal relationship but do not imply a particular direction (increase or decrease). Arrows point from genetic polymorphism to all malaria phenotypes, indicating that all hypotheses are possible until tested, and thus the causal framework can be thought of as summarizing the working hypotheses which can then be reduced and refined as study results are obtained. Parasitemia occurs early in the causal pathway, prior to clinical symptoms and can be considered a malaria endophenotype. Genetic association studies evaluating specific polymorphisms can be tested for association with any of the measurable phenotypes in the causal progression, including parasitemia and IgG levels. Previous parasitemia candidate gene studies will be reviewed with examples considered in the context of this causal framework for the complex scenarios involving mediation through IgG levels. The candidate gene studies of parasitemia involve evaluation of the causal arrow from genetic polymorphism to parasitemia.Parasitemia can be considered to be the result of two opposing forces in a tug-of-war, the pressure exerted by the malarial parasite in its multiplicative red blood cell stage, versus the pressure of anti-malarial immunity. The higher the parasitemia, the greater the malarial force, and the lower the force of anti-malarial immunity. For the study of human genetics, it would be optimal to focus on a measure of parasitemia that represents the global effectiveness of anti-malarial host immunity, which would assume factors contributing to malarial parasite pressure to be homogeneous across the study population, including parasite species and strain, and that all other parameters impacting on parasitemia, such as the initial parasite dose, are homogeneous or controlled for. Also, factors such as nutritional status, which affect host immunity without reflecting innate effectiveness should be constant in the population. These assumptions are met in part in the longitudinal study design that is limited geographically with contained vectorial transmission, and focused on a community with a similar lifestyle and socio-economic status as in a longitudinal study following an endemic community in Senegal for 22 years . Host imP. falciparum parasitemia as a binary trait in 1954 [The first genetic association studies with malaria parasitemia levels began with the observation of the co-occurrence of geographic regions of high malaria prevalence and high prevalence of certain hemoglobinopathies . Hemoglo in 1954 . He also in 1954 . Genetic in 1954 , where p in 1954 , others  in 1954  or even  in 1954 . Given tP. falciparum malaria-endemic regions displaying a protective effect. Following from this observation, lowest mean parasitemia was observed for group O compared to A, B or AB groups consistently across studies . Genetiiewed in , while oiewed in ). Severaiewed in ). Promotasitemia . In a faorphisms . In a li1 region . Interesod cells . Other pod cells . These sA key potential mediator of anti-malarial host immunity is IgG: passive transfer of IgG, purified from sera of semi-immune adults to non-immune patients resulted in clearance of parasitemia . The potHBB polymorphism on parasitemia levels. For example, a study in a population in Burkina Faso with a frequency of 0.14 of the C beta-globin allele , and a low frequency of the S allele, showed reduced maximum P. falciparum parasitemia levels among HbC subjects [HBB polymorphisms alter anti-malarial IgG production [P. falciparum erythrocyte membrane protein 1 (PfEMP1). Complex causal hypotheses , although which of the large family of human FcRs are optimally involved remains unclear . Importa FcγRIIa . Althougtrophils  and in itrophils ,33. Thustrophils . The speG6PD A- carriers had a lower increase of IgG3 levels to merozoite surface antigens during the transmission season than the non-carriers, which is consistent with carriers having a lower burden of merozoites, and indicative that protective immunity is likely to occur through other mechanisms than through humoral immunity. Such mechanisms could be cell-mediated immunity and/or physiological consequences of the blood disorder could explain clinical protection [G6PD-Mahidol (487A) on a population infected with both P. falciparum and P. vivax, the variant form was associated with protection from high P. vivax parasite density, but not high P. falciparum parasite density in humans, indicating Plasmodium spp. specificity for the protective effect [G6PD deficiency leading to protective or susceptibility effects depending on specific severe malaria phenotype as mentioned above, collectively, these studies demonstrate the causal relationship from G6PD deficiency to IgG subtype to parasite species specificity, to parasitemia levels and clinical malaria specificity. Population level longitudinal studies focused on quantitative traits, such as parasitemia levels and IgG subtype levels, and specification of the severe malaria phenotype, enabled the identification of genetic associations that also provide insight into the functional relationship between the polymorphisms at this gene and disease.Carriers of Glucose-6-phosphate dehydrogenase deficiency (G6PD deficiency) alleles are partially protected against malaria. In a study of children in an area with seasonal malaria transmission in Senegal, otection . In a ste effect . Along wPlasmodium spp. to more fully account for pathogen variability.Although to date, the main phenotypic focus of studies on the human genetics of malaria have been on severe malaria, we show here, through key examples, that malaria infection, viewed as a collection of related quantitative traits, particularly parasitemia and sub-type specific IgG levels, has high potential to increase understanding of the genetic architecture of malaria. Studies focusing on these phenotypes act as a bridge between genetic association studies and functional studies, and enable the elaboration and verification of highly specific hypotheses Figure A. Combin