Plasmodium falciparum. It is commonly found as a complication of infection traveling to the brain. CM has a poor prognosis unless promptly identified and treated. This case report describes a 15-year-old girl who suddenly started experiencing a tonic-clonic seizure while playing. At the time of arrival at the emergency department, her vital signs were consistent with shock. She had hepatomegaly on physical examination, a hallmark of malarial infection due to an immune response against the proliferation of the protozoa. Peripheral blood smear for malaria parasites was positive for P. falciparum and P. vivax. The patient was started on intravenous (IV) saline, IV phenytoin, and IV metoclopramide. She was also transfused with two units of packed red blood cells. The patient was subsequently diagnosed with CM. For most patients, the course of treatment includes aggressive therapy with anti-malarial medications. She was started on broad-spectrum antibiotics and anti-malarial medications. Following two weeks of treatment, her condition improved significantly and she was discharged.Cerebral malaria (CM) is a severe infection of the brain caused by the parasite  Plasmodium falciparum [Neisseria meningitidis, Streptococcus pneumoniae, Enteroviruses, and herpes simplex virus(HSV). Diagnosis of meningitis caused by CM is made by cerebrospinal fluid (CSF) analysis. Often, CSF cultures are negative in malaria-endemic areas regardless of the presence of low-density parasitemia [Cerebral malaria (CM) is the most severe pathology caused by the malarial parasite lciparum . Unlike lciparum . Other basitemia . Here, wA 15-year-old female resident of Nigeria was brought into the emergency room (ER) in an unconscious state by her mother. The informant was her aunt whom she lived with. She was well until three days prior when she complained of a frontal headache, and her aunt treated her with acetaminophen. Her headache worsened until she lapsed into unconsciousness, necessitating her presentation to the ER. There was no history of fever, neck pain or stiffness, jaundice or skin eruptions, fatigue, confusion, or dizziness. A few hours into admission, she developed a generalized tonic-clonic seizure lasting for five minutes.P. falciparum and P.vivax. As shown in the urinalysis report in Table On physical examination, the patient had a Glasgow Coma Scale (GCS) score of 6/15, she was pale with cold clammy extremities, severely dehydrated, afebrile with a temperature of 36.7°Celsius, tachycardic at 101 beats per minute, blood pressure of 90/60 mmHg, respiratory rate of 20 breaths per minute, and oxygen saturation of 99% on room air. No obvious jaundice and no enlarged lymph nodes were noted. Neither Kernig’s nor Budzinski’s signs were positive, and her pupils were equally reactive and equal in size. Abdominal examination revealed hepatomegaly, and there was no tenderness or other organ enlargement. A urethra catheter was passed for urine monitoring which drained coke-colored urine, consistent with hemoglobinuria. Other physical examinations were unremarkable. The laboratory work was significant for anemia of 7.4 g/dL, low hematocrit of 21.6%, white blood cell count of 3.85 with an elevated neutrophil count of 83.7%, lymphopenia of 12.5%, low platelet count of 63,000, and high procalcitonin of 3.46 ng/mL. The peripheral blood smear for malaria parasite was positive for Abdominopelvic ultrasound revealed hepatosplenomegaly and ascites. CSF analysis and a computed tomography (CT) scan of the brain were unremarkable. Soon after the diagnosis was made, the patient was admitted into the intensive care unit. A nasogastric tube was passed and aspirated about 900 mL of gastric content mixed with blood. She was started on IV quinine for 48 hours with alternate IV normal saline and dextrose. She was also treated with phenytoin to control the seizure activities, as well as IV paracetamol and imipenem. She was administered two units of packed red blood cells on admission.On day two, the patient slowly regained consciousness; however, she started running a high-grade fever with a temperature of 38°Celsius. In addition, she had three episodes of non-bilious and non-projectile vomiting, but there was no associated abdominal pain. She was treated with metoclopramide and the vomiting stopped. She was thereafter transitioned to oral quinine, and clindamycin was added to her medications. A subsequent repeat of deranged laboratory values such as packed cell volume, procalcitonin, full blood count, and malaria parasite was normal.On day 13, she did not show any signs of altered mental status and was discharged with carbamazepine and multivitamins. She was scheduled for a follow-up in two weeks.Plasmodium parasites, causes a life-threatening disease in humans called malaria [Plasmodium parasite, P. falciparum and P. vivax are the most life-threatening species [P. falciparum leads to irregular fever patterns among affected patients. Malaria can be diagnosed by determining Plasmodium in the peripheral blood smear or bone marrow [The female Anopheles mosquito, a carrier of  malaria . Among t species . Malariae marrow . As showCM, the most severe neurological presentation of acute falciparum malaria, is a syndrome of unconsciousness and coma  and presUsually, CM results in endothelial lesions of the blood-brain barrier, and the endothelial lesions contribute to various brain injuries through sequestration of parasitized red blood cells in brain capillaries, cytokine production, immune cell/platelet accumulation, and microparticle emission . MedianaTreatment of CM carries its own risk. Our patient was in the intensive care unit and was treated with quinidine. Quinidine can cause torsades de pointes related to prolonged QT intervals, as well as hypoglycemia due to hyperinsulinemia ,11. Freq P. falciparum and P. vivax are the most life-threatening species. Patients experience an increased risk of death even with appropriate treatment early in the disease course, and some patients develop neurological sequelae. Therefore, it is imperative to identify and treat CM promptly due to its poor prognosis. In this case, the patient presented in an unconscious state with a tonic-clonic seizure, and her blood smear showed both P. falciparum and P.vivax. She improved significantly after aggressive inpatient medical management, followed by discharge from hospital admission in stable condition. To make an early diagnosis of CM and efficiently treat the patient, clinicians should have a high index level of suspicion in patients presenting with unresolved headaches in a malaria-endemic area or patients who traveled to an endemic area. They should commence treatment as soon as possible to improve neurocognitive outcomes.CM is a leading cause of malaria mortality among African children. It results in endothelial lesions of the blood-brain barrier and is the most severe neurological presentation of acute falciparum malaria. It is a syndrome of unconsciousness and coma, and it usually presents with fever, anemia, hepatomegaly, and seizures. The incidence is higher in children than in adults, and


Plasmodium spp. infections are a significant barrier to eliminating malaria. In malaria‐endemic areas, naturally acquired antimalarial antibodies develop with repeated infection. These antibodies can confer protection against the clinical manifestations of Plasmodium spp. infection in highly exposed populations, and several distinct functional antibody mechanisms have been defined in the clearance of Plasmodium parasites. However, the role of antimalarial antibodies during subclinical infection is less well defined. In this review, we examine the development and maintenance of antibody responses and the functional mechanisms associated with clinical protection, highlighted by epidemiological studies investigating the association between human immunity and detection of subclinical infection across various malaria transmission intensities. Understanding the development and role of the antimalarial antibody response during subclinical Plasmodium spp. infection will be essential to furthering novel interventions including vaccines and immunological biomarkers that can be utilized for malaria surveillance and ultimately progress malaria elimination.Undetected subclinical  This review examines the role of antibodies in sub‐clinical malaria, focusing on antibody development, and functional mechanisms, and epidemiological studies to inform malaria control interventions. AMAapical membrane AgCSPcircumsporozoite proteinEBAerythrocyte binding antigenLMlight microscopyMSPmerozoite surface proteinP.PlasmodiumPCRpolymerase chain reactionRDTrapid diagnostic testSpp.species pluralisWHOWorld Health Organization1Plasmodium spp. infections. Reported annual malaria cases do not represent the true number of Plasmodium spp. infections occurring worldwide, the majority of which are clinically silent, yet contribute significantly to ongoing transmission. Subclinical infection has been attributed to the non‐sterilizing nature of the naturally acquired antimalarial immune response in highly exposed populations. Such immunity is generally measured by the presence and magnitude of antibody responses in an individual or population. There are a number of studies investigating antigen‐specific antibody responses in protection from clinical manifestations of Plasmodium spp. infection, overwhelmingly they have found that increased antibody level or seropositivity, is associated with protection from clinical malaria. However, evidence for antibody‐mediated immunity in sub‐clinical Plasmodium spp. infection remains unclear. Here we review the literature examining naturally acquired antimalarial antibodies in sub‐clinical Plasmodium spp. infection in naturally exposed populations. Understanding the role of immunity in sub‐clinical Plasmodium spp. infection is crucial for our knowledge of malaria epidemiology and appropriate prevention, control, and surveillance activities, including novel vaccines and sero‐surveillance strategies.Malaria control and elimination efforts have resulted in remarkable success over the past two decades, however, the latest World Health Organization (WHO) reports reveal that progress in malaria elimination has begun to plateau and that global cases increased in 2020.2Plasmodium spp. and transmitted by Anopheles mosquitoes. There are five species of Plasmodium known to cause disease in humans , the two most prevalent and pathogenic being P. falciparum and P. vivax. Malaria is endemic to regions of sub‐Saharan Africa, South America, and the Asia Pacific. In high transmission settings in sub‐Saharan Africa, P. falciparum is the dominant species and children are at significant risk of serious illness compared to adults.Plasmodium spp. occurs.Malaria is a vector‐borne disease caused by the protozoan parasite Anopheles mosquito as it takes a blood meal. Sporozoites (pre‐erythrocytic stages) then invade hepatocytes,P. vivax, may remain there for many months or years in a dormant stage referred to as hypnozoites.Human infection begins with the inoculation of sporozoites by an infected female Plasmodium spp. infections that occur without the characteristic symptoms of malaria are referred to as subclinical . Though sometimes detectable by conventional diagnostics such as rapid diagnostic tests (RDTs) and light microscopy (LM), subclinical infections often consist of parasite densities below the detection limit of these diagnostic tools available in the field and clinical settings. In many malaria‐endemic settings, RDTs and LM have missed the majority of subclinical infections detected by more sensitive molecular diagnostics such as polymerase chain reaction (PCR), these are often referred to as sub‐microscopic infections that are in most but not all cases, also subclinical  stage of infection, namely the dominant circumsporozoite protein (CSP), can limit subsequent blood‐stage parasite density, and have been associated with delayed time to reinfection.Naturally acquired Abs are among the most studied antimalarial‐immune response, and several of their functional mechanisms have been associated with protection from clinical falciparum malaria (reviewed in ref. 18). Induction of the antibody isotypes with functional capacity for complement deposition and Fc receptor interaction is dependent on Th2‐like T follicular helper cell subsets, as demonstrated in experimentally infected, previously malaria naïve individuals.P. falciparum infection. The most studied targets include members of the merozoite surface protein (MSP) family including , involved in attachment and re‐orientation of the merozoite, invasion ligands such as apical membrane antigen (AMA1), and erythrocyte‐binding Ags ,Plasmodium spp. This includes for P. vivax infection where several Ag‐specific Ab responses have been identified but few have been associated with clinical protection (reviewed in ref. 49). Ultimately, the functional mechanisms against these targets slow parasite multiplication and limit blood‐stage parasitemia, preventing clinical symptoms associated with high parasite density. Despite strong evidence and defined mechanisms for a protective role of Abs in ameliorating clinical disease, the role of these mechanisms in perpetuating subclinical Plasmodium spp. infection and conversely how subclinical infection maintains antibody responses is less clear.The blood stages, merozoites and the infected erythrocyte, are more widely studied in association with protection from clinical malaria, particularly during 4P. falciparum transmission.The persistence of subclinical infections is generally accepted to be a consequence of frequent exposure to the parasite and the acquisition of nonsterilizing immunity, controlling parasite density below a clinical threshold. The persistence of anti‐merozoite IgG responses associated with circulating memory B‐cells has been shown for several years following the interruption of Plasmodium infection. The underlying subclinical infection itself has been implicated in protection from future incident malaria,Plasmodium spp.P. falciparum infection.The development of robust Ab responses in low transmission settings may require fewer exposures due to the lower number of circulating strains, and incident infections have recently been shown to be more likely to occur when there is the introduction of new strains.Plasmodium spp. infection have also been identified that are independent of acquired antimalarial immunity. Recent reports of replicating P. falciparum and P. vivax parasites accumulated within the spleen of splenectomized patients with no peripheral Plasmodium spp. infection provided evidence of a hidden parasite biomass.P. falciparum parasites detected in the dry compared to wet (high‐transmission) season, accounting for longer circulation of infected erythrocytes and increased splenic clearance.P. falciparum‐specific IgG responses and memory B cell activity compared to uninfected children, indicating that some preexisting exposure is associated with carriage of subclinical infection in highly seasonal transmission settings.Parasite factors that explain the carriage of subclinical P. falciparum infections are regularly cleared in the absence of treatment. In high transmission settings, participants regularly clear and become infected with new parasite strains, presenting as a single infection over a long period of time unless molecularly genotyped.P. falciparum and P. vivax infections oscillates frequently and that many infections are spontaneously cleared in a matter of months.Plasmodium spp. infection. Recent studies have demonstrated an association between Abs and protection from PCR‐detectable subclinical infection.Whether the development of protective Ab responses is responsible for the persistence of a low‐density infection, or if chronic low‐density infections are responsible for the maintenance of antimalarial antibodies remains unclear. Acquired antimalarial immunity is not generally thought to be sterilizing, and as discussed, chronic reservoirs of subclinical infection in intensely exposed populations are thought to be maintained by acquired immunity. Recent evidence suggests, however, that peripheral subclinical infections may not be as chronic as once thought, and that subclinical 5Plasmodium spp. infection comes from large‐scale serological surveys in malaria‐endemic populations. In many cross‐sectional surveys, antimalarial Abs are found at greater levels or in a greater proportion of subclinically infected individuals compared to uninfected community members for both P. falciparum and P. vivax infection across a range of transmission intensities (summarized in Table Plasmodium spp. infections, many studies have focused on sporozoite and merozoite‐specific responses with a lack of investigations into the role of Abs in response to the infected erythrocyte (Table Plasmodium spp. infection. Indeed, Abs can predict hot spots of infection in discrete regions,Evidence that Ag‐specific Ab responses are implicated in subclinical te Table . CollectPlasmodium spp. infection. Recent longitudinal investigations in naturally exposed populations, some implementing highly sensitive molecular diagnostics, have found antibody seroprevalence is associated with protection from clinical malaria and thus, an increased likelihood of subclinical parasitemia to some but not all parasite targets (Table Plasmodium spp. infection is significant.Longitudinal studies can provide stronger evidence of the role of antibodies in subclinical ts Table .86, 8ts Table .14, 96Plasmodium spp. infections will advance our understanding of the epidemiology of malaria. To progress this field, identifying the functional immune mechanisms associated with both chronicity and clearance of subclinical infection is required, as are well‐designed, longitudinal serological surveys with highly sensitive molecular diagnostics and inclusion of diverse malaria‐endemic populations. Most evidence to date has focused on the response to merozoite‐specific IgG responses, and future inclusion of a greater diversity of parasite targets including variant surface Ags may provide further insight into the role of Ab responses in the prevalence and maintenance of subclinical Plasmodium spp. infection. Additionally, a greater focus on non‐falciparum Plasmodium spp. infection is required, particularly for P. vivax where protective functional mechanisms are yet to be elucidated. Knowledge gained from these in‐depth investigations will be vital to the development of malaria elimination tools, including novel vaccine targets and sero‐surveillance markers.Furthering our knowledge of antimalarial immunity and the natural course of subclinical K.O. and M.S.R. performed the literature searches. K.O. wrote the first draft of the manuscript. M.S.R. and F.J.I.F. contributed to the writing of the manuscript. F.J.I.F. conceived the manuscript. All authors agreed to the final submission of the manuscript.The authors declare no conflicts of interest.


Plasmodium species, and even if most attention is directed to those species that cause human malaria, Plasmodium parasites can infect several other vertebrates, including birds. This Special Research Topic of Frontiers in Immunology brings original, mini-reviews, perspectives, and hypothesis and theory articles that explore the parasite-host relationship during malaria infection in humans and birds.The interaction between a pathogen and its host can lead to health or disease. Malaria is caused by different P. falciparum and predominantly affects children in the first years of life and pregnant women. The course of infection depends on the interaction between the host’s immune response and the parasite virulence. In severe cases, a dysregulated immune response is present. Furthermore, an interaction between the parasite and the endothelium could lead to endothelial activation. Even if diagnostics of malaria are quite well established, the prognosis of severe and fatal malaria cases is very challenging. A host factor that could be useful in this prediction is the soluble urokinase-type plasminogen activate receptor (suPAR), as reviewed by Stefanova et al..In humans, malaria is mainly caused by Sakoguchi and Arase reviewed recent discoveries of how surface erythrocytic antigens of P. falciparum can interact with human inhibitory immune receptors.Parasite surface antigens, such as PfEMP1, STEVOR, and RIFIN proteins, interact with the host and are important for parasite evasion. In this Research Topic, P. falciparum. In this special Research Topic, Barateiro et al. wrote a Perspective article, reviewing the main findings concerning placental autophagy in malaria, and hypothesized that placental autophagy plays a cytoprotective role during infection in pregnancy.The interaction between the parasite and the host was also discussed in the context of malaria in pregnancy (MiP). MiP can be a severe disease and various factors can be important for the final outcome. Despite that, the molecular mechanisms involved in gestational malaria are still not clear. Placental autophagy is modulated primarily by immune signals and inflammation and is dysregulated in pregnant women infected with Plasmodium cycle, heme is released upon red blood cell rupture, which can lead to inflammation and tissue damage. The main mechanism of heme catalysis is through the activity of the enzyme heme oxygenase 1 (HO-1). HO-1 plays a dual role: while it is associated with successful pregnancy; in inflammatory conditions, it can induce iron overload and cell death. In this Research Topic, Cariaco et al. investigated the role of HO-1 in gestational malaria using a murine model. They found that HO-1 expression and activity are enhanced in Plasmodium infection during early pregnancy. They also showed that ZnPPIX treatment decreased the ability of HO-1 to develop its deleterious effect when in excess.During the erythrocytic Palinauskas et al.). Microbiota was also evaluated in the context of an avian malaria vaccine .Host-microbiota interactions can lead to resistance or susceptibility to an infection. In this Research Topic a ‘Hypothesis and Theory’ paper discusses the use of bird-malaria-microbiota system to understand the implications of gut microbiota diversity to anti-α-Gal (Understanding how the parasite interacts with the host and how the host responds to this interaction is fundamental for developing new tools for malaria interventions. In this Research Topic, aspects of the parasite-host relationship were discussed.All authors listed have made a substantial, direct, and intellectual contribution to the work and approved it for publication.The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.All claims expressed in this article are solely those of the authors and do not necessarily represent those of their affiliated organizations, or those of the publisher, the editors and the reviewers. Any product that may be evaluated in this article, or claim that may be made by its manufacturer, is not guaranteed or endorsed by the publisher.


The recent worldwide increase in malaria cases highlights the need for renewed efforts to eliminate malaria. The World Health Organization advocates that malaria surveillance becomes a core intervention. Current methods to estimate the malaria burden rely on clinical malaria case reports and surveys of asymptomatic parasite infection mainly from children  < 5 years. In this study the hypothesis was that screening blood donors for malaria parasites would provide real-time information on the asymptomatic reservoir of parasites in the adult population and mirror other surveillance data.This study was conducted in Malawi, a high malaria burden country, at the Malawi Blood Transfusion Service, which collects blood units at donation sites countrywide. A secondary analysis was conducted on data obtained from a prior Sysmex XN-31 analyser malaria diagnostic evaluation study utilizing residual donor blood samples. XN-31 malaria results, donor age, sex, geographical location, and collection date, were analysed using standard statistical methods.The malaria parasite prevalence in blood donors was 11.6% (614/5281 samples) increasing seasonally from December (8.6%) to April (18.3%). The median age was 21 years and 45.9% of donors were from urban areas, which showed a lower prevalence compared to non-urban regions. The Central administrative region had the highest and the Northern region the lowest malaria parasite prevalence. The donors were predominantly male (80.2%), 13.1% of whom had malaria parasites, which was significantly higher (p < 0.0001) than for female donors (7.4%). Multivariable logistic regression analysis showed that age, location, and collection month were significant predictors of malaria positivity in males, whereas in females only location was significant. There was no gender difference in parasite density nor gametocyte carriage.This study demonstrates the powerful utility of screening blood donors for malaria parasites using the XN-31, which not only improves the safety of blood transfusion, but provides valuable complementary surveillance data for malaria control, especially targeting males, who are generally excluded from periodic household surveys. Blood donations are sourced countrywide, year-round, and thus provide dynamic, real-time information on the malaria burden. Furthermore, the XN-31 identifies the asymptomatic human reservoir of infectious gametocytes, which must be targeted to eliminate malaria. Malaria, despite being preventable and treatable, with an estimated 241 million cases in 2020 in 85 malaria-endemic countries, continues to be a major contributor to the global health burden . As the Key elements of an effective malaria surveillance strategy are to systematically and continuously collect high quality data on the burden of malaria and to integrate these findings into national health information systems . These dAnopheles mosquitoes, it is also readily transmissible via transfusion of infected blood [Whilst malaria is fundamentally viewed as a vector borne parasitic disease, transmitted by the bite of infected female ed blood –7. The aed blood , notablyPlasmodium falciparum [The Sysmex XN-31 analyser (XN-31), a new higher diagnostic sensitivity technology for automated malaria detection, was recently evaluated at the Malawi Blood Transfusion Service (MBTS) with the specific objectives to determine the sensitivity and specificity of XN-31 for malaria screening and to estimate true malaria parasite prevalence in the MBTS donor pool. The study confirmed that the XN-31 had excellent sensitivity and specificity, revealing an overall malaria parasite prevalence of 11.6% amongst blood donors, in contrast to the 6.5% detection rate for thick smear microscopy, the method in routine use for malaria screening at the time of the study . Malawi lciparum . HoweverThe objective of this secondary data analysis was to provide a detailed analysis of the donor demographics and assess whether the observed patterns and trends mirror those obtained from standard malaria surveillance activities, such as household malaria indicator surveys (MIS). The ultimate aim was to show that donor malaria screening could be used as a source of real-time surveillance data, in line with the third GTS pillar, namely, to transform malaria surveillance into a core intervention. Understanding the nature of the reservoir of malaria parasites in asymptomatic individuals is a key component along the path towards malaria elimination.Malawi is a high malaria burden country with perennial transmission and the entire population at risk of infection. This study was conducted at MBTS, a centralized service which collects blood units at numerous donation sites countrywide, covering 27 of the 28 administrative districts (excluding Likoma island). There are 4 static (permanent) donation sites in Blantyre, Balaka, Lilongwe and Mzuzu which are in the Southern, South-Eastern, Central and Northern regions of Malawi respectively and mobile units in all 27 districts. Blood samples were transported to one of these 4 permanent sites and then to the central laboratory in Blantyre for centralized blood-borne pathogen screening and blood typing.All blood donations collected by MBTS are from voluntary unpaid donors. Pre-donation screening consists of two stages: (1) educating blood donors on the eligibility criteria for blood donation aimed at encouraging self-deferral among individuals willing to donate blood and (2) administering a blood donation form which contains the national blood donation acceptability criteria. This helps screen out those who are not eligible to donate blood. The exclusion criteria for blood donation are extensive and include the following: age below 16 or above 65 years; body weight below 42 kg; a feeling of unwellness; engagement in behaviour deemed to increase risk for transfusion transmissible infections ; past medical history suggestive of HIV, hepatitis or syphilis; past or present history of renal, cardiovascular, central nervous system and metabolic disorders such as insulin-dependent diabetes; and haemoglobin below 12.5 g/dl. Only individuals who satisfy the national blood donation acceptability criteria are accepted as blood donors.Most blood collection (~ 65%) takes place at mobile collection sites. MBTS staff visit donation sites 1–2 weeks prior to the scheduled donation day to sensitize potential donors with regards to the donation process and eligibility criteria. MBTS also provides an incentive to encourage repeat donation through their milestone rewards programme, which entails standardized gifts, comprised of MBTS branded items, such as caps, T shirts and certificates, depending on number of donations made, from the third donation onwards.This study is a secondary data analysis of a prior study  that wasPlasmodium species causing human malaria. The analyser also provides a concurrent complete blood count measurement for each sample analysed.The measurement principle of XN-31, an analyser essentially identical to its research-use-only predecessor the XN-30, has been detailed elsewhere . BrieflyThe XN-31 malaria results were segregated based on the administrative district location of the individual blood collection sites, weather zones as defined by the Department of Climate Change and Meteorological Services, Ministry of Forestry and Natural Resources, Malawi  as well Donor age, sex and geographic location were analysed to ascertain the primary determinants of malaria prevalence.® Statistical Software version 19.8 . A descriptive analysis was done to summarize donor demographic data such as sex and age. Continuous data were expressed as medians and interquartile ranges (IQR) while categorical variables were summarized as percentages. Prevalence was estimated by dividing the number of malaria positive samples by the total sample size. An acceptable minimum sample size for prevalence estimation was set at 30 [Data analysis was done using MedCalcet at 30 . MalariaThe details of comparative microscopy and XN-31 malaria screening results are described in detail elsewhere . In brieThe donor group was comprised of 80.2% (4236/5281) male, 17.4% (918/5281) female and 2.4% (127/5281) for whom this information was not captured.The median age of blood donors was 21 years (IQR 19–28), with 95% of donors between 16 and 47 years, and the oldest 64 years. Female donors were significantly younger  with median age of 19 years (IQR 18–25) compared with males . There were also differences in age distribution based on collection centre. Donors, whose samples were sent to the Lilongwe blood collection centre were the oldest  while donors from Mzuzu were the youngest . The median age of donors from Balaka and Blantyre was 20 years (IQR 19–25) and 21 years (IQR 19–27), respectively. Donors from exclusively urban areas  with median age of 25 years (IQR 20–32) were significantly older  compared with 20 years (IQR 18–23) for donors from predominantly non-urban collection sites .Blood samples included in this study were from blood donations collected in 24 of the 28 Malawian administrative districts, which are grouped into the Northern, Central and Southern administrative regions. The weather zones are comprised of the northern areas, central areas, lakeshore areas, Southern Highlands, and the Shire Valley. The 5281 samples included in the study comprised ~ 25% of the total donations collected during the study period, with 45.9% coming from urban donors. Although urban donors seem “oversampled”, considering that the Malawi population is 16% urban and 84% non-urban , no weigThis secondary data analysis incorporated data from blood donor samples collected from December 2019 to April 2020. The trends of the monthly malaria prevalence averages for all districts; urban versus non-urban donation site locations; low prevalence zones ; and high prevalence zones  are shown in Fig. The sex distribution of malaria positive donors overall and by prevalence zones is shown in Fig. To investigate what drives the difference between male and female donors, a multivariable regression analysis was undertaken to quantify differences of potential confounding factors such as age, urban vs non-urban location and month of collection in males and females Table . For fem3/µL (IQR 0.066 to 0.440) and 0.155 × 103/µL (IQR 0.057–0.626) for male and female donors respectively. There was also no difference (p = 0.9472) in gametocyte carriage amongst male (12.4%) and female (12.7%) parasitaemic donors. The median parasite density was however significantly higher (p = 0.0027) in donors in whom gametocytes were detected  by the XN-31 analyser, compared with malaria-infected donors where gametocytes were not detected  , whereas 94.7% (72/76) had both ring forms and gametocytes (median MI-RBC 0.252 x 103 p/μL). The median MI-RBC value differences for these two groups was statistically significant (p = 0.0023).There was no difference (p = 0.7933) in parasite density between male and female infected donors Fig. a. The meIn 2020 the Sysmex XN-31 automated malaria detection analyser was evaluated at the MBTS, utilising samples from 5281 blood donors, collected nationally from 24 of the 28 administrative districts in Malawi . In thisFor malaria elimination to succeed, it is essential that countries generate and use high quality surveillance data to accurately define malaria incidence over time and space . DependiThe most recent Malawi MIS took place in 2021, and prior to that in 2017 . The MISThe overall donor malaria prevalence rate in this study was 11.6%, increasing progressively from December (8.6%) to April 18.3%) Fig. a. This s% Fig. a.These observations highlight that the episodic nature and short duration (1–2 months) of data collection utilizing epidemiological household surveys, and the delay in such results becoming available, makes accurate tracking of rapid, year-to-year changes in malaria burden a difficult task .The current study data concur with the generally reported higher malaria risk associated with non-urban areas , 27. TheP. falciparum infections in Eastern Uganda [The study blood donor pool was 80.2% male with 95% of donors between 16 and 47 years. Female donors were slightly younger (median age 19 years) compared with males (median age 21 years). Urban donors comprised 45.9% of the pool with an even stronger male representation (90.2%), and significantly (p < 0.001) older (median age 25 years) compared with non-urban donors (median age 20 years). This urban male dominance and older age, accounted for the age difference between male and female donors overall. Parasitaemia rates were significantly (p < 0.001) and consistently higher in male donors (13.1%) compared with female donors (7.4%), irrespective of location or month of collection. This male bias of parasitologically confirmed malaria infection prevalence is in keeping with several other studies conducted in Africa , 29, 30.n Uganda , utilizin Uganda  and incrn Uganda , these fn Uganda .For surveillance data to be translated into meaningful interventions and to monitor their impact, it needs to be representative of the community at risk of malaria. Sample size determination and sampling for a typical MIS is a highly complex process, and whilst the bigger the better, it is a balancing act between the demands of the analysis, implementation team capacity and budget constraints . The 201Besides providing quantitative parasite counts, the XN-31 analyser also provides information on the presence of gametocytes, as a research-use-only parameter. Gametocytes were observed equally in malaria-infected male and female donors, with the parasite burden (MI-RBC count) being significantly higher in gametocyte carriers compared with malaria-infected donors where gametocytes were not detected Fig.  in line This study has limitations. Firstly, the original study was designed exclusively to confirm analytical performance of the XN-31 analyser as a malaria diagnostic test, hence the focus was on inclusion of a sufficient number of blood donors over a certain time-period with no regard for proportional representativity of donors based on districts (blood collection site), urban versus non-urban location nor month of collection. Consequently, reliable parasitaemia prevalence rates could not be established for administrative districts with too few or no donor inclusions in this study nor could weighting be applied to adjust for this disproportional representation. Also, only donor samples received on low workload days , were included, meaning that the study sample composition may differ from the complete donor pool. Furthermore, the data collection did not span a full year hence it does not cover the full seasonal range of malaria transmission. Topazian et al. , using qBlood donors in malaria-endemic countries are an untapped source for the systematic measurement of the asymptomatic parasite reservoir. The XN-31 detects malaria with high diagnostic accuracy and provides parasite burden quantification. Thus, the XN-31 automated analyser is an ideal tool for malaria screening of blood donors, which could not only contribute towards increased blood safety, but also serve as a complementary source of surveillance data for malaria control. Blood collection is a vital healthcare activity in all countries, and perhaps more so in malaria-endemic countries, such as Malawi, where malarial anaemia  is a key


Sri Lanka, an island nation, has eliminated endemic malaria transmission. Maintaining elimination in the continued presence of vectors requires vigilance in screening people travelling from high malaria-risk areas and a rapid response with focal screening for infections identified in the community. Such screening requires accurate and very rapid assays that enable an immediate response. Both microscopy and rapid diagnostic tests (RDTs) have limitations including sensitivity and speed in screening large numbers, while polymerase chain reaction (PCR) is practical only as laboratory confirmation. This study assessed the utility of ‘Gazelle’, a novel rapid malaria assay based on magneto-optical detection of haemozoin, a by-product of malaria parasite metabolism.Between October 2020 and March 2021, two groups of individuals were screened for malaria by four methods, namely, microscopy, Rapid Diagnostic Test (RDT), Gazelle and PCR. Passive case detection was carried out for confirmation of diagnosis amongst individuals suspected of having malaria. Individuals at high-risk of acquiring malaria, namely persons returning from malaria endemic countries, were screened by active case detection.Plasmodium falciparum infections, two Plasmodium ovale, and one each of Plasmodium vivax and Plasmodium malariae. Gazelle correctly detected the P. vivax, P. ovale and P. malariae infections within the 2 min test time, but did not detect two P. falciparum infections giving a sensitivity of 77.8%. Specificity was 100%.Of the 440 individuals screened for malaria, nine malaria positives were diagnosed by PCR, microscopy and the HRP2 band of RDT, which included five P. falciparum detection, will be important to broaden utility.The Gazelle, a portable bench top device proved useful to screen a large number of blood samples for non-falciparum parasites within 5 minutes of sample input. Species differentiation, and improvement in The online version contains supplementary material available at 10.1186/s12936-022-04283-7. Plasmodium vivax infections accounted for over 90% of malaria infections diagnosed. Plasmodium falciparum was present but its incidence varied over time and Plasmodium malariae was also prevalent till about the late 1960’s after which transmission was interrupted. The Anti Malaria Campaign (AMC) began to classify cases as indigenous and imported since 2008 [Sri Lanka received malaria-free certification from the World Health Organization (WHO) in September 2016, becoming the second country in the WHO South East Asia region to be declared malaria-free [nce 2008 , 4, therP. vivax cases, 84 P. falciparum, 26 Plasmodium ovale, 6 P. malariae, and one mixed infection. The high number of imported malaria cases being diagnosed combined with high receptivity due to the presence of the primary vector Anopheles culicifacies and emergence of Anopheles stephensi [The number of imported malaria cases reported over 5 years, between 2017–2021, included 97 Two malaria case surveillance strategies play a key role to prevent the reestablishment of malaria in Sri Lanka. Passive Case Detection (PCD), which is the detection of malaria cases amongst people who seek health care on their own, usually for fever; and Active Case Detection (ACD), which involves actively searching for malaria infections, such as in people or populations at high risk but who may or may not be obviously ill , 8. LaboP. falciparum (due to high sensitivity of HRP2) and P. vivax but a lower sensitivity has been recorded for the pan specific pLDH for P. falciparum, P. ovale and P. malariae [Microscopy remains the traditional gold standard for malaria diagnosis in Sri Lanka. Microscopy is performed by Public Health Laboratory Technicians (PHLT), who are posted at government hospitals across the country . Challenmalariae . This hamalariae . Howevermalariae , 12. SucHaemozoin, produced when the malaria parasite digests haemoglobin, which is its primary nutrient source found as component of red blood cells, has been identified as a bio-marker for malaria diagnosis given its magnetic and birefringent properties –15. As tThis study aimed to compare a new haemozoin-detecting device, the Gazelle, with microscopy, Carestart™ RDT and nested PCR for screening and diagnosis of malaria in people suspected of having malaria over a period of six months.This study was carried out between October 2020 and March 2021 at the Central Laboratory of the AMC Headquarters, Colombo, Sri Lanka. Two groups of individuals were screened for malaria (a) the first group comprised individuals referred to the AMC Headquarters (AMC HQ) Central Laboratory for confirmation of malaria based on clinical suspicion of disease. The reasons for clinicians suspecting and referring for confirmation of malaria included (i) travel history to a malaria endemic country, (ii) prolonged fever of unknown origin and, (iii) signs and symptoms suggestive of malaria, (b) the second group comprised individuals at high risk of malaria, namely security forces personnel returning from United Nations peacekeeping missions in Africa and individuals repatriated from African destinations who were screened by active case detection (ACD) by the AMC.Proportion of agreement between the screening test and the gold standard was assumed as 50% to obtain the maximum sample size. Desired precision considered was 5%. α error was taken as 0.05. The calculated minimal sample size was 418 after allowing for a non-response rate of 10% .Two mL of intravenous blood was collected under aseptic conditions by trained nurses in to EDTA tubes after obtaining written informed consent from the participants to be included in the study. Blood samples of individuals suspected of having malaria were referred to the AMC by clinicians in state and government hospitals. In the case of high-risk individuals who had arrived from malaria endemic countries, the samples were collected within 24 h of arrival in Sri Lanka at the port of entry and transported to the AMC Headquarters.The samples were screened for malaria by microscopy, Carestart™ RDT and Gazelle within 24 h of collection and PCR were carried out thereafter for confirmation of diagnosis. The assays were performed in a blinded manner using coded samples.Thick and thin blood films were prepared from the collected blood samples. The slides were stained with Giemsa and screened for malaria parasites using microscopy, independently in a blinded manner by a PHLT, who is an expert microscopist and a parasitologist. A smear was interpreted as negative only after examination of 100 fields with an oil immersion lens at 1,000 magnification , 19. If CareStart™ Malaria Pf/PAN (HRP2/pLDH) Ag Combo Test, product G0131 (Access Bio Inc. USA) was used for malaria diagnosis. Performance of the tests and interpretation of the results were done according to the manufacturer’s instructions and the Standard Operating Procedures prepared by the AMC [P. falciparum and pan-specific lactate dehydrogenase, pLDH) was recorded for all positive test results. the AMC . BrieflyA point-of-care portable magneto-optical device, Gazelle was also used for malaria diagnosis. Gazelle comprises a reader  and single use disposable cartridges Fig. . Prior tPlasmodium genus specific nested PCR was carried out for all samples. Primers rPLU1/rPLU5 were used for the genus specific nest 1 reactions. The product of the first amplification reaction (1 µL) was used as the template for the genus specific second amplification reaction (nest 2) with the oligonucleotide primers rPLU 3 and rPLU 4. The base pair position of these primers with respect to the 18S rRNA A-type gene of P. falciparum (GenBank accession no. M19173) are for rPLU3 132–161 and for rPLU4 353–364 respectively. This gave a PCR product of approximately 235 bp. Whenever the genus-specific nest 2 PCR revealed positive results, the following species-specific nest 2 primers were used to determine the Plasmodium species: rFAL1/rFAL2 , rVIV1/rVIV2 (P. vivax), rMAL1/rMAL2 , rOVA1/rPLU2 , and Pmk8/Pmkr9 (P. knowlesi). PCR products were electrophoresed for 30 min on a 2% (w/v) agarose gel at constant voltage of 120 V in 0.5 × Tris Borate EDTA buffer. The separated bands were visualized under an ENduro GDS UV trans illuminator (312 nm wave length) after staining for 15 min with ethidium bromide. Fragment sizes were estimated relative to the 100 bp ladder marker. A sample was considered as positive for Plasmodium if a DNA fragment of the expected size of 235 bp was visible. The products of the primary genus specific PCR reaction was used as the template for four separate species specific nested PCR assays.To confirm the presence or absence of parasites and parasite species, all samples were analysed by nested PCR assay at the AMC HQ Central Laboratory according to standardized protocols and published methods . In brieSTARD (Standards for Reporting of Diagnostic Accuracy Studies) shown in the Additional file The clinical data and laboratory results were collected on record forms and later entered into a Microsoft Excel database. Data recorded on the Gazelle was transferred to the parent company after each batch of samples was run.The performance of Gazelle was calculated and compared with microscopy, RDT and PCR (the reference standard) with 95% confidence intervals for the following values: sensitivity, specificity, negative likelihood reactions, odds ratio and positive and negative predictive values . The total number of samples is those samples on which all four diagnostic tests were successfully performed.440 individuals were screened for malaria over a six month period between October 2020 and March 2021. Of the 21 individuals that were screened by passive case detection eight gave a history of travel to a malaria endemic country . The other 13 patients had not visited any overseas country within the past 1 year but were referred to exclude malaria in the differential diagnosis. The balance 419 were classified as a high-risk population for malaria and screened by ACD  Nine malaria positive patients were diagnosed by microscopy Table , which iP. falciparum, two P. ovale and one each of P. vivax and P. malariae infections. The results of Carestart RDT (HRP2 band) and PCR were consistent with microscopy . Specificity was 100% , sensitivity 100%) and specificity (100%) between microscopy, RDT and PCR methods. The results of comparison of Gazelle with other methods are given in Table 0% Table . One mis00% and sP. falciparum malaria positives. However, during the study period, the Gazelle reader, which is a portable device, weighing less than 5 kg proved rapid and otherwise accurate in the rapid screening of blood samples. The benefits of using Gazelle in this study was identified as the need for low level of user training to perform the test, the ability to screen a large volume of samples in a short period of time and the rapidity of obtaining results.This study aimed to determine the utility of the Gazelle as a rapid screening device for malaria infection, using typical populations where rapid screening would be an advantage. The low positive rate in this population limited the power to determine test accuracy. Compared to microscopy, RDT (HRP2) and PCR, Gazelle did not detect two P. falciparum is a concern. Haemozoin has been demonstrated in early ring stages [P. falciparum at parasite densities below 40 parasites/uL at the ring stage [P. vivax has also been shown to be similar to microscopy and better than RDTs [The failure to detect the two ring stage infections of g stages , but inag stages , 27. Othng stage , 29. Gazhan RDTs .Plasmodium species [P. ovale and P. malariae (noting the low numbers of these species in this study) which could prove useful due to the fact that Carestart RDT (pan specific-LDH) shows a lower sensitivity to diagnose both these species in Sri Lanka [Treatment in Sri Lanka is based on species identification and thus, until species identification is possible, Gazelle will not be able to be used as a sole test for diagnosis but it has the potential to be used as a screening test at special locations such as ports of arrival. The developer is developing a species-specific algorithm based on the variation in haemozoin crystal morphology between  species . Gazelleri Lanka .The blood samples collected for this study were transported to the AMC central laboratory where Gazelle was carried out. Should the test be carried out at a port of entry, it may prove to be cumbersome due to the requirement of pipettes to measure and insert the required blood volume and diluent into the Gazelle cartridge. This could be overcome by preparation of a one-step cartridge for Gazelle that could directly collect finger prick blood samples. This method is in development and should be available in the market in the near future.P. falciparum can be addressed, it would be a better tool considering the very short time required for testing and giving the result.In the absence of its ability to quantify and demonstrate the parasite species, Gazelle will be of advantage to be used in combination with HRP2 Carestart RDTs as a screening tool to screen large numbers of high-risk individuals arriving at ports of entry. Compared to the RDT which takes approximately 15–20 min, Gazelle can be performed in a very short time hence if the limitation regarding detection of Gazelle, a haemozoin based malaria detecting device has potential for rapid throughput malaria screening in Sri Lanka due to its ease of use and availability of results within a short period of time. Sri Lanka is in the phase of prevention of re-establishment, where not only a limited number of malaria cases are diagnosed, but accurate diagnosis of each and every case is essential to prevent onward transmission. Thus, the technical challenges such as species identification, quantifying parasite density and ability to perform the test using finger prick blood should be addressed and overcome if it is to be used as a stand-alone screening device in a country such as Sri Lanka.Additional file 1: Figure S1. STARD Diagram.Additional file 2:Table S1. STARD checklist for the reporting of studies of diagnostic accuracy. STARD Checklist.


Malaria presents a diagnostic challenge in most tropical countries including Rwanda. Microscopy remains the gold standard for diagnosing malaria, however, it is labour intensive and depends upon the skill of the examiner. Malaria rapid diagnostic tests (MRDTs) have been developed as an easy, convenient alternative to microscopy.A cross sectional study was conducted from October to November 2019 on 130 febrile patients who were directed to the laboratory department for blood screening for malaria parasites at Byumba Health centre. The main objective of this study was to correlate Microscopy and MRDTs in diagnosis of malaria.After signing a consent form, blood samples were collected and screened for malaria parasites microscopically and by using MRDTs. Data collection forms were filled with relevant information and obtained results for MRDTs and for peripheral blood smear were recorded. The collected data were statistically analyzed using GraphPad Prism 9 software. The mean age found to be 16 years old. In this study peripheral blood smear microscopy was considered as a reference method. The sensitivity and specificity of RDT Histidine–Rich Protein 2 (HRP-2) were calculated and found to be 96.6% and 60% respectively. The negative predictive value was found to be 92.85% where positive predictive value was 73.3%.MRDTs should be used along with microscopy to avert complications associated with delayed diagnosis and similar studies are required to identify alternative techniques with high specificity for the diagnosis of malaria. The traditional method of microscopic identification of parasite however, is not only daunting in poor power setting, but also time consuming and requiring a lot of expertise/training. Thus, the peripheral blood smear examination technique is generally used, however, it is limited to large clinics/tertiary centres. This conventional staining of peripheral blood smears/microscopy still remains the gold standard in laboratory diagnosis of malaria.4 Malaria Rapid Diagnostic Tests (MRDTs) are commercially available in kit forms with all necessary reagents and the ease of performance of the procedures does not require extensive training or equipment to perform or to interpret the results. Results are read in 12 to 15 minutes. MRDT mainly come in two forms. One is antigen based and normally requires the use of haemolysed red blood cells while the other is antibody based and normally requires the use of extracted serum. Generally speaking, antibodies are better expressed in serum otherwise plasma could also stand in place of serum for antibody-based method.6 This study aimed to correlate Microscopy and MRDT (Histidine – rich protein 2 (HRP-2), Ag of plasmodium falciparum) in diagnosis of malaria at Byumba Health centre.Malaria is one of the highest killer diseases affecting people in tropical countries especially in Africa. The 2019 World Health Organization (WHO) Malaria Report estimates that there were 228 million malaria cases and 405,000 deaths in 2018. According to WHO, the first priority for all countries where transmission rates of malaria are high or moderate is to ensure maximal reduction of morbidity and mortality through sustained provision of universal access to quality-assured and appropriate vector control measures, diagnostics and antimalarial medicines, while retaining the long-term vision of malaria eradication.This study was conducted out in the laboratory department of Byumba Health Centre, located in Northern Province, Byumba Sector, Gicumbi District, Rwanda. The city lies about 60 kilometres (37 mi), north of the capital Kigali. This location lies approximately 30 kilometres (19 mi), south of the International border with Uganda at Gatuna.A cross sectional study design was conducted among patients of Byumba Health Centre. Data was collected from October to November 2019.130 Samples that tested for Malaria were used. The samples were taken from patients who attended Byumba Health Centre between October and November 2019.All male and female patients attending Byumba Health Centre between October and November 2019 with clinical suspicion of malaria based on fever and or history of fever within the previous 48 hours were eligible for inclusion in the study. Lack of consent and incomplete data constituted the exclusion criteria.The study was approved by the research committee of Byumba Health Centre, accredited by Byumba District Hospital. The objectives and procedure were carefully controlled according to set Standard Operating Procedures (SOPs). Written consent was sought for from study participants or study participant's caretaker for minors. To ensure confidentiality, numbers were used as study participants' ID instead of names on patient's data extraction forms.After filling the consent form, Blood sample was collected from the middle or ring finger of the patient by using lancet. 5μl of whole blood from the finger was added into MRDT, then 4 drops of assay diluents were added into MRDT according to the manufacture's protocol and test to detect malaria parasite/antibody detection method. The results were read after 15 min. Similarly, peripheral blood smears were made on a clean slide and allowed to air dry before being sent to the Parasitology laboratory. 10% Giemsa stain was used to stain thick smear for 15 minutes and tested with right microscopy. The results from both MRDT and microscopy were reported qualitatively (Positive or Negative). MRDT results and thick smear results were recorded on the data collection sheet. This was done within one hour from the collection time. Materials were consisted of Giemsa stain, microscopic slides, and light microscopy with good 100X objectives, MRDTs kits and lancets.Collected data was checked and analysed using GraphPad Prism 9 software. The validity of diagnostic test was used in calculation and then the measurements were reported in number and percentage.The study considered a total of 130 participants; 70(54%) females and 60(46.1%) males. The mean age of the participants was found to be 16 years. The demographic characteristics of the study subjects are shown in Proportions of malaria by MRDT and peripheral blood smear microscopy are presented in 15In this study, microscopy was considered as a method of reference. The sensitivity of MRDTs HRP2 in diagnosis of malaria is reported at 96.6%, this is presented in 1613 These results mean that if you tested negative for Malaria by MRDT (HRP-2), you would have 92.85% chances of not having the disease. When you tested positive for Malaria with MRDT (HRP-2), you would have a chance of 73.3% of truly having the disease.Predictive values of MRDT (HRP-2), positive and negative were calculated as shown in 7 The disease has a worldwide distribution and is found throughout the tropics, sub-Saharan Africa, South East Asia, the Pacific islands, India, Central and South America. Malaria caused by Plasmodium falciparum predominates in Africa where the mortality attributed to it approaches 1 million annually, and accounts for 90% of the global malaria burden.8 Majority of these deaths are of children under the age of 5 years. Thus, one child dies of malaria in Africa every 30 seconds, which translates into a tragic 3000 children each day. Many of the children who survive an episode of severe malaria suffer from brain damage and cognitive disability, consequently crippling these families with its debilitating aftermath.9 Malaria presents a diagnostic challenge in most tropical countries like Rwanda. Microscopy remains the gold standard for diagnosing malaria, but it is labour intensive and depends upon the skill of the examiner. RDTs have been developed as an easy, convenient alternative to microscopy.10 Poor diagnosis of malaria implies under diagnosis and inappropriate treatment procedures.3 MRDTs are known to capture at least 3 target antigens: lactate dehydrogenase (LDH), Plasmodium falciparum histidine-rich protein 2 (PfHRP2) and pan-plasmodial aldolase. HRP-2 MRDTs are the most sensitive for parasite detection and are heat-stable under field conditions compared to the other antigen tests.12 However, HRP-2 MRDTs have limitations, as their performance has been shown to be affected by product quality and parasite-related factors such as pfhrp2/3 gene deletion, non-P.falciparum species and prozone effects that may lead to false-negative MRDTs.13 Microscopy is the most widely tool used to diagnose malaria at peripheral levels. In capable hands it is very sensitive for parasitaemia ≤50/μL (0.001%)19 and it can give important information to the clinician like species, parasites stages and parasite density.Malaria affects a significant number of people across the world each year and is the most wide-spread parasitic disease encountered.18 The study also observed high negative predictive values of MRDTs compared to the positive predictive values. This finding is in contradiction with findings of a similar study conducted in Egypt.13 Several studies conducted elsewhere have shown varrying degree of false negative result for MRDT because of hyperparasitaemia, deletion or mutation of HRP-2 gene and the prozone effect .21The observed high sensitivity (96.6%) and specificity in this study is similar to reports of other studies conducted elsewhere.This research on the correlation of malaria rapid test and peripheral blood smear microscopy has been carried out on patients who attended Byumba Health Centre, suspected to have malaria. The sensitivity of HRP-2 based Rapid diagnostic test for malaria was (96.6%) and high to the specificity of this type of MRDT with 76.4%. The negative predictive values of MRDTs were high compared to the positive predictive values. If you tested negative for Malaria by MRDT (HRP-2), you would have 92.87% chances of not having the disease. When you tested positive for Malaria with MRDT (HRP-2), you would have a chance of 78.38% of truly having the disease. In this study, there aretests that were considered as false positive  while Other tests were reported as false negative . The results obtained by MRDT (HRP-2), for malaria parasites should be confirmed with other Tests of high specificity such as microscopy and Polymerase Chain Reaction. Health professionals are recommended to confirm MRDTs results with microscopy before administering treatment and precautions on the uses of MRDTs, transportation of kits and samples correction should be taken into consideration. Further studies should determine the most appropriate type of malaria diagnostic test to be used in combination with microscopy and MRDTs.


Malaria causes anemia by destruction of red blood cells and inhibition of erythropoiesis.We assessed whether the magnitude of the malaria‐specific effect on anemia differs by age, during low and high malaria seasons.In rural Zambian children participating in a pro‐vitamin A efficacy trial, we estimated differences in the prevalence of anemia (defined as hemoglobin < 110 g/L for children < 60 months. and < 115 g/L in older children) by malaria status and assessed malaria‐age interactions. Regression models (with anemia as the outcome) were used to model malaria‐age interaction in both the low and high malaria seasons, controlling for potential confounders.n = 820 children). In the low malaria season, anemia prevalence was 29% in malaria‐negative children and 54% in malaria‐positive children (p < 0.001), with no malaria‐age interactions (p = 0.44). In the high malaria season, anemia prevalence was 41% in malaria‐negative children and 54% in malaria‐positive children (p < 0.001), with significant malaria‐age interactions (p = 0.02 for anemia). Age‐stratified prevalence of anemia in malaria positive versus negative children was 67.0% versus 37.1% (in children < 60 months); 57.0% versus 37.2% (in 60–69 months.); 46.8% versus 37.2% (in 70–79 months.); 37.0% versus 37.3% (in 80–89 months) and 28.0% versus 37.4% (in 90+ months).Average age was 68 months at baseline (Malarial anemia is most severe in younger children, especially when transmission is intense. Anemia control programs must prioritize this vulnerable group. WorlMalaria‐related anemia is characterized by destruction of parasitized and especially non‐parasitized erythrocytes, and impaired erythrocyte production during malaria episodes , 14. The22.1This study included 4‐ to 8‐year‐old children from Mkushi District in Central Province, Zambia, an area with a high burden of malaria and anemia , 21, 22.Briefly, we conducted baseline and endline surveys in September 2012 and March 2013, respectively, which corresponded with the low and high malaria transmission seasons, respectively. At both timepoints, we collected socioeconomic data and morbidity history and measured height to the nearest 0.1 cm using a Shorr board, weight to the nearest 0.1 kg with an SECA 874 digital scale and axillary temperature using a digital thermometer. Children presenting with a high fever (axillary temperature > 39.5°C) were referred to the nearest health center. Using a syringe, we collected from each child, ∼7 mL of venous blood into a 10 mL collection tube . A malaria rapid diagnostic test (RDT)  was used to test for malaria in the field. Children who had positive RDT results during the baseline and endline assessments were treated with artemether and lumefantrine  in accordance with Zambian guidelines . Thick aEthics approval was obtained from the Institutional Review Board of the Johns Hopkins Bloomberg School of Public Health in Baltimore, Maryland, USA, and the Ethics Review Committee of the Tropical Disease Research Center (TDRC) in Ndola, Zambia.2.21‐acid glycoprotein (AGP) , ferritin , and soluble transferrin receptor  at TDRC. We measured C‐reactive protein (CRP) using an Immulite 1000 analyzer  at JHSPH. We used reverse‐phase high performance liquid chromatography to measure serum retinol concentration at Craft Technologies, Inc., in Wilson, North Carolina, USA. All baseline assessments were repeated at the endline, 6 months later.Thick and thin malaria microscopic slides were read at the TDRC laboratory. Slides were stained with 3% Giemsa, washed, dried, and read by two independent microscopists. Where necessary, a third independent reading was performed to resolve discordant results. Parasite density was estimated using thick films by counting the number of parasites per 200 white blood cells. A slide was considered negative if no parasites were detected after counting 200 oil immersion fields. For each positive thick slide, the corresponding thin film was read to determine the Plasmodium species. Commercial ELISA kits were used to determine serum concentrations of α2.3Anemia was defined as hemoglobin < 110 g/L for children < 60 months and < 115 g/L for children ≥ 60 months . Malaria2.4n = 820). With a sample size of 820 (274 anemic cases and 546 non‐anemic cases) and 5% type‐1 error rate, we had 80% power to detect a difference of 10% in anemia prevalence comparing malaria‐positive children and malaria‐negative children.Because the study intervention had no impact on hemoglobin concentration, we pooled data from all children who had a hemoglobin measurement and malaria microscopy at both time points, regardless of treatment allocation  < 0.1. In the event of significant age interactions, plots were constructed to visualize how the association between malaria and anemia (or hemoglobin) changed by age. Ferritin, retinol, and sTfr were log‐normalized.Indicators of nutritional status, socio‐demographic status, inflammations, anemia, and malaria at baseline and endline are summarized as means or proportions Tables  and 2. Pp = 0.05. All data analyses were conducted with STATA 13 software (StataCorp). In adjusted models, iron status (defined by ferritin and sTfR), vitamin A status (defined by serum retinol), and inflammation (defined by CRP and AGP) were included as covariates.For all other tests, statistical significance was set at 3n = 820) who had complete baseline and endline data for hemoglobin, ferritin, sTfR, malaria, CRP, and AGP at both baseline and endline. The children in this subsample were statistically similar to those in the overall trial with respect to age and gender. The mean (± SD) age of the study population was ∼68 months (±15) with ∼35% below the age of 5 years . In addition, no effect modification by age was observed (p‐interaction = 0.45). In the high malaria season, however, both hemoglobin and anemia differed significantly by malaria status, and in addition, a significant age interaction was observed. After adjusting for covariates, mean hemoglobin was 17 g/L lower in children diagnosed with malaria compared to children without malaria (p = 0.01). As shown in Table p = 0.02). Furthermore, significant age interactions were observed in the high malaria season . Figures In the low malaria season, there was no statistically significant difference in the hemoglobin concentrations among children with or without malaria after controlling for age, iron status, vitamin A status, and inflammation , the malaria‐specific effect on anemia was age‐specific. In the youngest age group (<60 months), we observed ∼30 percentage point difference in anemia prevalence between children with and without malaria. However, this difference became narrower with age. Beyond the age of 80 months, this difference was no longer apparent.The observed high risk of malarial anemia among younger children could be due to a number of factors, including the degree of micronutrient deficiency, outdoor exposure, and malaria‐specific immunity, which improves with age. We previously showed that both iron and vitamin A status influenced malaria and anemia risk in this population , 27. BecThere are several strengths, but also some limitations with the current study. This study is limited by its observational nature, particularly considering the complex etiology of anemia in children. Hemoglobin levels are driven by a complex interplay of nutritional factors (particularly iron metabolism), inflammation and in the context of malaria, by factors which modulate erythrocyte destruction and synthesis . BecauseIn conclusion, our results have important implications for anemia control programs and research in settings with endemic malaria. Our data suggest a need to focus limited resources in preventing and treating malaria in vulnerable young children. In Zambia, preventative strategies such as distribution of insecticide treated bed nets free of charge to children under five are available. In this study, we provided mosquito nets to all children after the baseline survey. Our baseline and endline surveys (data not shown) found that a substantial proportion of children  did not sleep under mosquito nets the previous night. Furthermore, because the incidence of malaria was generally high in older children, it plausible that the degree of outdoor exposure is an important risk factor in malaria transmission. Hence, it is important that existing interventions are coupled with educational programs designed to increase awareness about optimal health behavioral practices in the context of malaria prevention and control. Our results suggest that, particularly for young children, anemia control interventions should be integrated into existing malaria control programs. Additional research is needed to understand the pathophysiologic pathways which make young children particularly vulnerable to malarial anemia.Clement O. Acheampong drafted the manuscript. Clement O. Acheampong and Maxwell A. Barffour analyzed the data. All authors reviewed and approved the content of the manuscript. Clement O. Acheampong, Maxwell A. Barffour, and Amanda C. Palmer take responsibility for the final content of this paper.The authors declare that there is no conflict of interest that could be perceived as prejudicing the impartiality of the research reported.Table S5. Age distribution of anemia prevalence by malaria status in Zambian children during the high malaria seasonClick here for additional data file.