This study examined the prevalence of dhfr and dhps polymorphisms in P. falciparum found in pregnant women of Nchelenge district.Sulphadoxine-pyrimethamine (SP) is the recommended drug for intermittent preventive treatment in pregnancy (IPTp) in most African countries, including Zambia. However, malaria is still one of the leading causes of morbidity and mortality in pregnant women despite reports of greater than 50% of women taking at least two doses of SP in IPTp. Studies have shown that resistance to SP is associated with mutations in the This cross-sectional study was conducted in 2013 in Nchelenge, a holoendemic area with malaria prevalence estimated at 50% throughout the year. Three rural health centres were randomly selected and a census survey carried out at each health centre. A questionnaire was administered and malaria testing done using RDT and microscopy, with collection of a dried blood spot. A chelex extraction was done to extract parasite DNA from dried blood spots followed by nested PCR and enzyme restriction digestion.dhfr triple (Asn-108 + Arg-59 + Ile-59) mutant (68%) and dhps double (Gly -437 + Glu-540) mutant (21%). The quintuple haplotype was found in 17% with 2 samples with an additional Gly-581mutation. In addition 6% mutations at Val-16 were found and none found at Thr-108 respectively, these both confer resistance to cycloguanil. Multivariate analysis showed that there was an association between malaria and women aged 30-34 years old p < 0.05(AOR: 0.36) at 95% CI.Of the enrolled participants (n = 375), the median age of the women was 23. The prevalence of malaria by PCR was 22%. The PCR positive samples examined (n = 72) showed a high prevalence of dhfr and dhps genes. The high malaria endemicity in the general population of this area may have contributed to the high prevalence of resistant parasites in pregnant women, suggesting a need to examine the efficacy of SP given that it is the only approved drug for IPTp in Zambia.This study showed a high number of mutations in the  Plasmodium falciparum malaria in sub-Saharan Africa annually [Malaria remains a major cause of death predominantly in sub-Saharan Africa. Statistics by the World Health Organization attribute an estimated 627,000 deaths to this disease in 2012 alone [annually -5. Epideannually .Zambia has achieved over 50% reduction in malaria prevalence in the past decade . HoweverIn Zambia, IPTp was adopted in 2003. It is administered presumptively three months after quickening 16-26 weeks) -26 weeks. Prior tdhfr) and dihydpteroate synthetase (dhps) genes. Mutations in dhfr confer resistance to pyrimethamine while mutations in dhps confer resistance to sulphadoxine and other sulpha drugs. There are variations in SP mutations, single, double or triple - the more mutations, the stronger the resistance. The dhfr triple mutant (Asn-108 + Ile-51 + Arg-59) and dhps double mutant (Gly-437 + Glu-540) have been strongly associated with potential resistance in sub-Saharan Africa [Drug resistance is one of the greatest challenges of malaria control programmes. Hence, while SP remains ideal for IPTp, in stable transmission areas, increasing resistance has been reported ,22. SP rn Africa ,24.dhfr frequency ranging from 71-92% and 39-71% frequency for the double mutant dhps, respectively [A study done in 2006 in six districts in the general population of Zambia showed variation of rates of resistance to SP with mutated ectively . TherefoThis cross-sectional study was conducted in Nchelenge district, Luapula province, Zambia in February to April 2013. The population of Nchelenge is estimated at 180,000 and the inhabitants are mostly fishermen and peasant farmers. Nchelenge has one first level referral hospital St Paul’s Mission hospital, ten rural health centres and three health posts. This is a holoendemic area of malaria transmission with an estimated 50% confirmed cases throughout the year, hence transmission is throughout the year. An estimated 80% of recorded deaths are attributed to malaria with the most affected being children under five years of age and pregnant women. The target population of this study was pregnant women who were residents of Nchelenge and attended antenatal care at any of the three health facilities selected for the study  out of 14 in the district and these three were the sites chosen for the study. The rural health centres selected were Kabuta, Nchelenge and Kashikishi RHC. Consenting individuals were included in a census survey from the rural health centres. Hence, all pregnant women who came to the centre for antenatal care, regardless of whether for re-visits or first attendance, were eligible to be included in the study.The prevalence of malaria in pregnant women was assumed to be 50%, which is the prevalence for the general population in Nchelenge. The calculated total sample size was 384 with a power of 80 at 95% CI.The data collection tool used in this study was a structured questionnaire to document demographic data and risk factors for the presence of malaria. To validate the data collection tool pre-testing of the questionnaire was done in two ways: the tool was pre-tested to ensure clarity of questions in English and then for consistency, precision, clarity of questions and correct translations in Bemba. Necessary adjustments and revisions were made based on the pre-test results.P. falciparum (RDT and/or smear and/or PCR). Approximately 100 microlitres of venous blood was collected from each participant by a single finger prick. A rapid diagnostic test (RDT) and microscopy was used to test for malaria. The RDT used detects the antigen histidine rich protein II (HRP II) found on the parasite surface, the test kits brand was the Standard Diagnostics Bioline Malaria antigen Pf test (SD-p.fTM) manufactured by Standard Diagnostics Inc., South Korea. The test results were ready within 15 minutes. Thick blood smears were collected on microscope slides for later reading by a microscopist.The eligible pregnant women were screened for malaria and anaemia on the scheduled ante-natal care day. Malaria was defined as a positive laboratory test for Plasmodium falciparum DNA was extracted from the DBS by a chelex protocol and amplified by polymerase chain reaction (PCR) followed by restriction enzyme digestion. Mutations were defined as the detection of polymorphisms in the dhfr gene specifically at Asn-108, Arg-59, Ile-59 and dhps Gly-437, Glu-540 and Gly-581 mutation. Mixed infections were those that showed both sensitive and resistant parasite strains while sensitive strains were parasites that showed no polymorphisms in the region of interest after restriction digestion. Quality assurance was done at Tulane University, School of Public Health and Tropical Medicine in the Kumar Laboratory. Positive samples (n = 10) were randomly selected and PCR was done followed by restriction digestion with enzymes BStNI, AluI and BSrI. The results were similar to those initially obtained hence results were reproducible and reliable.Dried blood spots (DBS) were also collected on Whatman 3MM filter paper at the same time as the RDT and thick smear. Analysis was done using Stata version 12.2. Variables were defined, descriptive statistics; means, medians, standard deviations and odds ratios were used to assess the data. The factors examined were age, marital status, gravidity, education level, ITN ownership and usage, malaria knowledge, medical history and SP intake. These were self reported and SP intake was confirmed by medical records. Chi square was used to determine the association between malaria and each of the associated factors. Significance  was set at 5% with a confidence interval at 95%. Multivariate analysis using logistic regression was done for adjustments, to control for confounding and to examine the odds ratios for malaria prevalence in relation to each of the risk factors.Ethical approval was sought from the University of Zambia Biomedical Research Ethics Committee (UNZABREC) and granted in February, 2013 (IRB004-11-12).A total number of 400 participants were enrolled in this study, of these 375 had complete data. The median age of the participants was 23 years old (IQR 20-24). The median gestational stage was 24-36 weeks. Of the women enrolled 57% reported to have had a primary education while only 3.2% reported a college/university attainment. Of these women 15% had not received any SP at the time of enrollment. The major source of non-participation was refusal (<5%) and incomplete data.The prevalence of malaria by rapid diagnostic test was 30%, by microscopy prevalence was 15% and 22% by PCR. The rapid diagnostic test is an antigen-based test and may remain positive two weeks after treatment, hence may result in false positives. Using Chi square analysis, significant results were obtained for malaria symptom knowledge and age. Over 75% of women with symptom knowledge of malaria had taken medication in the last malaria infection, with the greatest proportion (50%) having taken artemether-lumefantrine (Coartem®). In terms of insecticide-treated net distribution, 74% of the women reported to own at least one net in their household although the results suggest that there was no association with malaria. Using multivariate analysis none of the factors were significantly associated with malaria except age. Women aged 30-34 were 64% less likely to have malaria compared to those in the reference group 14-18 years old (AOR: 0.36 95% CI).dhfr and dhps mutations are shown in Figures dhfr triple mutant (Asn-108 + Ile-51 + Arg-59) was 68% and dhps double mutant (Gly-437 + Glu-540) was 21%. The quintuple mutant was present in 17% (n = 12) of the samples and the sextuple mutant was present in 3% (n = 2) of the samples. The figures show additional codons at Val-16 and dhps mutation at codon 436. There was no association between SP intake and genotypes present.Of the PCR positives, 72 were genotyped using restriction fragment length polymorphism (RFLP) for the presence of mutations at the different loci. The prevalence of This study documents high malaria prevalence and antifolate resistance mutations in pregnant women in a holo endemic area. It is important to note that >60% of women who were found with malaria had taken at least one dose of SP. However, with multivariate analysis there was no association between presence of malaria and any of the associated risk factors except age, hence the association found with chi square may be due to confounding. The multivariate analysis on predictors of malaria prevalence suggests an association between malaria prevalence and women aged 30-34 years compared to those women aged 14-18 years old. This may be due to a protective effect reported in other studies that older women who are likely to have had more than one previous pregnancy (multigravidae) are less likely to have malaria compared to those who have not had children previously (primigravidae) .Possible bias due to non-participation may have occurred as this study was based exclusively at the health centres thereby excluding those pregnant women who did not attend antenatal clinic and who may have a different profile from those that do. Another study limitation was that clinical efficacy of SP was not determined in order to assess treatment failure, and the data collected including when the last malaria medication was taken may be subject to recall bias.dhfr mutations indicating mostly pyrimethamine resistance but also a higher emergence of mutations Val-16 which is an indication of resistance to cycloguanil and a higher emergence of mutations at dhps 581 than previously reported. Another study conducted in Mansa, Zambia in pregnant women showed 63% had quintuple mutants while only two had an additional mutation at the Gly-581 loci. The treatment failure recorded in the study was 22%. This shows that the dhps mutations are becoming more common and there has been an escalation in the mutations in this population. This is consistent with other studies that have shown that in hyperendemic areas with high rate of transmission as is the case for Nchelenge, resistant parasites have a tendency to increase and contribute to treatment failure [dhfr and dhps mutants had escalated rapidly since 1988, and that the quintuple (dhfr triple + dhps double) mutant associated with highest levels of SP clinical failure was starting to be seen (6.5%) [The results obtained show high levels of  failure . SP has n (6.5%) .dhps triple mutant and suggested that IPTp with SP may not improve overall pregnancy outcome [dhfr and dhps mutations while still others have found that SP IPTp is effective [Studies done in Tanzania have shown an emergence of the presence of the  outcome . Furtherffective ,30.High prevalence of multiple mutants raises concerns as these have been shown to correlate with therapeutic or clinical failures. Therefore there is need to explore other drugs as options for IPTp.This study shows a high prevalence of malaria in pregnant women of Nchelenge district, despite reported use of SP-IPTp. This may be due to the high parasite resistance mutations to the drug as was evident in this study. The research implications are that there is need to conduct further high powered studies to determine efficacy and parasite clearance rates using SP-IPTp and also begin to explore alternative avenues for drugs to be used in IPTp if it is to be beneficial to this population.


Plasmodium infection, helminth carriage and the risk of anaemia.Anaemia in children under five years old is associated with poor health, growth and developmental outcomes. In Papua, Indonesia, where the burden of anaemia in infants is high, we conducted a community survey to assess the association between A cross sectional household survey was carried out between April and July 2013 in 16 villages in the District of Mimika using a multistage sampling procedure. A total of 629 children aged 1–59 months from 800 households were included in the study. Demographic, symptom and anthropometry data were recorded using a standardized questionnaire. Blood and stool samples were collected for examination.P. vivax parasitaemia and 3.9% (21) had P. falciparum; the majority of children with malaria were asymptomatic . Soil transmitted helminth (STH) infection was present in 43% (105/269) of children assessed; those with STH were at significantly greater risk of P. vivax parasitaemia compared to those without STH . Anaemia (Hb<10 g/dl) was present in 24.5% (122/497) of children and associated with P. vivax parasitaemia , P. falciparum parasitaemia , hookworm carriage , Plasmodium–helminth coinfection  and severe stunting .Of the 533 children with blood film examination, 8.8% (47) had P. vivax and P. falciparum infections and hookworm all contribute to risk of paediatric anaemia in coendemic areas and should be targeted with prevention and treatment programs. The relationship between helminth infections and the increased risk of P. vivax parasitaemia should be explored prospectively.Asymptomatic  Anaemia remains a critical determinant of global morbidity and mortality in children under five years old –3. WhilsP. falciparum endemic areas of Africa . , 9. 2, 9f Africa , 14–16, P. vivax .P. falciparum and P. vivax , p<0.001), fever in the last 24 hours , and anaemia  (Three risk factors for p<0.001) .Ascaris lumbricoides (AL), Trichuris trichiura (TT) and hookworm were present in 28%, 21% and 13% of the children respectively.Stool examination was undertaken in 269 (43%) children of whom 39% (105) had soil transmitted helminths (STHs); almost half of these children were infested with multiple STHs . The prevalence of helminthiasis was 19.4% 7/36) in infants compared to 40.8% (95/233) in children aged 1 to 5 years (Tables 6 in infaP. vivax parasitaemia with 15.3% (15/98) of children with STHs having parasitaemia recorded compared to 4.6% (8/174) of children without STH . The risks of P. vivax infections remained higher compared to those without STH after stratifying by species of STH: Ascaris lumbricoides, Trichuris trichiura and hookworm with the OR of 3.7 , 3.8  and 3.5  respectively.Overall children with STHs were at significantly greater risk of parasitaemia  although the trend remained in multivariable analysis it was no longer significant . The difData on haemoglobin concentration were available in 60 (55.5%) infants (mean Hb 10.9 g/dl [95%CI 10.6–11.3]) and 437 (83.8%) young children (mean Hb 11.2 g/dl [95%CI 10.1–11.4]); Of all the nutritional status indices, only severe stunting was associated with an increased risk of anaemia . The prevalence of anaemia was 34.1% (28/82) in children with severe stunting and 21.1% (65/308) in those with normal height according to age; p = 0.02.Plasmodium and STHs occurred in 7.1% (19/269) of the children with available data. Compared to monoinfection the presence of both Plasmodium and STH was associated with a greater risk of anaemia  and severe anaemia .Falciparum and vivax malaria were both significant risk factors for anaemia in children under five with ORs of 16.3  and 4.7  respectively, p<0.001; Plasmodium infection and the importance of soil transmitted helminths. Plasmodium infection was present in 12% of children under 5 years old, most of whom were asymptomatic, with P. vivax the dominant species of infection. Helminth carriage was common and when present with malaria doubled the risk of anaemia.In Papua, Indonesia, anaemia exerts a significant morbidity particularly in young children. Previous hospital based studies have highlighted the importance of undetected, untreated or inadequately treated malaria as a key contributor to reduction in haemoglobin concentrations, , 9. The Plasmodium infection and may also play a role in P. vivax infections [The aetiology of anaemia from malaria and helminth carriage is well described and is attributable to a combination of chronic blood loss, haemolysis and haemopoietic suppression. Associated nutritional deficiency is also likely to play a key role. Although in our study general measures of stunting and wasting were not significant risk factors, in the subset of individuals tested, microcytosis was present in almost 75%, suggesting a potential contribution of iron deficiency. Recent studies have demonstrated that increased hepcidin level and associated impaired intestinal iron absorption may play an important role in the development of anaemia in children with asymptomatic fections .P. falciparum and P. vivax infections [P. vivax is particularly apparent in children under 5 years old, with the prevalence (8.8%) more than twice that of P. falciparum (3.9%). Plasmodium vivax peripheral parasitaemias were lower than in P. falciparum infections and children with P. vivax more likely to be asymptomatic. However the relapsing nature of P. vivax infections is an important determinant of anaemia in the first year of life [The high proportion of asymptomatic malaria in young children in this region suggests that clinical immunity develops at an early age in both fections , 31, alt of life . In addi of life .Soil transmitted helminths (STHs) were present in almost half of the children by 5 years of age, and of these, almost half had multiple infestations. Stool sampling was undertaken in less than half of the children. The characteristics of those with and without stool samples  was similar except for ethnicity, with 60% of Papuans were without stool sampling compared to 51% in non Papuan children. Papuan ethnicity was a major independent risk factor for helminthiasis raising the prevalence 2 to 4 fold, hence our analysis is likely to have underestimated the true burden of helminth infection.The presence of hookworm was particularly important, increasing the risk of anaemia 2.6 fold. Once again the true burden is likely to be actually higher since Kato Katz has low sensitivity in diagnosing hookworm eggs . HookworP. falciparum and hookworm comorbidity in preschool children was associated with lower haemoglobin concentrations compared to those having single infection [P. falciparum infections and hookworm infestation had lower risk of developing iron deficiency anaemia [In Papua, children with malaria and STH co-infection were four times as likely to have anaemia compared to those having mono-infection with either pathogen, and at almost nine fold greater risk of severe anaemia. Although the frequency of co-infection was relatively low (7.1%), the presence of co-infection and its association with anaemia is likely to have a significant detrimental effect on the child’s health and development . Similarnfection . On the  anaemia . AlthougPlasmodium spp. in a complex way, increasing susceptibility to falciparum malaria without resulting in severe symptoms [P. vivax infection may be different. Heavy ascariasis infestation reduced the risk of symptomatic P. vivax malaria in one study of children aged 2–14 years old [P. vivax since most have been conducted in sub-Saharan Africa where P. vivax is not endemic [Dysregulation of cellular immune responses in STH infection –40 may dsymptoms , 41, 42.ears old . In anotears old . Studies endemic –49.Plasmodium infection and STHs to the development of anaemia could not be examined in this cross sectional study. However in this population, individuals with a history of fever in the previous 30 days is a useful indicator of symptomatic malaria and in the cross sectional surveys was associated with a 3.6 fold higher risk of anaemia, suggesting a likely contribution of recurrent malaria. Secondly, iron status was not quantified so the contribution of iron deficiency could not be addressed. However in the subset of patients in whom data were available, three quarters of children with anaemia had microcytic erythrocytes suggesting iron deficiency and/or infections in the underlying aetiology.The sample size of our study was designed to evaluate the impact of antimalarial treatment policy by comparison with a prior survey conducted in 20015. The extensive data collected in the second survey enabled us to define the effect of asymptomatic malaria and STH infections to anaemia in children under-fives. The sample size of this study was sufficient to test the association of malaria, STH and anaemia . HoweverPlasmodium infection and malaria/helminth coinfection are important risk factors for anaemia in young children. STHs appear to increase a child’s risk of P. vivax parasitaemia. Operational studies are needed to define the effectiveness of combined malaria and STH control programs to prevent their combined contribution to anaemia in early life and associated morbidity, mortality and cognitive impairment. Our study suggests that deworming has potential to reduce P. vivax infections. In the case of malaria, the risks and benefits of malaria screening or preventive treatment in infancy in areas endemic for P. falciparum and P. vivax warrants careful examination [In conclusion, asymptomatic mination .S1 Dataset(XLSX)Click here for additional data file.


P. falciparum, S. mansoni and soil transmitted helminths (STH) infections and their correlation with schoolchildren’s anemia in the Democratic Republic of Congo (DRC), we collect these data.Anaemia reduces cognitive potential in school children, retards their growth and predisposes them to other diseases. As there is a paucity of data on the current burden of S. mansoni and STH infections were assessed using kato-katz technique. Malaria infection and hemoglobin concentration were assessed using Blood smear and Hemocontrol device, respectively.This study reports baseline data collected from a randomized controlled trial investigating the impact of IPT with SP and SP-PQ on anemia and malaria morbidity in Congolese schoolchildren . Plasmodium spp. infection was 18.5% (95%CI:15.6–21.9). Amongst those infected, 24 (21%), 40 (35.1%), 40 (35.1%), 10 (8.8%), had light, moderate, heavy, very high malaria parasite density, respectively. Above 9 years of age (p = 0.02), male and history of fever (p = 0.04) were both associated with malaria infection. The overall prevalence of S. mansoni infection was 6.4% (95%CI:4.4–9.1). Girls were associated with S. mansoni infection (p = 0.04). T. trichiura was the most prevalent STH infection (26.3%), followed by A. lumbricoides (20.1%). Co-infection with malaria-S. mansoni and malaria-STH was, respectively, 1.5% (CI95%:0.7–3.3) and 6.4% (CI95% 4.4–9.1). The prevalence of anemia was found to be 41.6% (95%CI:37.7–45.6) and anemia was strongly related with Plasmodium ssp infection  and S. mansoni infection .A total of 616 primary schoolchildren from 4 to 13 years old were enrolled in the study. The prevalence of S. mansoni infection were strongly associated with high prevalence of anemia in schoolchildren. Therefore, specific school-based interventions, such as intermittent preventive treatment or prophylaxis, LLITN distribution, anthelminthic mass treatment and micronutrient supplementation are needed to improve school children’s health.Malaria and  P.falciparum, schistosomiasis and STH infections have been reported from various epidemiological settings in Africa Malaria, Schistosomiasis and Soil Transmitted Helminth (STH) infections compose a considerable disease burden in schoolchildren in developing countries. These parasitic infections show a similar geographic distribution and polyparasitism of P.falciparum triggers the development of acquired immunity that controls blood-stage parasitaemia, thereby reducing clinical symptoms and life-threatening complications in older children and adults Plasmodium infections, if untreated, persist and maintain malaria-induced inflammation that is commonly associated with iron deficiency anaemia (IDA) due to impaired intestinal iron absorption, impaired iron release from hepatocytes, and impairment of the recycling of iron derived from phagocytosis of parasitized red blood cells In endemic malaria areas, the prolonged carriage of P. falciparum, Schistosomiasis and/or STH infection on childhood anemia has been reported Findings on the interaction of malaria and helminth infections on the health of children are controversial. Some studies have shown a protective effect of helminths on symptomatic malaria. Severe worm burden seems to suppress malaria symptoms The Democratic Republic of Congo (DRC) is at present one of the most affected countries by malaria P. falciparum, Schistosomiasis and STH infections, or their relationship with anemia in primary schoolchildren. The aim of this study was therefore to determine the prevalence of asymptomatic malaria, Schistosomiasis and STH infections in apparently healthy children attending schools in a semirural area in Kinshasa, DRC, as well as their relationship with anemia.While a hand full of studies are dedicated to the groups most vulnerable for malaria such as children under the age of five and pregnant women, very little is known about the ongoing burden of asymptomatic Kinshasa is one of the biggest cities in Africa and divided into 25 administrative zones or municipalities that cover about 9,965 km2. According to the national health policy, 25 Health Zones (HZs) roughly cover the 25 administrative zones. These HZs are also divided into Health Areas (HAs). Kimbanseke is the most densely populated peripheral municipality of Kinshasa housing 946, 372 inhabitants in 2004 This study analyses cross-sectional, baseline data from an open label, randomised, controlled trial enrolling asymptomatic school children and investigating the impact of intermittent preventive treatment (IPT) with Sulfadoxine-Pyrimethamine (SP) and SP plus Piperaquine (PQ) on anemia and malaria morbidity in Congolese schoolchildren . The trial design and protocol are described in detail elsewhere th to the 24th November 2012. Children whose parents did not provide written informed consent or presenting fever, muscle aches or other symptoms suggestive for malaria at the time of enrolment were excluded. Children participating in another clinical trial were also excluded from the study. A structured questionnaire was used to obtain information on age, river contact, history of fever, diarrhea and abdominal pain.Briefly, asymptomatic primary school children were recruited from the 10Plasmodium infection using microscopic examination for malaria parasites. Giemsa-stained thick blood smears (TBS) were used for microscopy. Blood slides were examined using light microscopy at 1000 × magnification. Hundred microscopic fields were examined in the thick smear before concluding that a blood slide was negative. All slides were read twice by experienced microscopists. If the discrepancy was greater than 15%, a third reader was used to confirm diagnosis. The parasite density per microliter of blood was calculated using the following formula: (Number of trophozoites x 8000)/Number of leucocytes. Parasite counts were utilized to classify the intensity of Plasmodium spp. infection into light, moderate, heavy, or very heavy infections respectively as followed: 1–499 parasites/µL, 500–1,999 parasites/µL, 2,000–9,999 parasites/µL and ≥10,000 parasites/µL.Finger-prick blood specimens were taken for analysis of S. mansoni and/or helminth eggs were counted and the number obtained was multiplied by the factor 24 in order to get the number of eggs per gram of feces (epg). Egg counts were utilized to classify infection intensities into light, moderate, or heavy infections respectively as followed: for S. mansoni 1–99 epg, 100–399 epg, ≥400 epg; A. lumbricoides, 1–4,999 epg, 5,000–49,999 epg and ≥50,000 epg; and for T. trichiura, 1–999 epg, 1,000–9,999 epg and ≥10,000 epg Fecal samples were collected and taken to the laboratory of Parasitology of the University of Kinshasa for analysis using the Kato-Katz technique Urine samples were analysed using the centrifugation method as described by Okanla Hemoglobin (Hb) levels were determined using a HemoControl device . Anemia was defined by a hemoglobin concentration <11, <11.5 and <12 g/dL, respectively for schoolchildren of less than 5 years, 5–11.9 years and 12–14.9 years old. Anemia was classified as severe anaemia Hb <7 g/dl, moderate anaemia Hb: 7–9.9 g/dl and mild anaemia Hb: 10–11.4 g/dl. Weight and height were measured in each child. With these measurements we calculate the following indicators using the WHO Anthroplus software: a) height-for-age Z-score (HAZ) to assess stunting in children 6 to 240 months old; b) weight-for-age Z-score (WAZ) to assess underweight in children of 6 to 120 months of age; and c) BMI-for-age Z-score (BAZ) to assess thinness in children of 6 to 60 months of age The sample size was estimated on the basis of the expected additional impact on anemia of the intervention arm versus the comparator The investigators agreed to conduct the present study in full agreement with the principles of the Declaration of Helsinki’ and subsequent relevant amendments. The study was approved by the Ethical Committees of the University of Antwerp, Belgium and of the School of Public Health, University of Kinshasa, DRCongo. Approval was also obtained from the Ministry of Health of DRCongo. Prior to the start of the project, special permission was obtained from the DRCongo Ministry of Education and the local health and education authorities of Biyela. A series of meetings were also held in the participating schools to explain the nature and purpose of the trial. Written informed consent (IC) was obtained from each parents or legal guardians of all children prior enrolment. An oral assent was obtained, as well, from children who were 12-years-old or older. All participants were administered albendazole  and praziquantel (40 mg/kg) against helminth infections. Children suffering from clinical malaria were treated according to national policy.95%:27.4–34.9), abdominal pain , diarrhea (15.3% CI95% 12.6– 18.4), and bloody feces (11.2% CI95% 8.9–14.0). Eleven percent of the children reported a history of blood transfusion . Two hundred seventy seven school children  were using the river as source of water to bathe or to wash their clothes.In total, 989 children aged between 4 and 13 years (median: 8 years and IQR:7.5–9.5 years) were assessed for eligibility in November 2012 of which 616 (62.3%) met the inclusion criteria and whose parents consented. A high frequency of refusal was found 324 (32.8%), with no real differences between schools in terms of percentage of parents refusing . Childre95%: 2.4–4.8) children presented fever with a mean temperature of 38.6 (SD 1.2). Blood smear, hemoglobin concentration and stool examination were performed, before they were treated according to the national policy. The prevalence of malaria parasitaemia amongst febrile children was 41.2% (CI95%:24.7–59.3). The median malaria parasite density was 9840/µL (CI95%:4660–14480). No S. mansoni infection was diagnosed in symptomatic children (95% :29.8–64.9)  .Plasmodium spp. infection was 18.5% (CI95%:15.6–21.9) and was found to be significantly lower than in symptomatic children. P. falciparum was the most prevalent species . P.malariae was diagnosed in two children. In those with parasitemia, the median parasite density was 1478/µl (IQR:557–5279) and was also significantly lower than 9840 parasites/µL found in symptomatic children (p = 0.01) , and girls  were less likely to have malaria infection. History of fever was an independent risk factor for malaria infection   .  . Schoolc = 0.03) . No signS. mansoni infection was 6.4% (CI95%:4.4–9.1) and the median parasite density was 48 eggs/g (IQR:24–76) and in girls . Other factors such as history of abdominal pain, diarrhea, bloody feces before enrolment did not demonstrate any association with S. mansoni infection . The liknfection .95%:28.6–37.4). Only two of the three majorSTH were found: Trichuris trichiura (26.3%) and Ascaris lumbricoides (20.1%). Hookworms were not found. However one case of a non-STH, Enterobius vermicularis was diagnosed. Girls  and younger children  had a significant lower risk for STH infection  and 6.4% (CI95% 4.4–9.1)  was found in 79 participants .Co-infection with malaria and a helminth was common though we did not observe any 4.4–9.1) . There wThe nutritional status of most children was within healthy parameters but a few cases of stunting (HAZ<−2) 0.16%, thinness (BAZ<−2) 0.65% and underweight (WAZ<2) 0.97% were observed .S.mansoni, and a 0.66 g/dl Hb lower when co-infected with STH (p = 0.02) than all not co-infected (The mean hemoglobin concentration was 11.6 g/dL (SD±1.4) and was found to be lower in children with asymptomatic malaria infection compared to uninfected children (p<0.001) . Hb concinfected . Other f95%:37.7–45.6) with no difference between sexes (p = 0.6). It was more common in P.falciparum-infected children (71.0%) than in uninfected children (38.8%) (p<0.001) . The multivariate regression model shows that anemia was associated with asymptomatic malaria infection  and S. mansoni infection  independently  . Of thesp<0.001) . In thisp<0.001) . No signendently . No inteP. falciparum in endemic areas is supposed to decrease significantly with age, because children would gradually developed some degree of immunity against the malaria parasite, as a result of repeated infections P. falciparum infection in endemic areas.In the Mokali Health Area, a semi-rural area of Kinshasa located in the Health Zone of Kimbanseke, the prevalence of asymptomatic malaria infection in schoolchildren was found to be 18.5%. Similar observations were made in 1981–1983 in Kinshasa, and 2000 in Kimbanseke T. trichiura having the highest prevalence, followed by A. lumbricoïdes (20.1%). These values are significantly lower than 90% and 83.3% respectively for A. lumbricoïdes and T. trichiura reported by Vandepitte in 1960 in Kinshasa A. lumbricoides infection, however improved sanitary, access to adequate water supply and access to health care should further decrease the prevalence of STH infections.Like malaria, STH were highly prevalent in the study population (32.8%). This could be the result of poor sanitary conditions in the Health Area of Mokali. This study recorded a prevalence of 26.2% for S. mansoni infection to be 6.4%. This prevalence is significantly lower compared to 89.3% reported in 2012 in Kasansa Health Zone, another endemic setting for S. mansoni in DRC S. mansoni infection. Similar observation was found in Yemeni in California This study also estimated the prevalence of P. falciparum and S. mansoni occurred at very low levels (1.5%). This is consistent with findings from Kenya in 2008–09 and Ethiopia 2008–09 and Uganda 2006 P. falciparum and STH co-infections were more frequent (6.4%). No association was found between malaria infection and S. mansoni infection neither between malaria infection and STH infection. This is in total agreement with previously reported data from Tanzania in 2010 Co-infection with S. mansoni infection was also found to be an independent risk factor for anemia in schoolchildren. No interaction was found between asymptomatic malaria infection and S. mansoni in regard to anaemia.On the other hand, the prevalence of anemia in primary schoolchildren was found to be 41.6%. This was lower than 67% observed in Kasansa, DRC in 2012 plasmodium spp carriage. Finally, no hookworms were found in Mokali’s schoolchildren. This could be at least partly explained by the fact that, the Kato-Katz slides were examined after 24 hours, which situation may lead to overclearance of hookworm eggs by glycerol.The study has a number of limitations. First, given the high rate of refusal (32.8%), which may lead to a selection bias, the reported data may not be representative of the schools surveyed. However, given that children whose parents did not consent were similar to those included in regard to age, sex and class, we have no reason to suspect that children in these two groups differed greatly in regard to other characteristics not assessed. This high proportion of refusal may indirectly suggest a negative perception of IPT or other malaria intervention in schoolchildren by the community. This underlines the urgent need to assess the perception and potential social and cultural barriers that can prevent an efficient implementation of malaria control strategies in schoolchildren. Second, asymptomatic malaria infection is mostly characterized by low grade parasitemia P. falciparum infection was highly prevalent in schoolchildren of Biyela Health Zone, and along with S. mansoni infection, they contribute to a great extent to the occurrence of anemia. These results highlight the important role of school-based interventions, which may include: deworming, micronutrients and intermittent preventive treatment for malaria for the control of anemia among African schoolchildren.This study demonstrated that


P. falciparum infection and its complications. This study investigated the prevalence and risk factors for P. falciparum infection and malaria among pregnant women reporting for first antenatal care (ANC) clinic visit in the mount Cameroon area.Pregnant women in malaria endemic areas are at high risk of Venous blood samples from consented pregnant women were screened for malaria parasitaemia by light microscopy. Haemoglobin levels, white blood cell (WBC) counts, lymphocyte counts and percentage were determined using an automated haematology analyser. Socio-demographic/economic data, environmental factors and use of malaria control measures were documented. Univariate and multivariate statistical analyses were used.P. falciparum parasitaemia. Malaria parasitaemia was significantly (P < 0.001) associated with febrile illness. The overall prevalence of malaria and asymptomatic infection was 16.0 % (95 % CI = 11-20 %) and 10.5 % (95 % CI = 7.3-15 %) respectively. A greater proportion of the malaria cases (61 %) reported at the clinic during unscheduled days meanwhile women with asymptomatic parasitaemia mostly (92.8 %) seek for ANC during scheduled clinic days. Lower lymphocyte percentage was significantly associated with increase parasite density  and febrile status . While age and gravidity were significant factors associated with P. falciparum infection and/or malaria, the presence of bush and/or standing water around human residence was an independent risk factor of P. falciparum parasitaemia  and malaria . Being unmarried was significantly associated with increase risk  of P. falciparum parasitaemia. Similarly, single women (938) had a significantly higher  geometric mean parasite density (GMPD) compared with married women (338).Sixty-eight  of the women enrolled were positive for P. falciparum infection and malaria. Education on early ANC attendance and environmental sanitation are important public health targets for malaria control in pregnancy in this setting.Marital status and human residence in areas with bushes and/or standing water modify risk of  P. falciparum infection and its complications which are severe and multiple [Pregnant women in malaria endemic areas are at high risk of multiple . These mmultiple . It is gmultiple . Howevermultiple –5. It ismultiple . In areamultiple . Treatmemultiple , 8.Longitudinal cohort studies investigating the clinical presentation of malaria parasite infection in pregnant women showed that during routine ANC (scheduled) visits, malaria is less common , 9, 10. P.falciparum infection may largely depend on the intensity and stability of malaria transmission in the local environment [Clinical manifestations of ironment . Other fironment , 10, levironment  and rainironment . Malariaironment , 14. Mosironment –15. MalaP. falciparum infection and malaria among pregnant women reporting for first ANC clinic visit during scheduled and unscheduled clinic days at some health facilities in the study area. Secondly, the study assessed demographic, socio-economic, environmental, haematological factors to identify risk factors associated with P. falciparum infection and malaria in pregnancy in this setting.Previous findings in the mount Cameroon area show that fever history is common among pregnant women and this usually correlates with the time of first clinic visit , 17. PreThis study was carried out in Mutengene and Muea Integrated Medical Centres located in the Mt. Cameroon Area, Fako Division, South West Region. These medical centres are government-owned institutions that offer antenatal care, preventive and curative services at affordable costs for the middle and low- income population. Both health facilities are highly accessible, facilitating utilisation of ANC services.P. falciparium accounts for up to 96 % of malaria parasite infections in this area [Anopheles gambiae s.l. (56.86 %), An. funestus s.l. (32.57 %), An. hancocki (9.38 %) and An. nili (1.18 %). All anopheline species are highly anthropophilic and exophagic (exceedingly for An. gambiae) with a human blood index (HBI) of 99.05 % [Anopheles gambiae is proportionately more abundant throughout the year and show peak of abundance towards the rainy season. Equally, the high survival rates of the malaria vectors  suggest a high vector potential for the species [An. gambiae s.s is defined as total annual precipitation of 230–9224 mm, maximum annual temperature range of 25–31 °C and minimum annual temperature range of 18–20 °C. An. funestus thrives in zones with a total annual precipitation of 230–2817 mm, maximum annual temperature of 25–29 °C and minimum annual temperature of 20–23 °C. The mean maximum annual relative humidity is 88 % [The Mt. Cameroon Area has an equatorial climate made up of a long rainy season which runs from March to October with maximum rainfall usually recorded in the months of August and September. The dry season runs from November to February. Malaria parasite transmission is perennial and his area . Malariahis area . Entomol 99.05 % . Anophel species . The ove species . In the  is 88 % .Mutengene is a semi-urban, road junction town located at about 220 m above sea level with a highly heterogonous population of approximately 40,000 inhabitants comprising people from most ethnic communities in Cameroon, some parts of neighbouring Nigeria, Niger and Ghana in search of fertile farmland and business opportunities . MutengeMuea is a semi-rural setting located at an altitude of 562 m above sea level on the Eastern flank of the active volcanic Mt Cameroon with relative humidity of above 80 %, temperature range of 18–28 °C and an annual rainfall of about 4096 mm. Muea has a heterogeneous and multiethnic population of approximately 9,000 inhabitants, majority of whom are farmers .Ethical clearance (No2013/0107/UB/FHS/IRB) was obtained from the University of Buea, Faculty of Health Sciences Institutional Review Board and administrative authorisation from the South West Regional Delegation of Public Health, Buea. Written informed consent was obtained before enrolment into the study.Pregnant women who reported for first ANC clinic visit on scheduled or unscheduled clinic days at the Mutengene or Muea Medical Centres and who gave their consent to participate in the study were enrolled. Pregnant women included those with or without fever.This prospective cross sectional study was carried out from March to August 2013 which corresponds to the period of intense malaria transmission in the mount Cameroon area. In the study area, routine ANC clinic visits (scheduled) are on Mondays and Thursdays while Tuesdays, Wednesday and Fridays are unscheduled days. Women reporting for first ANC clinic visit on scheduled or unscheduled clinic days and who volunteered to participate in the study were enrolled consecutively. At first scheduled or unscheduled visit, a structured questionnaire was used to document demographic information , gynaecologic/obstetric history , pregnancy complications, trimester of first visit), socio-economic indicators  and environmental factors . Monthly income recorded was self reported. IPTp-SP/dosage and ITN usage were recorded. Temperature was recorded using a digital thermometer and fever was defined as temperature > 37.5 °C. Maternal peripheral venous blood (2 ml) was collected by venipuncture into EDTA tubes for haematological assessment and malaria parasite determination. Human immunodeficiency virus (HIV) infection status was determined for mothers enrolled. Women in this setting are offered free routine confidential HIV testing and counselling at first ANC clinic visit. All samples were transported on ice bath to the Malaria Research Laboratory, University of Buea for analysis.Thick and thin blood films were prepared, stained with 5 % Giemsa for 30 minutes  and examA complete blood count was ran using a Beckman coulter counter® (URIT 3000) following the manufacturer’s instructions, that automatically generated values for haemoglobin (Hb) levels, WBCs, lymphocytes and platelets. Anaemia was defined as Hb levels < 11.0 g/dl .P. falciparum parasitaemia while asymptomatic infection was defined as malaria parasitaemia with temperature of ≤ 37.5 °C. All data collected were entered into SPSS  version 19  for analyses. Malaria parasite densities were log transformed before analysis. Associations between P. falciparum parasitaemia and malaria with type of clinic visit, house surrounding, educational level, age, gravity, trimester of pregnancy, insecticide treated nets (ITNs) usage, IPTp-SP uptake and haematological parameters were evaluated using Pearson Chi-Square (χ2) test. Differences in group means were compared using ANOVA, Student’s-t-test, Mann Whitney U or Kruskall Wahlis test. Multinomial logistic regression model was used to determine risk factors associated with P. Falciparum parasitaemia and malaria. Statistical significance was set at P < 0.05.Malaria (symptomatic infection) was defined as fever associated with A total of 303 women were enrolled. The mean age of the women was 24.1 ± 5.3 (range: 15–40) years while the mean gestational age was 23.4 ± 5.6 (range: 10–38) weeks. There was a significant positive correlation  between maternal age and gravidity. Most of the women attended their first ANC clinic in the second (58.1 %) and third trimester (36.2 %). The majority (68 %) of the women were married. About 90 % of the study participants had at least a primary education and only 11 % had obtained tertiary education. The greater percentage of the women reported a monthly income of less than 30.000 FCFA (franc des Communautés Financières d’Afrique) (approximately 60 USD). Anaemia was frequent (57.3 %) among pregnant women while HIV prevalence of 5.9 % was recorded. ITN usage was 49 % while IPTp-SP uptake was low (11.6 %)  parasites/μl. Geometric mean parasite density differed significantly with marital status, with single women (938) having significantly higher  GMPD when compared with married women (338). This difference was independent of age or gravidity status. Temperature was recorded for 257 women, out of whom, 24.9 % (64/257) had febrile illness (temperature >37.5 °C). Malaria parasitaemia was significantly  associated with febrile status, where a significantly higher percentage of women with febrile illness  were parasitaemic for P. falciparum infection when compared with aparasitaemic febrile cases . The overall prevalence of malaria and asymptomatic infection was 16.0 % (41/257) (95 % CI =11-20 %) and 10.5 % (27/257) (95 % CI = 7.3-15 %) respectively. The prevalence of malaria and asymptomatic infection differed significantly  with type of clinic visit  women enrolled were positive for P. falciparum parasitaemia or malaria. Nonetheless, a statistical significant negative correlation  was observed between percentage lymphocyte and malaria parasite density. More so, women who presented with fever had a significantly lower  mean percentage lymphocyte (28.63 ± 9.1) when compared with afebrile cases (34.52 ± 13.8).Haemoglobin levels, WBC counts, lymphocyte (counts and percentage) and platelet counts did not differ with the presence of P. falciparum parasitaemia and malaria. Younger women (≤20 years) were frequently infected (P = 0.002) with malaria parasitaemia and commonly had malaria (P = 0.018) when compared with older women (>20 years)  and/or standing water near houses were significant factors associated with r women ≤0 years wr women ≤0 years w2 = 57.73; P < 0.001). Similarly, there was a significant association  between marital and gravidity status where the majority of the unmarried women  were primigravidae and the married women predominantly  multigravidae. The presence of bush and/or standing water around house surroundings was a significant independent risk factor associated with P. falciparum parasitaemia  and malaria   among women of different gravidity status. Multigravid women  were more informed about MiP than secundigravidae  and primigravidae . On the contrary, women with knowledge on MiP (GA = 23.7 ± 5.6 weeks) had their first ANC clinic visit later  than those without knowledge on MiP (GA = 21.9 ± 5.1 weeks).To investigate the risk factors of malaria parasite infection and disease among pregnant women in the study area, multinomial logistic regression model was performed allowing adjustment for possible confounders. Being unmarried  was a significant independent risk factor associated with malaria parasitaemia . The majority (86 %) of these breeding sites are found within 20 m from the nearest inhabited house while 21 % are located between 20 and 50 m from the nearest inhabited houses. Breeding sites are less likely to occur beyond a distance of 50 m. Furthermore, Tanga et al. [This study investigated the risk factors for i et al.  revealedi et al. . The stua et al.  reportedP. falciparum infection and malaria suggests that the environment may modify the risk of infection and disease in pregnant women in this setting rather than gravidity. Areas of intense malaria transmission risk have been associated with greater risk of infection [In malaria endemic settings, studies have consistently shown that primigravid women are at increased risk of malaria , 6, 10. nfection . The assnfection . Women onfection . SP uptanfection . SP is anfection  and misuP. falciparum among single women might be due to the fact they were significantly younger (≤20 years) and the majority were primigravidae. In areas of high and stable transmission, age and pregnancy-associated anti-parasite immunity may play an important role in limiting P. falciparum infection to low parasite densities [In this study, more than twice (OR = 2.6) as many unmarried pregnant women were infected compared with women with a husband or partner. More so, single women had higher parasite densities than did married women. Similarly, in Yaoundé, Mbu et al.  showed tensities , 37 in oensities  reportedensities . Marriagensities , health ensities . Chepkemensities  suggesteensities . This inAlphavirus (Chikungunya and O’nyong-nyong) in sera of patients from Mt Cameroon area. Secondly, the study did not survey the environment where people stay to ensure that the reported presence of standing water and bushes around human residence faithfully relate to the risk of P. falciparum and malaria. While anopheline mosquitoes are predominant (82.73 %) and diverse in the Mt Cameroon area, Tanga et al. [Aedes, Culex and Mansonia species in some mosquito breeding sites and thus the possibility of DENV transmission in the area cannot be rule out. It is imperative that future studies investigate the prevalence of arboviral infections among pregnant women in this study area, as these infections are important and overlooked public health problems.This study had few limitations. First, the study did not investigate other arthropod- borne infections that cause fever. In Africa, fever episodes are most often considered as malaria (without any laboratory confirmation) and some of the supposed malaria attacks could be actual dengue fever which remains largely unknown . Two sera et al.  identifiP. falciparum parasitaemia is a leading cause of febrile illness among pregnant women before first ANC clinic visit and women with malaria frequently seek for medical attention during unscheduled clinic days. Being unmarried and the presence of bush and/or standing water in house surroundings modify the risk of P. falciparum infection and malaria in pregnancy. Prompt malaria diagnosis and proper treatment as well as early ANC care attendance to avoid adverse consequences of early infection in pregnancy is justified. Education on early ANC attendance and environmental sanitation are important public health targets for malaria control in pregnancy in this setting.In the study area,


Our laser device rapidly and noninvasively detected malaria in a patient and identified parasite-positive mosquitoes. Plasmodium falciparum infection in a malaria patient without drawing blood or using any reagent. This method uses transdermal optical excitation and acoustic detection of vapor nanobubbles around intraparasite hemozoin. The same device also identified individual malaria parasite–infected Anopheles mosquitoes in a few seconds and can be realized as a low-cost universal tool for clinical and field diagnoses.A fast, precise, noninvasive, high-throughput, and simple approach for detecting malaria in humans and mosquitoes is not possible with current techniques that depend on blood sampling, reagents, facilities, tedious procedures, and trained personnel. We designed a device for rapid (20-second) noninvasive diagnosis of  Malaria control and elimination would benefit greatly from an efficient and universal diagnostic tool that is fast (provides results in seconds), noninvasive and safe (uses no blood sampling or reagents), simple to use , sensitive and specific (detects low-level asymptomatic infections), and inexpensive and that detects malarial infection in humans and in mosquitoes –positive. To quantify the infection, we counted the incidence rate of malarial infection–positive traces IR  and calculated the Hemozoin Index (HI)  for 24 h. During hospitalization, the patient had mild hemolytic anemia and thrombocytopenia, neither of which required transfusion of blood products. In addition to the antimalarial medications and to provide symptomatic relief, the patient received intravenous fluids, antiemetics, and antipyretics.The patient was admitted to Ben Taub General Hospital, Harris Health System , with fever, myalgia, abdominal pain, nausea, and vomiting for the previous 4 days and no history of malaria chemoprophylaxis. Malaria identification and speciation was done by microscopy  and a rapid malaria antigen test . Both tests confirmed a We found wrist and ear lobe veins to be the optimal location for the test. Fingertips were also explored but were inadequate because some persons develop very thick and rigid skin patterns that prevent efficient transdermal delivery of a laser pulse to the blood vessels.The difference between the amplitudes of the in vitro background traces from intact blood , panel FAnopheles gambiae mosquitoes were fasted for 6 hours and fed on infected blood by using jacketed membrane feeders warmed to 37°C by a circulating water bath. Briefly, cultured P. falciparum (NF54 strain) gametocytes were diluted to 0.3% gametocytemia, 50% normal human erythrocytes and human serum for 15 min. After removing unfed mosquitoes, blood-fed mosquitoes were maintained at 26°C and 70%–80% relative humidity on 10% dextrose. Ten days later, 15 randomly selected mosquitoes were dissected to determine the oocyst numbers, and 2 other groups were used for the device evaluation. Midguts were stained with 0.1% mercurochrome and oocysts counted microscopically. Each of the 15 mosquitoes had 9–151 oocysts (median 50). The remaining mosquitoes were killed by freezing at −20°C before analysis using the prototype device. Mosquitoes fed on uninfected blood and maintained as described were used as negative controls (uninfected and oocyst negative). For further use as positive controls, mosquitoes were fed on uninfected blood containing 60 μg/mL hemozoin.For infection in mosquitos, female All procedures were approved by the corresponding internal review board committees at Rice University and Baylor College of Medicine . The patient and the volunteers provided informed consent.We have prototyped a diagnostic device  and evalTo obtain proof of the device feasibility, the prototype was further tested in human volunteers who did not have malaria and on a malaria patient with a similar skin tone (dark). We applied the probe to wrists and earlobes and positioned it over subcutaneous vessels. Thus, laser pulses were delivered to blood through the skin. No blood samples were taken, and no reagents were applied. Acoustic traces in response to each of 400 laser pulses (of the same fluence as described earlier) were collected simultaneously with laser irradiation  and analyzed statistically in real time. In the malaria patient, the parasitemia (percentage of infected erythrocytes) was determined by microscopy (thin blood film) and varied from 2%  4 hours before the device test to 0.3%  9 hours after the device test. As malaria-negative controls, we used healthy volunteers with similar skin tone and under the conditions and procedure applied to the malaria patient. Acoustic traces from the wrist of the malaria patient , panel D2, which is considered to be skin-safe according to federal regulations  by scanning the body across the probe under the same laser pulse fluence as described for the human studies. The negative control group  returned acoustic traces similar to those for hemozoin- and malaria parasite–negative human blood  an blood , panel HThese results provide a proof-of-principle for the H-VNB technology as a unique noninvasive transdermal diagnostic tool for malarial infection in humans and mosquitoes. The next step is to optimize the prototype with a malaria-specific laser operating at 672 nm for a better sensitivity (


Severe malaria may influence inner ear function, although this possibility has not been examined prospectively. In a retrospective analysis, hearing impairment was found in 9 of 23 patients with cerebral malaria. An objective method to quickly evaluate the function of the inner ear are the otoacoustic emissions. Negative transient otoacoustic emissions are associated with a threshold shift of 20 dB and above.This prospective multicenter study analyses otoacoustic emissions in patients with severe malaria up to the age of 10 years. In three study sites  144 patients with severe malaria and 108 control children were included. All malaria patients were treated with parental artesunate.p < 0.001, 95 % confidence interval 48.4–67.9 %) passed otoacoustic emission screening at the baseline measurement. The value increased to 65.2 %  at follow up 14–28 days after diagnosis of malaria.In the control group, 92.6 %  passed otoacoustic emission screening. In malaria patients, 58.5 % .The study population was divided into severe non-cerebral malaria and severe malaria with neurological symptoms . Whereas otoacoustic emissions in severe malaria improved to a passing percentage of 72.9 %  at follow-up, the patients with cerebral malaria showed a drop in the passing percentage to 33 % (n = 18) 3–7 days after diagnosis. This shows a significant impairment in the cerebral malaria group (The presented data show that 40 % of children have involvement of the inner ear early in severe malaria. In children, audiological screening after severe malaria infection is not currently recommended, but is worth investigating in larger studies. Hearingisorders . In chilisorders . So far,In a murine model, severe malaria causes significant hearing impairment . HistomoOAE testing is a specific objective hearing test, which has been used in neonatal hearing screening programs. The presence of OAE confirms a regular function of the inner ear. In transient OAE testing, a click stimulus is applied to the outer ear canal. This stimulus provokes a contraction of the outer hair cells in the inner ear, which can be detected afterwards in the outer ear canal. Transient OAE is generally evaluated as the cross-correlation or “wave reproducibility” of multiple measurements. Wave reproducibility is expressed as a ratio from –100 % to +100 %. A high cross-correlation or wave reproducibility is generally accepted as a measure of normal hearing . ReproduThe goal of this prospective multicenter study was to evaluate the inner ear function of children with severe malaria by using transitory evoked OAE and assessing the proportion of children who failed OAE examination.The present study was performed as a sub-study of “Severe Malaria in African Children II” (SMAC) (registered at PACTR201102000277177). The study and the sub-study have been approved by the local ethic committees in Kumasi, Ghana; Kisumu, Kenya; and Lambaréné, Gabon. The studies have been performed in three study sites: Komfo Anokye Teaching Hospital, Kumasi, Ghana; Centre de Recherche Mèdicale de Lambaréné, Lambaréné, Gabon; and Kenya Medical Research Institute, Kisumu, Kenya. Guardian informed consent was obtained from all participants.The study was designed as a prospective multicenter cohort study, comparing OAE measured in children with severe malaria to a healthy, local, age-matched control population. The malaria cohort was divided into a severe malaria group and a cerebral malaria group. OAE were measured in the control group once. In the malaria groups they were measured at the following time points: immediately after diagnosis of severe malaria prior to starting medication, 12–24 h after the first medication, after recovery, 3–7 days post diagnosis, and 14–28 days after the diagnosis. OAE were considered as absent if the reproducibility was below 60 %.The healthy control population was recruited at the study sites outside the hospital in schools and kindergartens. Children aged up to 10 years with no medical history of hearing impairment and no history of fevere related admission to hospital as an in-patient qualified for the control group . No furtPlasmodium falciparum malaria confirmed with a minimum parasitemia of >5000 parasites/μL on the initial blood smear; asexual forms of P. falciparum; and clinical manifestations that required hospitalization, like hyperlactatemia or metabolic acidosis, severe anemia, dark urine, hypoglycemia, jaundice, respiratory distress, severe vomiting, shock, abnormal bleeding, and/or renal failure [The severe malaria group was selected according to the following parameters: children up to 10 years; a diagnosis of  failure –21. AntiCerebral malaria was defined as severe malaria with neurological signs and symptoms like coma with a Blantyre Coma score of ≤ 2, repeated generalized seizures, focal neurological findings, or prostration . ProstraFor all groups, the ear inspection had to show a transparent tympanic membrane. Children with pathologic findings in the ear canal did not qualify. The baseline OAE testing was done immediately after confirmation of severe or cerebral malaria.Further clinical and laboratory examinations were performed in the severe malaria and the cerebral malaria group at the time of inclusion in the study. On admission the vital signs—heart rate, temperature, respiratory rate—were documented. The following clinical and laboratory parameters were examined: respiratory distress, deep breathing, severe vomiting, prostration, coma, repeated generalized seizures, jaundice, parasitemia, hemoglobin, platelets, glucose, creatinine, alaninaminotransferase, bilirubin, glucose, hemoglobinuria.As part of the SMAC II study, all patients were treated with parenteral artesunate after the first baseline measurement. The total dose of artesunate was 12 mg/kg, given as five intramuscular injections of 2.4 mg/kg at 0, 12, 24, 48, and 72 h or as three injections of 4 mg/kg at 0, 24, and 48 h either intramuscularly or intravenously. Allocation to one of the three schedules was random. Upon discharge patients received a weight-adapted standard regimen of artemether and lumefantrine .The OAE measurements were performed at each study site by two operators. Transient OAE were measured using a Madsen Capella Otoacoustic emissions machine . Both ears were measured five times and the best result was taken. If the reproducibility was below 60 % on one ear the OAE were considered absent. Each measurement consisted of 2080 repetitions. A broadband click ranging from 1 kHz to 4 kHz for 2 ms was used as a stimulus. The measurement was considered of a poor quality if the rejection rate exceeded more than 2000 sweeps. Therefore measurements were excluded with more than 2000 rejected sweeps. If the measurement at baseline did not work, the OAE testing was abandoned, and the treatment of the severely sick child immediately initiated. A second try was performed at the other measurement time points.t-test or Mann–Whitney U test for continuous variables and by chi-square test or Fisher’s exact test for proportions as appropriate. The proportions of patients passing the OAE test were compared between groups  by chi-square test. The study collective was categorized into severe non-cerebral malaria and cerebral malaria and statistically compared as described above. In order to identify prognostic factors for failing the OAE test at last follow-up, a univariate comparison of admission data was done between patients failing and patients passing on the last follow-up as described above. Variables with association at a significance level of p < 0.05 in the univariate analysis were entered into a stepwise logistic regression model with conditional forward selection as the selection method and age and sex as mandatory variables. The statistical evaluation was done by IBM SPSS statistics 21 .Otoacoustic measurements with a reproducibility below 60 % on one ear or both ears were considered a fail. Demographic and clinical data were compared between groups depending on data type and distribution either by Student’s p = 0.898). The mean age in the malaria group was 3.43 years (SD 2.384) and in the control group 3.82 years (SD 2.506).For the malaria group, 144 patients were recruited and 108 children as controls. Not all patients could be recruited for every measurement, partly because of prior administration of the study medication , or the patient being absolutely uncooperative , or due to loss to follow-up . No death was to be reported in the study population. The malaria group consisted of 67 female and 73 male patients (four were not documented), the sex distribution in the control group was 50 female and 56 male (two were not documented), with no significant difference in the sex distribution , 48.4–67.9 %) of all malaria patients passed the OAE testing. The results of the OAE are listed in Fig. p < 0.001 compared to the control group . A further significant difference was observed at the follow-up 14–28 days after inclusion  . Within 12 to 24 h, the passing percentage dropped slightly in the cerebral malaria group to 52.9 %  in contrast to a slightly improved passing percentage of 62.4 %  in the severe malaria population. The gap further increased at the 3–7 day measurement to a statistical significant value  were associated with hearing impairment at last follow-up. These variables were entered into a stepwise binary logistic regression model with age and sex as mandatory predictors. In this model the significant predictors associated with hearing impairment were age and the malaria severity group. Younger children had a significantly higher risk of hearing impairment Table . ChildreThe present study shows that severe non-cerebral and cerebral malaria  may leadThe control group was recruited outside the hospital with no admission to hospital as an in-patient for fever. Hospitalization for fever had been shown to be associated with significant hearing impairment in sub-Saharan Africa . The malIn the control group 92.6 % of the evaluated children passed the otoacoustic testing, indicating a functioning inner ear. An evaluation of preschool children performed in the US showed a similar passing percentage of 91 % in a group comparable for age and sex .In the malaria group the evaluated passing percentage started at 58.5 % at the baseline measurement and increased to 65.2 % at the follow-up 14–28 days after diagnosis, showing highly significant differences. This finding indicates that severe malaria affects the inner ear in 40 % of cases. Transient OAE are absent at a cochlea with hearing loss of 20 dB or more . SimilarCerebral malaria is known to influence the blood–brain barrier . SimilarThe statistical evaluation showed a significant association with hearing loss of the following parameters: reticulocyte count, respiratory distress, height, and repeated generalized seizures. Patients with a high count of reticulocytes showed less inner ear impairment than patients with a low count of reticulocytes. This can be explained by the fact that the “anemic malaria” results in an increased count of reticulocytes, but is less likely associated with hearing impairment . This inA limitation of the study appears to be the lack of tympanometry as an objective evaluation method of the outer ear canal and the middle ear. Lee et al. compared pneumatic otoscopy and tympanometry as tools to diagnose otitis media with effusions. With a sensitivity of 97.2 % otoscopy was superior to tympanometry, which had 87.5 % sensitivity. The gold standard to diagnose otitis media with effusions would be otomicroscopy with a sensitivity of 100 %. Ear microscopy was not available in the study setting. Therefore the better method of the remaining two was used . OAE tesThe fundamental pathologic cause of malaria-induced hearing loss is not fully understood. Severe malaria animal models have detected induction of apoptosis in the fibrocytes of the spiral ligament , in whicThe present data show that severe malaria and cerebral malaria lead to a cochlea malfunction of 20 dB or more in > 40 % of patients. With 240 million cases per year, only a small percentage of individuals with persistent hearing impairment still results in a huge number of disabled individuals, with enormous developmental and socioeconomic impact . These d


Plasmodium falciparum infection, nutrition, and food security in a cross-sectional survey of 365 Batwa pygmies in Kanungu District, Uganda in January of 2013. We identified 4.1% parasite prevalence among individuals over 5 years old. Severe food insecurity was associated with increased risk for positive rapid immunochromatographic test outcome . High age/sex-adjusted mid-upper arm circumference was associated with decreased risk for positive test among individuals who were not severely food-insecure . Within Batwa pygmy communities, where malnutrition and food insecurity are common, individuals who are particularly undernourished or severely food-insecure may have elevated risk for P. falciparum parasitemia. This finding may motivate integrated control of malaria and malnutrition in low-transmission settings.Although malnutrition and malaria co-occur among individuals and populations globally, effects of nutritional status on risk for parasitemia and clinical illness remain poorly understood. We investigated associations between  Plasmodium falciparum malaria and malnutrition are co-occurring health risks within populations globally; however, nutritional modulation of individual risk for P. falciparum parasitemia is complex and poorly understood.Plasmodia may indicate that micronutrient supplementation could increase incidence of clinical malaria within an endemic populationPresently hypothesized associations between malaria risk and nutrition suggest that resolving a population's chronic and acute nutrient deficiencies may lead to any of several scenarios for malaria control. Generally, improved individual immune function with better nutrition may limit health burdens from malaria resurgenceHousehold food insecurity, defined here as constrained social, economic, and physical access to sufficient and nutritionally adequate foods for maintaining a healthy and active lifestyle, is a determinant for malnutrition and associated comorbid conditions globally46P. falciparum for labile micronutrients within a human host, reverse-causal effects of malarial anemia on anthropometric indicators for nutritional status, and external socioenvironmental factors predisposing individuals to risk for all three conditions. For instance, as a cause for poor mental health outcomes, including psychological stress, food insecurity may disrupt normal host endocrine function and increase susceptibility to P. falciparum or other infections.Plasmodium infection while also limiting nutrient availability for parasite reproduction and pathogenesis within the human host.P. falciparum and P. vivax, with parasite prevalence between 3% and 4% and high adult to children under 5 years case ratios (> 1) characteristic of unstable malaria epidemiology.Relationships among food insecurity, malnutrition, and risk for malaria have been sparsely investigated and are difficult to observe given the multifactorial host-level biology of malnutrition and immune function, the need of ratios >  characteP. falciparum transmission is generally low and volatile in Uganda's southwestern highlands,As a marginalized, semisubsistent Indigenous population with pre-existing burden from these climate-sensitive health risks, the Batwa pygmies of Uganda are expected to be vulnerable to increasing malaria, food insecurity, and malnutrition amidst climate change.P. falciparum infection among Batwa pygmies and (2) contribute observational evidence of whether risk for current P. falciparum infection covaries with food insecurity or nutritional status in a low-transmission setting.Here, we present the outcomes of a malariometic, anthropometric, and food security census among Batwa pygmies residing in Kanungu District, Uganda. Our objectives were to (1) characterize the distribution of The study was approved in accordance with the requirements of the McGill University Policy on the Ethical Conduct of Research Involving Human Subjects and the Canadian Tri-Council Policy Statement on Ethical Conduct for Research Involving Human Subjects; the latter affords particular attention to issues among Indigenous and disadvantaged peoples. We, moreover, followed best practice frameworks for working with Indigenous communities in climate change vulnerability research.P. falciparum diagnostic tests. Respondents were interviewed in the Rukiga language by paid interviewers from Kanungu District. We sought to achieve census-level representation across all ages within Kanungu District's 10 Batwa settlements  Food Security Supplement to the Current Population Survey (CPS).z) scores for the analysis.We assessed variation in acute nutritional status using individuals' relative standing within the observed distribution of age- and sex-specific mid-upper arm circumference (MUAC), which was measured on an individual's left arm halfway between the tip of the shoulder and the tip of the elbow. In undernourished populations, MUAC outperforms body mass index (BMI) as an anthropometric measure for acute nutritional status among children and adults and as a prognostic indicator for malnutrition-related outcomes.P. falciparum . The test additionally bound Plasmodium lactate dehydrogenase (pLDH) antigen to indicate infection by non-falciparum Plasmodia (sensitivity of 95.5%). Previous work has validated rapid diagnostic tests (RDTs) in low-prevalence settings within east Africa, including in Uganda's southwestern highlands, indicating that RDT sensitivity, specificity, and predictive values do not differ appreciably from gold standard polymerase chain reaction (PCR) procedures.Parasitemia was inferred from positive outcome to an SD-BIOLINE rapid immunochromatographic assay detecting Histidine-rich protein II (HRP-II) antigen from N = 365), we opted a priori to consider models with k < 10 parameters only. Before model-building, we ruled out collinearity among main effects by Spearman's rank correlation coefficient (ρ) with a cutoff of 10%. We evaluated component plus residual plots to determine appropriate functional forms of continuous and factor variables. After identifying optimal main effect terms, we evaluated potential interactions through a model-based recursive partitioning algorithm testing candidate main effect terms for joint significance.k < 10 parameters, we assessed whether incorporating additional significant and non-significant terms resulted in global model improvement through likelihood ratio tests. Because individuals' risks for infection were stratified by heterogeneous exposure to parasite-positive vectors domestically and within communities, we calculated Huber–White SEs robust to multilevel clustering by household and settlement.P < 0.05 to be statistically significant. We conducted analyses within the R environment .We estimated parasitemia prevalence by dividing the number of positive test outcomes by the number of tests administered, assuming that all individuals to be at risk for infection. To examine associations between relative risk for positive RDT outcome and individual- and household-level factors within the study population, we built a multivariable log-binomial generalized linear model accounting for demographics , relative nutritional status and food security, behavioral protective measures, knowledge of malaria, and vector exposure through a modified best subsets variable-entry procedure.We limited analysis to cases among older children (> 5 years) and adults to avoid confounding caused by two factors. First, cases among individuals over 5 years old are atypical in hyper- and holoendemic areas relative to cases among children under 5 years old.1,409 = 0.01; P > 0.9). By Little's test for missingness completely at random, we identified no pattern to data missingness in the variables under consideration  and expected complete case analysis to yield unbiased results.Our sampling frame consisted of 630 individuals, among whom 475 individuals were over 5 years old. Of these individuals, we reached 434 individuals, among whom 407 individuals (94%) consented to clinical evaluation, including anthropometric measurement and finger prick. Complete household-level, individual-level, and clinical data were obtained for 365 individuals, representing a 77% overall response rate. Parasite prevalence did not differ significantly between participants who dropped out or completed the full survey . An overall χ2 deviance test comparing the fitted model with a saturated model with k = n parameters indicated that deviance for the fitted model was not significantly larger than for the saturated model. Accordingly, we expected a maximally fitted model to offer no significant improvement relative to the present model in accounting for variation in RDT outcomes.Variables rejected because of statistical non-significance of additive main effects included sleeping under an insecticide-treated bed net (ITN) the night before the survey, knowledge about malaria and its prevention, time spent in environments other than the forest, knowledge of or reported engagement in domestic and peridomestic malaria prevention efforts, recent travel outside home settlements, and household crowding (measured as the number of children occupying a house). Because only four households reported indoor residual spraying with insecticide within the last 12 months, this variable was not considered. Despite statistical non-significance, retaining age served to reduce noise within component plus residual plots, decrease global deviation , and reduce the largest studentized residuals. Consequently, we force-included age as a continuous variable in candidate models. We identified no significant outlying observations in the model controlling for age  establishing whether malnutrition-associated wasting precedes or follows infection among individuals who are not severely food-secure. Characterizing livelihoods among individuals who are severely food-insecure may likewise identify behaviors or exposures associated with risk for P. falciparum, accounting for non-nutritional factors explaining why persons in severely food-insecure households here had elevated risk for infection. Such research could extend current evidence regarding the safety or suitability of food security and nutritional interventions in P. falciparum-endemic settings and address differential implications of supplementation for parasitemia and malaria morbidity.Our findings motivate future longitudinal studies suited to identifying temporal and causal associations among the variables assessed here, which may account for immunological factors underlying nutritional associations with parasitemia. Two important objectives for understanding present outcomes will be (1) characterizing whether risk for food insecurity varies in phase with risk for