The Home Management of Malaria (HMM) strategy was developed using chloroquine, a now obsolete drug, which has been replaced by artemisinin-based combination therapy (ACT) in health facility settings. Incorporation of ACT in HMM would greatly expand access to effective antimalarial therapy by the populations living in underserved areas in malaria endemic countries. The feasibility and acceptability of incorporating ACT in HMM needs to be evaluated.A multi-country study was performed in four district-size sites in Ghana (two sites), Nigeria and Uganda, with populations ranging between 38,000 and 60,000. Community medicine distributors (CMDs) were trained in each village to dispense pre-packaged ACT to febrile children aged 6–59 months, after exclusion of danger signs. A community mobilization campaign accompanied the programme. Artesunate-amodiaquine (AA) was used in Ghana and artemether-lumefantrine (AL) in Nigeria and Uganda. Harmonized qualitative and quantitative data collection methods were used to evaluate CMD performance, caregiver adherence and treatment coverage of febrile children with ACTs obtained from CMDs.Some 20,000 fever episodes in young children were treated with ACT by CMDs across the four study sites. Cross-sectional surveys identified 2,190 children with fever in the two preceding weeks, of whom 1,289 (59%) were reported to have received ACT from a CMD. Coverage varied from 52% in Nigeria to 75% in Ho District, Ghana. Coverage rates did not appear to vary greatly with the age of the child or with the educational level of the caregiver. A very high proportion of children were reported to have received the first dose on the day of onset or the next day in all four sites . The proportion of children correctly treated in terms of dose and duration was also high . Overall, the proportion of febrile children who received prompt treatment and the correct dose for the assigned duration of treatment ranged from 71% to 87% (average 77%). Almost all caregivers perceived ACT to be effective, and no severe adverse events were reported.ACTs can be successfully integrated into the HMM strategy. Home Management of Malaria (HMM) for children with uncomplicated malaria in high transmission areas in Africa is an integral part of malaria case management within the overall Roll Back Malaria (RBM) strategy . If the Plasmodium falciparum, 44 countries in Africa have adopted artemisinin-based combination therapy (ACT) as the first line treatment for uncomplicated malaria [So far, 18 African countries have adopted HMM as part of their malaria control programme . Due to  malaria . However malaria -8.A major concern with using ACT at the community level is the potential for poor adherence to the treatment schedule by both caregivers and community medicine distributors (CMDs). This could facilitate the development of parasite resistance to these expensive but currently highly efficacious drugs. Other concerns relate to acceptability by the community, the incidence of adverse events, cost and ability to provide adequate storage conditions to ensure drug stability in the community.A pilot study carried out in Ghana in 2004  providedThis multi-country study was performed in Ghana (two sites), Nigeria and Uganda all in sub-Saharan Africa. In Ghana two districts, Ejisu-Juaben and Ho were studied and in Ho district, an urban and a rural site were studied. In Nigeria, two districts Badeku and Ojoku/Ajia in Ona-ara local government were studied. The sites in Ejisu-Juaben, Ghana; Ona-ara Local Government, Nigeria; and Buguri and Iganga districts, Uganda were all rural areas. Ghana and Nigeria are located in West Africa, while Uganda is in East Africa. Table The study was conducted in three phases. The first phase involved advocacy, community mobilization, selection of CMDs and establishment of key baseline indicators. Information, Education and Communication (IEC) materials were developed and research staff recruited and trained. The intervention phase involved the training of CMDs and distribution of ACT. A year after commencement of ACT distribution, a household survey, interviewing eligible caregivers about child fevers in the past two weeks, was performed to evaluate the intervention.It was decided that to yield useful results in terms of feasibility, the study should cover populations with a minimum population of around 40,000 – the size of a typical district. The sample size for the household survey was calculated using country-specific fever prevalence rates to provide a precision of ± 5% for the estimates of coverage of ACT treatment through CMDs, assuming a design effect of two. The minimum number of households to be interviewed ranged from 700 to 768.Qualitative and quantitative data collection methods were used. Data collection tools for all the sites were harmonized prior to the intervention with some country specific modifications. The tools included the survey questionnaire and focus group discussion (FGD) and key informant interview (KII) guides, which were pre-tested and translated into the local languages. The format of CMDs' registers was also harmonized. The household survey focused on the health-seeking behaviour of caregivers of children with fever in the preceding two weeks, with an emphasis on timeliness of treatment, compliance with the prescribed treatment course, and perceptions of effectiveness and drug related adverse events. The FGD and KII guides focused on health-seeking behaviours, beliefs about the aetiology of malaria, treatment practices and community perceptions of the intervention.Community medicine distributors were trusted members of the community who were chosen by the community from a range of backgrounds, including farmers, chemical/medicine sellers, teachers, traders, community health workers, artisans, 'mother trainers', and opinion leaders Table . In GhanSelection criteria for CMDs included being a permanent resident (at least one year), trusted and respected by the community, able to keep simple records, and a willingness to serve. Uganda had a functioning pre-existing cadre of CMDs who were co-opted into the study. The number of CMDs per community depended on the community's population size. An average of two CMDs per community was used across the four sites (one CMD per 600 population).Although CMDs were not paid a salary, various motivation mechanisms were used. Some of these included rain coats, bicycles, boots, watches, T-shirts and certificates of participation. Quarterly monetary allowances varied between USD 4.5 (Uganda), USD 5 (Ejisu-Juaben) and USD 8 (Ho). In Nigeria, the CMDs received a commission of between USD 0.20 and USD 0.30 for each pack of AL dispensed in addition to periodic gifts and reimbursement of transportation fares.Two different ACTs were used in accordance with the national drug policies of the participating countries; AL was used in Uganda and Nigeria, and artesunate-amodiaquine (AS/AQ) was used in Ghana.For the AS/AQ combination, blister packs containing co-packed artesunate and amodiaquine tablets of 50 and 153 mgs respectively were used. For children aged 12 to 59 months the recommended dose of AS/AQ was one tablet of each drug once per day. However, for children of six to 11 months of age, tablets had to be broken in half and repackaged to comply with the recommended AS/AQ dose in that age group.In Nigeria and Uganda, a fixed combination of 20 mg of artemether and 120 mg of lumefantrine (AL) in two types of blister pack, one for children below three years of age and another for children three years and above, were used. The recommended dose for children aged six to 35 months was one tablet twice daily for three days, and for children aged 36 to 59 months two tablets twice daily for three days. Caregivers were advised to administer the drug after meals, preferably fatty food.In all the sites, drugs were provided through existing public health structures, at health district or sub-district level. In general, the distribution points for drugs were the local health facilities. In Uganda, CMDs replenished their stock at monthly meetings at health facility level. In Nigeria and Ghana, CMDs replenished their stocks from the health facilities as the need arose. In some instances drugs were delivered by the research team or by health staff during supervisory visits.Monthly supervision was undertaken in all sites, by health staff or by the research team Table . In eachEPI info version 6.02, SPSS version 11.0 and STATA version 9.2 were used by the four sites to enter and analyse data. A set of indicators and quantitative analyses to be performed were agreed in discussion with research staff from each site. Each research team then undertook the analysis of their own data using standard statistical methods. The FGD and KII information was transcribed and content analysis performed. From the transcripts and field notes, the responses to questions asked to explore issues of interest were grouped together, coded, categorized and analysed according to emerging themes. The findings were interpreted and reported in the form of narratives.Ethical approval for conduct of this study was obtained from the WHO Ethics Review Committee and at national level from the appropriate Ethical Review Boards. Informed consent was obtained from community heads, household heads and the caregivers who participated in the study.Based on the records maintained by the CMDs, some 20,000 fever episodes in young children were treated with ACTs by CMDs across the four study sites  were reported to have received ACT from a CMD Table . CoveragBased on the information recorded by the CMDs in their registers, a very high proportion of children received the correct dose of ACTs  each day. I did that and afterwards the child was well again. Happiness was then restored in my home".A local council chairperson of Bwalula village in Uganda explained:From 1–2 weeks during Homapak, it now takes about three months before my child gets sick. It is better than CQ when you complete the dose.The proportion of children for whom an adverse event was reported to the CMD was low (circa 1%) in both sites in which this information was available Table . At the "The change is that, previously, when children fall sick, we take some time to look for money and walk all the distance to go to the hospital. Sometimes before we get there the child's condition becomes worse. If you are lucky to get to the hospital early, after treatment you walk the same distance back or referred to the big hospital. The unlucky ones either die before they get to the clinic or die at the clinic; but ever since the introduction of this program, we have seen a change. For a year now, since the introduction of the new drug, no child has died and we have not sent any of them to the hospital".A caregiver added:"As for the drug, it works like magic!"There are two major findings from this study. First, it showed, at a larger scale than previously , that maThere was high utilization of CMDs. In an estimated population of children aged 6 to 59 months across the sites of 27,450, the CMDs treated 20,563 episodes of fever during the year of implementation (0.75 episodes of fever per child on average)). This contrasts with the figure of 0.12–0.34 episodes per year reported from African health facilities in available literature . FurtherConcerns have been raised by several authors about the potential misuse of ACT when deployed beyond the health system ,7, whichThese studies were designed for the CMDs to provide treatment on the basis of a clinical diagnosis. The possibility of moving away from a symptom based treatment to more complex therapeutic protocols based on parasitological confirmation is currently being investigated. To bring parasitological diagnosis to the community level will require reliable diagnostic tools, and evidence of feasibility and acceptability by the community.Adherence to the full treatment course has been highlighted as a matter of concern -16. AL hFurthermore, the majority of caregivers obtaining ACT from a CMD reported that they treated their children promptly  declare that they have no competing interests.All the authors except SC conceived the study; IA, ENB, BG and FB were principal investigators for their respective country's study site and together with FP, LD, BY and AB participated in the research designed and supervised data collection from the field. BY, PAB and SC performed most of the quantitative data analysis while KM analysed the qualitative data. FP, the WHO/TDR HMM research program manager monitored the four sites. All authors contributed to the draft. All authors read and approved the final manuscript.


Substantial decline in malaria transmission, morbidity and mortality has been reported in several countries where new malaria control strategies have been implemented. In Gabon, the national malaria policy changed in 2003, according to the WHO recommendations. The trend in malaria morbidity was evaluated among febrile children before and after their implementation in Libreville, the capital city of Gabon.From August 2000 to December 2008, febrile paediatric outpatients and inpatients, under 11 years of age, were screened for malaria by microscopic examination at the Malaria Clinical Research Unit (MCRU) located in the largest public hospital in Gabon. Climatic data were also collected.p < 0.01). Rainfall patterns had no impact on the decline observed throughout the study period.In total, 28,092 febrile children were examined; those under five years always represented more than 70%. The proportion of malaria-positive slides was 45% in 2000, and declined to 15% in 2008. The median age of children with a positive blood smear increased from 24(15-48) to 41(21-72) months over the study period (The decrease of malaria prevalence among febrile children during the last nine years is observed following the introduction of new strategies of malaria cases management, and may announce epidemiological changes. Moreover, preventive measures must be extended to children older than five years. Plasmodium falciparum has led to the introduction of new control strategies. Insecticide-treated bed nets (ITNs), along with prompt and effective treatment of clinical malaria cases, intermittent preventive treatment in pregnant women, and indoor residual spraying, are now being deployed widely across Africa with increasing amounts of coverage achieved . Resistance to chloroquine appeared in the 1980s and rose to 100% since 2001 [Malaria has, for a long time, been the major cause of febrile illness in sub-Saharan Africa, where 80% of the 247 million cases were reported in 2006 . The appachieved -8. One od 1990s [, Kombilance 2001 -14. In 2nce 2001 . Artemisnce 2001 . In thisP. falciparum. The population of Libreville was estimated to be 537,540 inhabitants in 2006, with children under five years of age representing 16%.This prospective and observational study was conducted from August 2000 to December 2008 in Libreville, the capital city of Gabon, where the climate is equatorial. In this urban area, malaria is predominantly caused by The Malaria Clinical Research Unit (MCRU) is a branch of the Department of Parasitology of the Medicine Faculty, located in the Centre Hospitalier de Libreville (CHL), the largest public hospital in Gabon. At MCRU, all febrile paediatric outpatients and inpatients, aged less than 11 years old, were prospectively screened for malaria. In this unit, children routinely benefit for free malaria diagnosis based on microscopic examination. Temperature, history of fever, age, sex, and location were collected prospectively.Throughout the study period, a thick blood film was done for each patient. Blood slides were processed according to the Lambaréné procedure, which is described elsewhere . Slides Monthly rainfall data of Libreville for the last decade were obtained from the meteorological services of the Agency for Air Navigation Safety in Africa and Madagascar (ASECNA).Demographic and laboratory data were recorded and given to the data manager. Data were entered and cleaned using Epi-info version 3.3.2  and were analysed with STATA version 10 . Medians with interquartile ranges (IQR) of age and geometric means of parasite densities (GMPD) are presented. Differences between groups were assessed using Pearson's chi-squared or Fisher's exact tests for proportions. For ordinal outcomes, the linear trend test (Cochran-Armitage test for trend) was used. The association between age and risk of malaria by year was assessed using odds ratio and the 95% confidence interval (OR (95%CI)). Linear regression was applied to test the significance of decreases for log-transformed data. To take into account the effect of auto-correlated observations and the season disturbances, an ARMAX model was built including covariates. We postulated a straightforward relationship between proportion of malaria cases and the monthly rainfall average measured by 20-point first-order moving average that was applied to filter seasonal variation and highlight the long-term movements in the data. Deviations in monthly rainfall were examined by comparing monthly values of the parameters  obtained during the study period (9 years).The significance level of all comparison was set at 0.05. All reported p-values are two-tailed.p < 0.01, Cochran-Armitage test for trend)  and in 2008 (n = 4684). There was a decrease in the yearly number of consultations for fever from 2002 to 2005, followed by a significant increase until 2008 (The median (IQR) age of children consulting for fever was globally of 2412-48) months. This did not significantly change from one year to another over the nine years . In 2000, the proportion of febrile illness due to malaria was 45%; this proportion declined to 12% in 2007 and 15% in 2008.There was a linear decrease in the annual number of malaria cases, from 1,785 in 2002 to 359 in 2007 with a slight increase in 2008 (p < 0.01). In 2000, parasite prevalence was highest and the median age of patients with positive slides the lowest ; from four years old, the proportion of malaria cases was similar in the different age groups  age of children with a positive blood smear significantly increased from 2415-48) to 41(21-72) months over the study period (5-48 to 4p < 0.01). A relative increase in malaria cases of 10% was observed between 2001 and 2002, followed by a drop of about two third in 2007 (10%) and 2008 (13%) . In older children (5-10 years), the proportion of malaria cases was significantly higher compared to the youngest; the risk of having malaria was 1.2 to 2.0 in older patients than younger ones (Table The study population was grouped into those less than five years and the oldest Table . In chilp < 0.01). From 2006, the same difference was only observed for patients aged 96 to 120 months old ((p < 0.02). During the study period, the median age did not significantly vary among the infected inpatients, ranging from 18 to 24 months old; whereas in the infected outpatients group, it ranged from 21 to 40 months (p < 0.01)According to the paediatric ward, data were compared between inpatients group (n = 15299) and outpatients group (n = 7502); origin was not known for 5291 children. Between 2000 and 2005, the proportion of malaria cases among children aged less than 24 months was always higher in the inpatients group (25-35%) compared to the outpatients (11-23%)  than in children aged 5-10 years old . No difference was observed when comparing GMPD between inpatient and outpatient groups (results not shown).Rainfall patterns were similar throughout the study period. June, July and August were the driest months with recorded rainfalls of 0 to 85 mm, except for the years 2000 and 2006 Figure . The resp = 0.48). Rainfall had no effect on the intercept of the trends in the proportion of malaria cases observed during the study period  or a delayed acquisition of immunity, as observed by other authors -27. HoweThe decline of malaria prevalence and the age shift towards the older children may indicate a change in the dynamics of malaria epidemiology in Libreville. It has been reported that the age at which patients experience malaria increases with a decline in exposure, and may precede changes in malaria transmission ,7,25. FuThis study was a health facility-based survey, probably not representative of the situation in the entire country; in several private health centres and rural areas, laboratory and/or well-trained microscopists are lacking. Moreover, the function of the MCRU in the CHL is to provide results of freely available microscopic diagnosis of malaria to clinicians working in the different wards, information on malaria clinical presentations, outcomes and treatments are missing.The proportion of malaria among febrile children has decreased during the last nine years, probably due to the impact of the utilization of more effective antimalarial drugs, artemisinin derivatives based therapies. After the wide-scale introduction of new effective prevention and disease management strategies, this decline was more pronounced. All these interventions are likely to be the main possible contributing factors to the observed malaria changes. Preventive measures must be further implemented and extended to older children. Studies on the prevalence of malaria in other parts of the country and record of entomological data are needed to characterize the transmission levels in Gabon.The authors declare that they have no competing interests.The study was initiated by MK and PGK; and conducted by MK with contribution of all authors. MKBA and DPMM interpreted the data and drafted the manuscript. Data were analysed by MKBA and EK. MMM, EBN, TP, ADE, MPM, EI, EEE and MCRU team participated to the field study. All authors revised and approved the final manuscript.


The planning and assessment of malaria interventions is complicated due to fluctuations in the burden of malaria over time. Recently, it has been reported that the burden of malaria in some parts of Africa has declined. However, community-based longitudinal data are sparse and the reasons for the apparent decline are not well understood.Malaria prevalence and morbidity have been monitored in two villages in north-eastern Tanzania; a lowland village and a highland village from 2003 to 2008. Trained village health workers treated presumptive malaria with the Tanzanian first-line anti-malarial drug and collected blood smears that were examined later. The prevalence of malaria parasitaemia across years was monitored through cross-sectional surveys.The prevalence of malaria parasitaemia in the lowland village decreased from 78.4% in 2003 to 13.0% in 2008, whereas in the highland village, the prevalence of parasitaemia dropped from 24.7% to 3.1% in the same period. Similarly, the incidence of febrile malaria episodes in the two villages dropped by almost 85% during the same period and there was a marked reduction in the number of young children who suffered from anaemia in the lowland village.There has been a marked decline in malaria in the study villages during the past few years. This decline is likely to be due to a combination of factors that include improved access to malaria treatment provided by the trained village helpers, protection from mosquitoes by increased availability of insecticide-impregnated bed nets and a reduced vector density. If this decline in malaria morbidity is sustained, it will have a marked effect on the disease burden in this part of Tanzania. There have been previous efforts to eradicate or, if this was not possible, to eliminate malaria, but these ambitious goals have not been achieved in large parts of Africa -5 despitAnopheles gambiae is the most prevalent vector in the lowlands, while Anopheles funestus predominates in the highlands. In previous studies, entomological inoculation rates (EIR) have been reported to be more than 100 in the lowlands, but less than 30 infective bites per person per year at intermediate altitudes. In the highlands, mosquito densities are too low to allow reliable EIR measurements, but an EIR of 0.03 infective bites per person per year has been extrapolated  up to 2006 and artemether-lumefantrine [ALu] from 2007 to date), paracetamol, slides, blood lancets, gloves, treatment charts, febrile case detection forms and storage boxes. CORPS were instructed how to collect blood smears under aseptic technique and to preserve these until collected by a team of supervisors composed of a clinical officer and a laboratory technician. CORPS were also instructed on how to treat patients with the first-line anti-malarial drug and an antipyretic, and how to refer patients who had severe symptoms or who did not respond adequately to the first-line anti-malarial treatment to Korogwe District Hospital. Villagers were informed that they could call on the CORPS at any time if they had symptoms of malaria. From suspected malaria cases, the CORPS would then collect basic clinical data, obtain a blood smear for malaria microscopy and institute presumptive malaria treatment. Patients with severe symptoms or symptoms suggestive of other diseases than malaria were referred to the local dispensary or the District Hospital.Malaria cross-sectional surveys were conducted in the two study villages from 2003 to 2008 Table . The sur® photometer . Whole blood was used to prepare thick and thin blood smears for malaria microscopy. These were stained with 10% Giemsa stain for 15-20 minutes after fixing thin smears with methanol. Asexual and sexual parasites were counted against 200 and 500 white blood cells, respectively. The differentiation of malaria parasite species was confirmed by microscopy of thin smears. A blood smear was declared negative only after examination of 200 high power fields. The density of asexual parasites was calculated assuming 8,000 leucocytes per μl of blood and expressed as parasites per μl. Slides were red retrospectively and during surveys malaria treatment was only given to patients complaining of malaria symptoms.Five millilitres of venous blood were collected from individuals aged three years or more. For children below three years, 300-400 μl of capillary blood was obtained by finger prick into an Eppendorf tube containing EDTA. The haemoglobin (Hb) concentration of each participant was measured using a HemoCuePlasmodium falciparum parasitaemia of any density. The summary measures consisted of percentage for categorical variables and means for continuous variables. Incidence was calculated as documented number of febrile malaria episodes to the population size in each age group and/or village per year.All data were double-entered into a Microsoft Access database, cleaned, validated and transferred into Stata version 9  and R 2.7.0 for statistical analysis. Fever was defined as an observed axillary temperature of ≥37.5°C and/or a history of fever within the previous 24 hours. A febrile malaria episode was defined as an episode of fever with a positive blood smear for asexual Plasmodium falciparum, parasiteamia across the years, number of individuals with parasitaemia during cross-sectional surveys were modelled as a binomial random variable using logistic regression. The model was fitted for each village separately and was adjusted for the effect of age and use of bednets. To compare the risk of Plasmodium falciparum infection between the two villages, a similar model was fitted for the two villages. In addition, linear regression models were used to determine the changes in mean haemoglobin (Hb) concentration across years and effect of Plasmodium falciparum parasiteamia and season on Hb levels while controlling for age and reported use of insecticide-treated or non-treated bed nets. In this model the effect of year was fitted as a categorical variable. A statistical significance level was considered at P-value < 0.05.In modelling the risk of Ethical clearance was granted by the Medical Research Co-ordinating Committee of the National Institute for Medical Research, Tanzania. Prior to the study, meetings were held with local authorities and with the villagers in each study village, during which the aims of the study were explained. Informed consent documents for the study were prepared in English and translated into Kiswahili before administration to both village leaders and participants. Written informed consent to participate was obtained from each study individual or from his or her parent or guardian. Villagers were free not to participate in the study without giving any reasons, or being disqualified from any medical services that were provided to all villagers throughout the study period. Feedback to the village has been performed continuously during the study.P. falciparum was detected in 78% and 25% of the residents of lowland and highland villages, respectively. In the subsequent cross-sectional surveys conducted each year, the prevalence of P. falciparum decreased steadily in both villages as shown Figures P. falciparum infection was higher in residents of lowland village, OR = 12.28  compared to highland village. The risk of infection decreased by year, with an OR of 0.564  and 0.660 , per year compared with the baseline, for lowland and highland villages, respectively. The decline in the point prevalence of parasiteamia was seen in all age groups in both villages . When the effect of the use of bed nets was assessed in each village separately, it was found that the use of bed nets was not associated with a reduction in the risk of malaria parasitaemia in highland village , but was significant in lowland village .The age specific incidence of malaria episodes by village is shown in Figure Plasmodium falciparum parasitaemia was associated with a lower mean haemoglobin concentration (Table The mean haemoglobin concentration of individuals living in highland village was higher than that of those living in lowland village. In highland village, mean haemoglobin concentration was highest in 2003, whereas in lowland village it was the lowest in this year. Similarly, the mean haemoglobin concentration of children aged 0-3 from lowland village was markedly lower in 2003 than in the following years Figure . AdjustiThis community based study assessed the malaria burden in two villages in Korogwe district, north-eastern Tanzania by collecting data in repeated cross-sectional surveys and by employing CORPS to monitor malaria morbidity over six consecutive years. In this study, lowland residents had almost a twelve-fold increase in the risk of malaria parasitaemia compared to their counterparts in the highland village. At the start of the study, the lowland village could be categorized as high transmission whereas transmission in the highland village could be categorized as moderate. During the six years of the study, there was a marked and progressive decline in the malaria burden and this was reflected in the point prevalence of parasitaemia, the incidence of clinical episodes of malaria and in the mean haemoglobin concentration.It is well established that immunity to malaria is a function of age, and that in areas of high transmission the malaria burden mainly is carried mainly by children and infants, while in areas of low transmission, all age groups are affected . As malaDuring the study, the malaria burden also decreased among inhabitants of highland village and among the older age groups in lowland village, but because the decline in burden was more pronounced among the young children in lowland village, the net effect was that, at the end of the study the malaria burden appeared to be more equally distributed among the different age groups. During the study, many cases of severe malaria were not documented. This is probably due to the fact that a relatively low number of individuals were surveyed, and that these individuals had access to prompt and effective treatment. Thus, how the decline in transmission has affected the incidence of severe malaria in its different manifestations could not be estimated. This, is an important question as cerebral malaria has been reported to affect slightly older children than those affected by severe anaemia, and a declining incidence might lead to an increase in this severe form of malaria ,26.Other studies mainly based on data collected at health facilities have documented that malaria transmission seems to be falling in other parts of East Africa and in some of these studies the decrease has been attributed to specific interventions -18. In tThere is no simple explanation for the decline in malaria in the study villages, and it is rather an unsatisfactory situation of having to assign this to a combination of different factors such as climatic changes, better health provision, increased coverage of bed nets and socio economical development. It is very important to establish the relative contribution of these and other factors to the decline in malaria, as this will shed light on the risk that malaria might strike back. Entomological studies in the study villages were not done during the study period and whether there have been changes in mosquitoes distribution across years is not established.In conclusion, there has been a dramatic progressive decline in point prevalence of malaria in the study villages during the study period, and in parallel the malaria disease burden has fallen, especially among those most affected previously, the infants and young children living in the village where malaria transmission was high previously.The authors declare that they have no competing interests.All authors were involved in the design of the study. LV, BM, AK, ML, TT and JL conceived the study. BM, LV and JL and were responsible for the implementation of the study in the field. BM coded the data and supervised data entry. BM and JL analysed data. JL wrote the first draft of the manuscript. All authors were involved in revising the manuscript, and read and approved the final version.


Plasmodium knowlesi is a cause of symptomatic and potentially fatal infections in humans. There are no studies assessing the detailed parasitological response to treatment of knowlesi malaria infections in man and whether antimalarial resistance occurs.P. knowlesi infections. These patients were given oral chloroquine for three days, and at 24 hours oral primaquine was administered for two consecutive days, primarily as a gametocidal agent. Clinical and parasitological responses were recorded at 6-hourly intervals during the first 24 hours, daily until discharge and then weekly to day 28. Vivax malaria patients were studied as a comparator group.A prospective observational study of oral chloroquine and primaquine therapy was conducted in consecutive patients admitted to Kapit Hospital, Sarawak, Malaysian Borneo with PCR-confirmed single 50) and 90% (PCT90) parasite clearance were 3.1  hours and 10.3 (9.4-11.4) hours. These were more rapid than in a group of 23 patients with vivax malaria 6.3 (5.3-7.8) hours and 20.9 (17.6-25.9) hours; P = 0.02). It was difficult to assess the effect of primaquine on P. knowlesi parasites, due to the rapid anti-malarial properties of chloroquine and since primaquine was administered 24 hours after chloroquine. No P. knowlesi recrudescences or re-infections were detected by PCR.Of 96 knowlesi malaria patients who met the study criteria, 73 were recruited to an assessment of the acute response to treatment and 60 completed follow-up over 28 days. On admission, the mean parasite stage distributions were 49.5%, 41.5%, 4.0% and 5.6% for early trophozoites, late trophozoites, schizonts and gametocytes respectively. The median fever clearance time was 26.5 [inter-quartile range 16-34] hours. The mean times to 50% (PCTP. knowlesi.Chloroquine plus primaqine is an inexpensive and highly effective treatment for uncomplicated knowlesi malaria infections in humans and there is no evidence of drug resistance. Further studies using alternative anti-malarial drugs, including artemisinin derivatives, would be desirable to define optimal management strategies for  Plasmodium knowlesi causes symptomatic infections in humans throughout South-east Asia in areas inhabited by its natural macaque hosts (Macaca fascicularis and M. nemestrina) and mosquito vectors of the Anopheles leucosphyrus group .To explore 50 and PCT90 for late trophozoites to be shorter (PCT50 2.5 [95% CI: 2.3-2.9]) than other asexual stages, while gametocytes took longer to clear .Three hourly slides performed in nine selected patients to determine stage specific clearance times found the PCTP < 0.01). The fever and parasites cleared at a similar time after treatment in knowlesi infections, while the fever cleared sooner than parasites in the vivax group.The calculated parasitaemia at the time of fever clearance was significantly lower in the knowlesi group . This suggests a low level of knowlesi malaria transmission and, therefore, low background levels of immunity in the population. The successful response to treatment with chloroquine and primaquine also implies that these infections are chloroquine and primaquine naïve, supporting a zoonotic mode of transmission.P. knowlesi is difficult to quantify. Using primaquine would seem unnecessary since gametocytes appeared sensitive to chloroquine in the first 24 hours. P. knowlesi is not known to have a hypnozoite stage [FCTs and PCTs were similar in the knowlesi group, while fever cleared before parasitaemia in the vivax group. This suggests that knowlesi malaria has a higher pyrogenic density than vivax malaria but it may also reflect differences in immune status between the two groups. Under Malaysian Ministry of Health treatment guidelines, oral primaquine was administered to knowlesi malaria patients for two days in this study as a gametocidal drug. The effect of this on both asexual and sexual forms of te stage . If perste stage ,22,23.P. knowlesi is sensitive to most antimalarial drugs including clindamycin [P. knowlesi W1 strain, while proguanil and pyrimethamine resistance were inducible in experiments with the P. knowlesi Nuri strain [Besides chloroquine, other anti-malarial agents have been used with apparent success for knowlesi malaria. These include mefloquine , quininendamycin , naphthondamycin , sulphonndamycin , tetracyndamycin , trimethndamycin  and erytndamycin . In Rhesi strain -35. Althi strain ,36,37.P. falciparum or P. vivax co-infection had been excluded. Further studies using alternative anti-malarial drugs, including artemisinin derivatives, would be desirable to define optimal management strategies for P. knowlesi.In the 1980's, oral chloroquine was thought to be more effective than oral quinine as treatment for chloroquine sensitive falciparum malaria . ChloroqP. knowlesi, reflecting a zoonotic mode of transmission and thus an absence of prior drug pressure. Further carefully-designed studies are required to determine whether chloroquine is effective in severe knowlesi malaria.Chloroquine is an inexpensive, highly effective and well-tolerated anti-malarial drug for the treatment of naturally-acquired uncomplicated knowlesi malaria in humans. There is currently no evidence of drug-resistant FCT: fever clearance time; PCT: parasite clearance time; 95% CI: 95% confidence interval; PCR: polymerase chain reaction.The authors declare that they have no competing interests.BS and TMED designed this study and wrote the paper with CD and JC-S. CD and MZR recruited and managed the patients. CD was responsible for collection and entry of clinical and laboratory data, and together with TMED, analysed the data. SKZ and PCSD, supervised by JC-S, conducted molecular detection assays. All authors read and approved the final manuscript.


Former studies have pointed to a monocyte-dependant effect of antibodies in protection against malaria and thereby to cytophilic antibodies IgG1 and IgG3, which trigger monocyte receptors. Field investigations have further documented that a switch from non-cytophilic to cytophilic classes of antimalarial antibodies was associated with protection. The hypothesis that the non-cytophilic isotype imbalance could be related to concomittant helminthic infections was supported by several interventions and case-control studies.We investigated here the hypothesis that the delayed acquisition of immunity to malaria could be related to a worm-induced Th2 drive on antimalarial immune responses. IgG1 to IgG4 responses against 6 different parasite-derived antigens were analyzed in sera from 203 Senegalese children, half carrying intestinal worms, presenting 421 clinical malaria attacks over 51 months. Results show a significant correlation between the occurrence of malaria attacks, worm carriage (particularly that of hookworms) and a decrease in cytophilic IgG1 and IgG3 responses and an increase in non-cytophilic IgG4 response to the merozoite stage protein 3 (MSP3) vaccine candidate.The results confirm the association with protection of anti-MSP3 cytophilic responses, confirm in one additional setting that worms increase malaria morbidity and show a Th2 worm-driven pattern of anti-malarial immune responses. They document why large anthelminthic mass treatments may be worth being assessed as malaria control policies. Deciphering the interactions between the malaria parasite with the human host and understanding how we acquire immunity to malaria has been fundamental to our malaria vaccine research program. Over the course of the program, we have extensively studied the characteristics of immune responses of children and adults to malaria The pattern of acquisition of resistance to malaria and the role of antibodies should be reminded first. Children in hyperendemic areas are at high risk of dying from malaria until the age of 5. Though this risk then decreases, they remain susceptible to malaria attacks until they reach the ages of 15–20 years This immunity is due to antibodies. Protection can be induced by passive transfer of IgG from malaria-immune adults to malaria patients These studies, the first of their kind, showed that, in contrast to current beliefs, immune responses were not absent in non-protected individuals. In fact, they were present and abundant, but were qualitatively different. Among non-protected individuals, aged 1–10 and up to 15 years, non-cytophilic classes of antibodies IgG2, IgG4, and IgM were the most abundant. This stood in stark contrast to responses in protected adults who had twice as much cytophilic IgG1 and IgG3 sub-classes compared to the noncytophilic classes This, in turn raised the question of why children and adolescents were producing a majority of non-cytophilic classes of antibodies. The possibility that this was merely due to an immaturity of the immune system during childhood and adolescence seemed unlikely. We considered the fact that, children often differ from adults in malaria endemic areas by the outstandingly high loads of worms they harbour To explore the possibility that helminthic infections may have an effect on acquisition of clinical immunity against malaria, we initiated several studies on worm-malaria interactions. Most of them indicated a deleterious role of worms on malaria immunity Ancylostoma, Ascaris, Trichuris, Enterobius, or systemic parasites such as Schistosoma mansonii and haematobium , confirmed a markedly higher incidence of clinical malaria in wormy individuals and showed that all types of helminths could be involved, either intestinal parasites, such as Although the results from the clinical studies indicated a clear interaction between worm infestations and occurrence of malaria attacks, they could not address the immunological hypothesis of a Thelper type 2 imbalance due to worms, either because the groups were too small to provide significant immunological data or simply because no biological specimens were available in those retrospective studies.We therefore undertook a prospective study in Casamance, Southern Senegal, in a cohort of 203 children who were assessed at the beginning of the study during the non-transmission season, and closely followed-up for a total of 51 months. Immunological, epidemiological and clinical data gathered in this manner provide, for the first time, the grounds to our immunological hypothesis of a worm-induced Th2 drive.The study was designed to be a prospective study aimed at following a cohort of children 1–14 years of age (mean±1SD: 6.9±3.1) for the occurrence of worms and attacks of malaria over a 4 year period (51 months).It took place in Mlomp, a village of ca. 8,000 inhabitants in Casamance, South West Senegal, where the local economy is based on subsistence farming, rice culture and fishing. Three similar adjoining suburbs surrounding the local Health care Unit were selected. All households are built on an identical model  and scattered in a similarly forested environment characterized by a few high trees. The village is surrounded by numerous rice fields which are a major source of intestinal helminth infections. At the start of the survey, bednets were not used by the children, and this was frequently verified throughout the 51 month follow-up.Plasmodium (P). falciparum was present in more than 95% of the clinical malaria cases. The main vector was Anopheles gambiae with an estimated 25 infective bites per individual per year in this village.Malaria transmission is seasonal, with a low monthly incidence of malaria attacks in June, which gradually increases up to a maximum in September, approximately 3 months after the peak of rainfalls. During the study period, the incidence of malaria morbididy was 52% and This longitudinal study took place between November 2000 and January 2005. The cohort was constituded based on an initial stool examination of 455 children. In order to constitute two sub-groups of equal size, one with, the other without worms, and paired for age, i.e. with no age imbalance between the two groups, 238 out of the 455 children aged 1–14 years could be included in the study according to those criteria. Thirty-five children could not be followed-up continuously at each time point, and therefore, only 203 (85.3%) children fulfilled the requirement of continuous follow-up during the 51 months of survey and were included in the final analysisThis study received ethical approval from the National Ethical Committee of Senegal. All patients (or their parents) gave formal informed consent after detailed explanations given in Diola language by a local health care assistant and the agreement was systematically obtained in the presence of a witness from the village.Stool samples were obtained at enrolment and annually from each child and screened for eggs and parasites by trained parasitologists both by microscopic observation of stools, as well as after using the eggs concentration technique described by Bailenger Briefly, stools were diluted 10 times in a dilution buffer . After 1 minute of incubation, the stools suspension was filtered through a metalic mesh and a volume of sulfuric ether was added to the same volume of filtered material in a centrifuge tube, and then emulsified by vigorous agitation. Following centrifugation at 1500 rpm for 1 minute, the pellet was sampled for microscopic examination. Stools found negative were re-tested a second time, a few days later.In Mlomp village, the parasite loads, mostly due to hookworms, reached an average of 1526 eggs/gramme of faeces, i.e. can be considered moderate to high.Stool examinations were repeated every year and individuals from the negative group who had become positive on year 1 and 2 were treated with 400 mg Albendazol®. In addition, it was agreed that, if clinical symptoms associated with helminth infection would occur, the child would be treated of his parasite infection by the nurse running the local dispensary.Finally, on year 3 and 4, in agreement with our initial protocol and in order to comply with ethical considerations, an anti-helminthic treatment of 400 mg Albendazol® was given once a year to all participants.The incidence of malaria attacks was measured by both active and passive case detection. Parents were asked to refer children to the dispensary in case of any abnormal symptom. In addition, households were visited at least weekly by the medical team continuously stationed in the village during the transmission season, so as to ascertain that suspected cases had been referred to the dispensary. Compliance estimated by weekly visits was high, particularly as nurses from the dispensary deliver free medical care. For each single case of fever (i.e. axillary temperature ≧37.5°C), a complete clinical examination was performed by a trained medical staff and under the supervision of the principal investigator (Dr. P.B.). A thick blood smear was systematically made and rapidly examined for the presence of malaria parasites. Slides were stained with Giemsa and examined against 200 leukocytes if positive, or against 400 leukocytes prior to being declared negative.P. falciparum malaria attack was defined by the presence of both fever and P. falciparum parasites with no clinically apparent other cause of fever. Children presenting with a malaria attack were given an antimalarial curative treatment. Intramuscular quinine injection was given in 95% of the clinical cases, followed by oral quinine for 4–5 days (25mg/kg/day in three doses). Over the two last years of the survey quinine was progressively replaced by artesunate-amodiaquine combination (arsumax™ and camoquin™), which accounted the last year for about 50% of the treatments. Recovery was considered as a further indication that the febrile episode was indeed an actual clinical malaria attack. From these data, prevalence, incidence density per time unit , and geometric mean annual incidence, used to express the risk of malaria in the study population were calculated.A No case of complicated malaria was recorded in the cohort of children enrolled for this study during the entire follow-up.P. falciparum antigens.Sera samples were obtained by venous puncture at the start of the survey (in November 2000) from each child of the cohort and the parasite-specific antibody contents from this first year blood samplings were evaluated by ELISA. Individual sera were tested for IgG1, IgG2, IgG3 and IgG4 subclasses against various P. falciparum-derived antigens (amino-acid numbers referring to 3D7 clone) tested were a) MSP3b(184–210) a 27 AA long synthetic peptide corresponding to the major B cell epitope of MSP3 (163–230), including the same region c) MSP3-CTerminus(191–354), AMA1 a gift of Dr Robin Anders, GLURP-R0, a kind gift of M. Theisen The IgG subclass antibodies to the different antigens were detected and quantified essentially as previously described Human subclass specific mouse monoclonal Abs were used and assayed using peroxidase-labelled goat anti-mouse IgG (at 1∶2000) as previously described Antibody responses are expressed as Arbitrary Units obtained by calculating the ratio of test OD values obtained for a given test sample divided by a background value defined as the mean plus 3 SD of the absorbance values obtained using a subsample of 40 negative control sera from malaria-naïve blood donors.P. falciparum-specific humoral responses were tested by two successive approaches. Firstly, the pattern of parasite-specific antibody responses was evaluated with regard to the presence or absence of intestinal helminths (used as a dichotomous variable) in univariate tests. Then, age and parasite-specific antibody responses were used as continuous explanatory variables in nominal logistic regressions. Secondly, the pattern of P. falciparum-specific humoral responses was determined with regard to the occurence or non-occurence of malaria attacks during the 51 months of follow-up. In additional studies, the (Log+1)-transformed total number of clinical episodes was used as a continuous variable and the explanatory variables included in multivariate analysis were age sex, household/family and the antibody responses, expressed as Log-transformed arbitrary units as in previous studies We analyzed the data using the JMP® software and Statview® 5 softwares, both from SAS Institute . We used univariate and multivariate analysis to explore factors potentially associated with susceptibility to malaria attacks. P. falciparum-specific antibody responses) on the significance of the comparisons. A stepwise logistic regression model retaining explanatory variables with P values less than 0.05 in the initial study was used to reduce the number of explanatory variables tested in the full model.We tested the hypothesis that a) children with high levels of antibodies to some of the parasite antigens tested were less susceptible to clinical malaria and b) children without worms differed from children with helminths in term of relative susceptibility to malaria. The different helminth infections observed were initially grouped in one single binary exposure variable corresponding to the presence or absence of helminth infection. We also looked for a potential individual effect of some specific type of infections after identification of the infecting parasite species. We first looked at the 2 subgroups separately using non-parametric tests to analyze non-Gaussian variables, whereas variables with normal distribution were tested by parametric tests. Multiple linear regression models were used to determine the potential effect of different explanatory variables (such as age and The 203 children from Mlomp had a mean age of 6.9±3.1 years (range: 1–14 years) at enrolment in the year 2000. At baseline, children without intestinal helminths (n = 98) were 6.8±3.4 years old (range: 1–13 years) whereas children with worm infections (n = 105) were 7.0±2.9 years old (range: 1–14 years), i.e. the 2 subgroups of children did not differ for age. Three years after the start of the follow-up, the weight of the children was comparable between the 2 subgroups . The sex ratio was similar in the two subgroups with 46.8% males.rate of malaria attacks per child was 0.614 (95% CI = 0.503–0.734).Over the duration of the study, the prevalence of malaria attacks peaked three months after the end of the rainy season (not shown). The monthly prevalence of symptomatic malaria episodes ranged from 0% in May 2001 up to 13% in November of the same year. The geometric mean annual incidence During the 51 months period of active and continuous survey, 47 out of 203 children (23.1%) had no detectable episode of symptomatic malaria diagnosed, whereas 421 malaria attacks were recorded in the 156 children who experienced between 1 to 9 clinical malaria episodes (mean ± 1SD = 2.7±1.7 attacks/child), and received first-line treatments. Fourty five children had 1 attack, 42 had 2 attacks, 29 had 3, and 40 had >4 attacks (range 4–9 attacks). The incidence of malaria episodes over the study period ranged from 81.13% (2.43±2.10 attacks/51 months) in 53 children aged 0–4 years, to 72.34% (1.81±1.85 attacks/51 months) in 47 children aged 10–14 years.P = 0.0014). This was most likely due to a combination of factors including improved access to medical treatment, introduction of artesunate-amodiaquine combination P = 0.069).During the study, and notably during the last two years of the survey, the occurrence of clinical malaria in the cohort of children was found to progressively decrease with time  were found infected at enrollment and with at least one intestinal helminth species. 60% of the remaining 98 children initially free of detectable helminthic infection remained so throughout the study. Fourty percent of helminth free children became helminth positive over the follow-up period , 14 being infected twice.Ascaris lumbricoides was found in 21, Strongyloïdes stercoralis in 17, Enterobius vermicularis and Trichuris trichiura in 6, and Hymenolepis nana in 3 individuals. Prevalence of each helminth species did not differ significantly depending on gender, age, or year of study. A mean number of 1526 eggs per gramme of stools (range: 40–5800) was determined by the concentration technique employed, and only 10% of the children with helminth infections had less than 80 eggs per g of stools, indicating moderate to high helminthic loads in this cohort of children.The most frequent species was hookworm, detected in 87 of the helminth-infected children. We addressed the existence of potential differences in spatial distribution of the prevalence of infections either as a function of neighbourhood, of household or of family links. Children were living in 3 main and geographically contiguous areas of Mlomp and belonged to 6 major family groups living in compounds similarly structured. As far as this could be evaluated, the everyday way of life, the social status, the housings and the socioeconomic indicators were very similar from one family to the other, with most parents working as agricultors in rice fields surrounding the village. On the basis of the above parameters, the prevalence of intestinal helminths, the occurrence of malaria attacks and/or the delay observed before occurrence of the first malaria attack, did not show any particular geographic clustering, i.e. these factors were equally distributed in the different geographic locations of the children. Of note, in several occasions, both helminth infected and non-infected children were identified in a same family.P = 0.0215). Over the first two years of follow-up, children with intestinal helminths had 1.943±0.309 malaria attacks, whereas children without helminths had 1.187±0.107 attacks (P = 0.0057).The results show that the presence of any helminth species was significantly associated with an increased risk of malaria attack . During P = 0.0218).When comparing 2 similar transmission periods of 6 months at 1 year interval, i.e. in 2000 and 2001, the likelihood of developing symptomatic malaria was markedly higher for children with intestinal helminths than for those without detectable helminth infection . OR and 95%CI were 2.72 (1.35–5.45).After 51 months of follow up, the higher incidence of malaria attacks remained in helminth-positive as compared to helminth-free individuals .At the end of the survey, Kaplan Meier estimates showed that 28.7% of worm free children had not suffered from clinical malaria, whereas only 11.7% of helminth positive children, i.e. 2.45 fold less individuals, remained free of symptomatic malaria, .All the available variables with a potential impact on malaria attacks were included in a logistic-regression model. Stepwise logistic regression analysis starting with a full model retained helminth infection [odds ratio = 2.69; and 95% confidence interval CI = 1.34–5.39; Thus the association between helminth infections and risk of malaria attacks remained significant after controlling for potential confounders. Given that all the children of the cohort lived in a similar environment and that anti-parasite immune responses were comparable between the children originating from the different compounds of Mlomp village (see below), it was considered that the risk of exposure to malaria was similar for all children.Therefore, in the present study we conclude that the major risk factor associated with clinical episodes of malaria was whether or not the children had detectable intestinal helminthic infection(s) at the beginning of the 51 months of follow-upAntibodies specific for each malaria antigen tested were detected in the children. For total IgG, there was no statistically significant difference in the magnitude of the responses to any antigen between the two groups, although responses tended to be higher in the “protected” group  than in the “susceptible” one . Futher studies analyzed isotype specific responses to each antigen.P.falciparum extract than children who had one or several malaria attacks (not shown). Similarly, children who remained free of malaria had higher levels of antibodies against Glurp-R0, SERP and the 3 MSP3-derived antigens than children of the same mean age who experienced malaria attacks. In an univariate type of analysis  or to the conserved region of MSP3 (DG210), targeted by biologically active antibodies. Protection from malaria attacks was also found associated with high anti-DG210 IgG1 levels , high anti-DG210 IgG3  and low anti-DG210 IgG4 levels , as well as with high anti-MSP3-CT IgG1 responses , high anti-MSP3-CT IgG3 responses , and with low anti-MSP3CT IgG4 responses . Anti-MSP3-CT IgE responses were also tested and found similar in children with (1.24±0.43) and in children without malaria attacks . No other antibody specificity showed a similar association with protection in Mlomp, as immune responses to AMA1, Glurp-R0 and Serp became no longer significant in the more stringent multivariate analysis.The potential association between these various antibody responses and the relative susceptibility to malaria attacks over the 51 months of follow-up was then tested by multivariate analysis. The variable that we wished to explain, i.e. the number of malaria attacks, was tested by taking into account several potentially explanatory variables including age, together with antibody responses used as continuous variables. Protection from malaria attacks was found significantly associated with high anti-MSP3b IgG1 responses [L-R Chisquare = 5.99; P = 0.056), anti-MSP3b IgG1 (P = 0.0013), anti-MSP3b IgG3 (P = 0.0581), anti-DG210 IgG1 (P = 0.0126), anti-DG210 IgG3 (P = 0.057), anti-MSP3CT IgG3 (P = 0.053). IgG4 isotype responses to DG210 (P = 0.018) and to MSP3CT (P = 0.026) followed an opposite pattern, being found at higher levels in children carrying worms. In contrast, no significant change in the pattern of anti-AMA1 antibody responses was found associated with the presence or absence of intestinal helminths.As shown in P = 0.013); lower levels of anti-MSP3b IgG3 , and conversely, with higher levels of anti-MSP3b IgG4 antibodies . This relationship was also found when antibodies to DG210 and MSP3CT were tested in similar conditions. The presence of intestinal helminths was found associated with lower levels of anti-DG210 IgG1  and higher levels of anti-DG210 IgG4 . When intestinal helminths were present, lower levels of anti-MSP3CT IgG1 , lower levels of anti-MSP3CT IgG3  and higher levels of anti-MSP3CT IgG4  were detected. Of note the changes in IgG4 responses were highly significant. The pattern of immune responses identified in helminth positive children, as compared to helminth negative children, is coherent with an increased Thelper 2 type of immune response associated with helminthic infection.However, in multivariate analysis, the presence of intestinal helminths was found to be associated with significantly lower levels of anti-MSP3b IgG1  worm carriage, ii) a decrease in cytophilic IgG1 and IgG3 antimalarial response together with an increase in non cytophilic IgG4, and iii) an increased occurrence of malarial attacks.Results obtained in Casamance confirm on the one hand, the association with protection of anti-MSP3 cytophilic responses th antigen, AMA-1 which is considered to act by antibodies inhibiting merozoite invasion. However, as has been observed in other settings, the association with MSP3 responses was the strongest and ultimately proved to be the only one remaining significantly associated with protection in multivariate analysis. This was confirmed using 3 different constructs derived from MSP3. As observed previously, the IgG3 isotype of MSP3 antibodies was associated with clinical protection Out of the different antigens studied, the association between clinical protection and cytophilic responses was found in univariate analysis for 3 of them, MSP3, Glurp and Serp/Sera, three molecules known to trigger the Monocyte-dependant mechanism, though not for the 4in vivo in exposed individuals. They also, once more, stress the potential relevance of antibodies to MSP3, as compared to other ADCI targets, and other vaccine candidates, a finding consistent with previous studies in other African and Asian settings In multivariate analysis, helminthic infection was found associated with an imbalance in favor of non-cytophilic antibodies directed to the molecules mediating ADCI, but not to that mediating merozoite invasion-inhibition. Here as well, and in agreement with the association with protection, the worm-induced imbalance was significant in multivariate analysis only for anti-MSP3 antibodies. In this case, both the cytophilic IgG1 and IgG3 responses were decreased, whereas the non-cytophilic IgG4, which can compete with the former two was significantly increased, relative to IgG1 and IgG3 levels. This was also consistent using 3 MSP3 constructs. These results indirectly support the importance of a Fcγ trigerred monocyte effect Among possible confounding factors, environmental factors can be likely excluded as in most families, there were both helminth free and helminth-infected children suggesting a similar exposure in the household and immediate surroundings. Conversely, the present observational study does not allow to formally conclude to a direct causal relationship between hookworm carriage, imbalance towards non-cytophilic antimalarial response and occurrence of malarial attacks, and does not exclude that genetic factors, for instance, could also contribute to either one of those 3 parameters.However, the hypothesis of a causal relation is supported by observations made in other fields than malaria. The influence of worms upon non-malaria specific immune responses in human is actually well documented P. yoelii strain becomes lethal), alteration of innate responses, and a profound decrease in Th1 responses P.falciparum extract in wormy individuals, leading to a contra-inflammatory profile of responses. These results confirm in humans our findings in rodents Findings in rodents, in which similar quantitative and qualitative immune responses changes were detected, are convergent. Higher parasite loads were reported in helminth-infected animals Our observations that intestinal worms can affect the quantity and quality of antimalaria immune responses have implications for malaria control  At least, the detection of every category of helminths, including a quantification of worms loads, should now be part of the biological investigations performed during malaria surveys or phase IIb vaccine trials and be included in the final bio-statistical model aimed at interpreting results.We have recently reported, in the village of Dielmo Even if this study substantiates the immunological role of worms in the isotype imbalance seen in the majority of children, it is likely that it is not the only influence. Several other factors may potentially contribute to the outcome; for instance, children are more prone to produce, early in life, IgM non-cytophilic classes of antibodies in contrast to adults who produce both IgG2 and IgM non-cytophilic responses. It is also plausible that there are genetic factors at the origin of a preferential switch towards a particular class or subclass of antibody. A detailed analysis of the underlying factors would require large cohort studies as there will be a need to combine an analysis of the genetic background of human populations, of parasite populations, and of the almost unlimited combinations of concomitant infections by parasites, viruses or bacteria, that are highly prevalent under tropical conditions P.falciparum/human immune interaction and for the very long delay required to acquire protective immunity against malaria related to a dominance of non-cytophilic responses, the latter being at least in part now attributable to worm carriageOur study and the underlying hypothesis bring new elements contributing to improving our understanding of the dynamics of


Rapid and correct diagnosis of malaria is considered an important strategy in the control of the disease. However, it remains to be determined how well these tests can perform in those who harbour the parasite, but are asymptomatic, so that rapid diagnostic tests (RDTs) could be used in rapid mass surveillance in malaria control programmes.Plasmodium species. In addition, the performance characteristics of the test were determined using microscopy as gold standard.Microscopic and immunochromatographic diagnosis of malaria were performed on blood samples from the hyperendemic Mount Cameroon region. Thin and thick blood films were stained with Giemsa and examined under light microscopy for malaria parasites. The RDT was performed on the blood samples for the detection of Plasmodium falciparum was the predominant Plasmodium species detected in the study population either by microscopy or by the RDT. Overall, the test recorded a sensitivity and specificity of 85.33% and 95.05% respectively, and an accuracy of 91.40%. The sensitivity and specificity of the RDT increased as parasite densities increased.Results revealed 40.32% to be positive for microscopy and 34.41% to be positive for the RDT. Parasites were detected in a greater proportion of samples as the parasite density increase. The Hexagon Malaria Combi™ test showed a high sensitivity and specificity in diagnosing malaria in asymptomatic subjects and so could be suitable for use in mass surveillance programmes for the management and control of malaria. Plasmodium species and the symptomless carrier status at any given time in children ranges from 60–100% [Plasmodium falciparum being the predominant Plasmodium species, although it co-exists with the non-falciparum species Plasmodium malariae, and Plasmodium ovale.The cosmopolitan populations of some areas of the Mt. Cameroon region are often infected with  60–100% , thrombo 60–100%  and a ne 60–100% ,1. The MGenerally in Cameroon, the best method of malaria control adopted by the National Malaria Control Programme is the early detection of parasites in symptomatic people followed by treatment with artemisinin combination therapies (ACTs) to curb the disease burden. Therefore, the malaria control programme in Cameroon usually leaves out individuals chronically infected with malaria parasites, but showing no symptoms.Plasmodium falciparum, can lead to a rapid deterioration in the state of the patient, it is important to find ways of diagnosing the disease in the shortest possible time.The detection of malaria parasites by light microscopy of Giemsa-stained thick and thin blood films remains the standard laboratory method for the diagnosis of malaria  in many Plasmodium lactate dehydrogenase (pLDH) have been used for several years and have been well received with RDTs showing high sensitivities and specificities compared to microscopy or polymerase chain reaction (PCR) [P. falciparum and cannot detect mixed infections.Rapid diagnostic tests (RDTs) have been used since the early 1990's for the detection of malaria parasites in humans. Several studies focusing on the detection of histidine rich protein-2 (HRP-2) and on (PCR) -18. NoneRDTS have been primarily evaluated for their performance in well-controlled research settings using sick people as subjects with either microscopy or PCR as gold standard. It remains to be determined how well these tests can perform in those who harbour the parasite but are not sick. Such that, these tests could be recommended for rapid mass surveillance in malaria control programmes so as to reduce the transmission and consequently burden of the disease. In order for any control programme to be effective in reducing disease prevalence, it should be both preventive and curative.P. falciparum only, non-falciparum species and mixed infections with P. falciparum in a population that was primarily asymptomatic for malaria in the hyperendemic Mt Cameroon region of South West, Cameroon.One of the RDTs that were recently introduced into the market is the Hexagon Malaria Combi™ and so far its performance characteristics and usefulness have not been evaluated in the Mt Cameroon Region. The main objective of this study was therefore to evaluate the performance characteristics and usefulness of Hexagon Malaria Combi™ test in detecting Samples were collected from school children at six different localities  in the in the Mt Cameroon region. This region stretches from the Atlantic Ocean at the Golf of Guinea and culminates at an altitude of 4,100 m above sea level. The equatorial climate has been modified by the double influence of the ocean and mountain. Temperatures range from 18°C to 35°C. Rainfall ranges from 1,500 mm–11,100 mm. This region has a mean humidity of 88%. About 1.3 million people live around the Mount Cameroon region. The population is cosmopolitan, made up of indigenous Bakweri people and immigrants from other areas around the country and foreigners (mostly Nigerians) attracted by the rich volcanic soil for agricultural processes and other job opportunities. The terrain is fairly hilly with many streams. At lower altitudes the extensive plantations of bananas, palms, rubber and the creation of agglomeration have totally replaced the primary forest.Ethical clearances for the study were obtained from the South West Provincial Delegations of Public Health and Basic Education. The ethics committee of the Tropical Medicine Research Station, Kumba, South-West Province, Cameroon, approved the study. All pupils found to be infected were treated with artesunate/amodiaquine.Sensitization visits were made by members of the research team to the localities to explain the objectives and advantages of the study to the teachers and the pupils. The study was carried out between March to June 2006. Children aged 4–16 years of both sexes in the six different localities were enrolled into the study. Informed consent was obtained from the parent/legal guardian of each child. The team of investigators moved to each of the six sites once every month for sample collection. Each child was observed for any clinical signs of malaria and his/her axillary was temperature recorded. Two mL of venous blood was collected from each consenting individual and used to prepare thick and thin blood films. The remaining blood was put into EDTA tubes, labeled accordingly, transported on ice blocks to the laboratory and stored at -20°C.Plasmodium species were detected with the help of bench aids for malaria diagnosis [The thick and thin blood films were stained with 10% Giemsa stain and read at ×100 (oil immersion objective) of the microscope. The iagnosis . Slides iagnosis . ParasitSlides were considered negative if no parasites were seen after observing 100 high-powered fields. The microscopist was blinded to the result of the rapid test.Plasmodium lactate dehydrogenase) released from Plasmodium species and is intended for the diagnosis of malaria. The test is designed to detect malaria caused by P. falciparum and the non-falciparum species . The expected species sensitivities are 90% for P. falciparum, 89% for P. m vivax while the specificity of the test is 99.5%.Briefly, the Hexagon Malaria Combi™ test is a rapid diagnostic test for the qualitative detection of pLDH  of diluent was dispensed into the conjugate wells and four drops into the washing wells. 20 μl of whole blood was added into each of the conjugate wells using disposable pipettes for each sample, mixed gently and allowed for one minute. Test strips were immersed into the conjugate wells and allowed for five minutes. Test strips were then transferred into the washing wells and allowed for 10–15 minutes for the clearance of haemoglobin. The results were then read using the manufacturers' guide. The standard reading time was 21 minutes.Once all the samples had been tested, sensitivity, specificity, accuracy and reliability of the Hexagon Malaria Combi™ test were estimated using microscopy as gold standard. The variables measured were the number of true positives (TP), number of true negatives (TN), number of false positives (FP), and the number of false negatives (FN).Sensitivity was calculated as TP/(TP+FN), specificity was calculated as TN/(TN+FP), the positive predictive value (PPV) was calculated as TP/(TP+FP), and the negative predictive value (NPV) was calculated as TN/(FN+TN) . Test acA total of 186 school children were recruited for the study. Of this 98 (52.69%) were females while 88 (47.31%) were males. The majority (52.69%) of the children belonged to the age group 9–12 years while the least (21.51%) belonged to the 13–16 years age group  were positive for non-falciparum species while 25 (33.33%) were positive for a combination of both P. falciparum and non-falciparum species.Of the 186 blood samples collected, 75 (40.32%) were found to be positive for malaria by microscopy. Of these, 40 (53.33%) of these were positive for P. falciparum, eight (12.50%) were positive for non- falciparum species and 21 (32.81%) were positive for P. falciparum and non-falciparum species  of the 186 blood samples were positive for malaria. 35 (54.69%) of these were positive for , eight 1.50% wereP. falciparum, non-falciparum species and mixed infections both microscopically and by using the rapid test. Plasmodium falciparum was the most predominant Plasmodium species detected either microscopically (86.67%) or using the rapid test (87.50%) in the study population  and specific (98.00%) for detecting ns Table .The recent development of rapid and accurate tests for the detection of malaria is highly commendable. One of the major goals of developing such rapid tests was that these rapid tests should be handled with ease and accurately by relatively unskilled staff for prompt diagnosis of malaria especially in areas where microscopy may not be available or available and pose problems because of its limitations ,6 especiThe strength of the present study is that it was performed in a population that was primarily asymptomatic in the hyperendemic Mt Cameroon region using microscopy as gold standard. Although asymptomatic malaria may be beneficial in inducing and sustaining partial immunity against malaria, it may give rise to severe disease complicated by coma, acidosis or severe anaemia . Thus toCurrently, the management of malaria by the National Malaria Control Programme in Cameroon is based on the detection of parasites in sick people followed by treatment with ACTs. This still places the population at risk because the majority of individuals infected without symptoms serve as a source of continuous malaria transmission in an area. In order for this control programme to be successful in curbing the disease burden, there should be no time waiting for people to fall sick before treatment, rather mass surveillance using rapid diagnostic tests to identify non-sick people and to map out areas which are potentially at risk would better reduce the burden of the disease.This study compared the diagnosis of malaria parasites by Hexagon Malaria Combi™ rapid test and traditional microscopy and found that the two methods yielded comparable results. A total of 186 asymptomatic children were tested; blood films identified 75 (40.32%) of these as positives while the rapid test identified 64 (34.41%). The fact that some malaria infections detected by blood films were not rapid test positive may be explained by the fact that Hexagon Malaria Combi™ rapid test detects parasite lactase dehydrogenase (pLDH), produced by life parasites only. Probably some of these children might have taken chemo-prophylaxis at home for malaria prevention. The possible explanations for discrepancies in test results obtained by microscopy and rapid test includes (i) insufficient detection of low parasitaemia by the rapid test (ii) Hexagon Malaria Combi™ rapid test detects only live parasites producing pLDH (iii) sequestration of parasites and (iv) false-positive reactions. Thus the five samples falsely positive by the rapid test could have been due to reasons (ii) and (iv) above while the 11 samples falsely negative for the rapid test could have been due to reason (i) above and probably the samples contained pLDH below the detection limit (20 ng/mL pLDH) of the rapid test.Plasmodium (P. vivax not found in this area) present in the Mt. Cameroon region with good sensitivities and specificities at different parasite densities, thereby giving this tool a greater potential for use as an epidemiological tool for the control of malaria. Also, this test performed better than the OptiMAL™ rapid malaria test, which failed to detect asymptomatic malaria in an area of Thailand endemic for P. falciparum and P. vivax [When compared to other commercially available rapid tests , the Hexagon Malaria Combi™ rapid test is highly recommendable because it can detect all the species of P. vivax . HoweverUnlike microscopy, which requires substantial training and experience, the Hexagon Malaria Combi™ rapid rest was performed well with minimal training. In addition, the rapid test can be performed in areas where electricity or microscopes are not available and the results can be obtained rapidly.Plasmodium falciparum kills within a short period. Adults or the parents/legal guardians of children may be ignorant of the symptoms of malaria and may associate these symptoms to witchcraft or other superstitious beliefs.There are compelling reasons to justify the implementation of rapid tests for malaria diagnosis for asymptomatic carriers. First of all, microscopy when used for active malaria surveillance may not be accurate as active surveillance itself is a highly labour intensive activity that may provide minimal returns . SecondlHowever, savings in technician time, including time to prepare stained blood smears and microscopically examination of the slides, the lack of the need to purchase and maintain expensive equipment, and the ability to work in remote areas where electricity and other infrastructure is lacking, make the use of antigen detection test methods tremendously appealing.The test has considerable potential for use in the management of malaria, provided it is made affordable. The cost of the test will clearly influence the extent to which it is deployed in any setting. To ensure that this useful tool can be made available to those in greatest need, the cost of the rapid test should not exceed U.S.$0.40/sample . UnfortuPlasmodium co-exist. The combination of its high sensitivity and specificity for P. falciparum, non-falciparum species and mixed infections at the different parasite densities suggests that the majority of malaria cases will be detected using this rapid test.In conclusion, the results obtained from this study demonstrate that the Hexagon Malaria Combi™ rapid test is suitable for use in mass surveillance programmes for the management and control of malaria in Cameroon, particularly in areas where the different species of HRP-2: histidine rich protein 2; PCR: polymerase chain reaction; RDT: rapid diagnostic test; FN: false negative; TN: rue negative; FP: false positive; TP: true positive.EEJE: Data collection, analysis, interpretation and, preparation of manuscript. JLN: Data collection and reading of manuscript for important intellectual content. NT: Contributed in data analysis and reading manuscript for important intellectual content. HKK: Coordination and critically reading of manuscript for important intellectual content. SW: Conception of study and experimental design, provision of finances for the study, coordination, interpretation of data and critically reading the manuscript for intellectual content. All authors read and approved the manuscript.