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Introduction : The emergence of Plasmodium falciparum resistance to artemisinin combination therapy (ACT) is a worrying development. It calls for close surveillance to monitor the efficacy of the drugs. The objectives of this study were to determine the performance of SD Bioline malaria AgPf(HRP-2/pLDH) 3 band Rapid Diagnostic Test (RDT) against Giemsa-stained blood smear and evaluate the suitability of this test in assessing the therapeutic efficacy of ACT in pediatric malaria patients in the Democratic Republic of the Congo (DRC). Methods : Five hundred and one patients with malaria symptoms were screened for P. falciparum in Kinshasa, DRC. Of the 166 patients who tested positive for P. falciparum at recruitment (day 0), 103 consented to participate in this study and were followed up and retested for P. falciparum on day 3, day 7, day 14, day 21 and day 28. Results : Sensitivity and specificity of the test were significantly high on day 0 and so were their positive and negative predictive values. Higher proportions of false positive cases were observed on the HRP-2 band irrespective of patient parasite densities during the follow up but these were barely seen on the pLDH band. Some patients turned positive during follow up but pLDH readings remained consistent with blood smear readings. Conclusion : SD Bioline malaria AgPf(HRP-2/pLDH) RDT demonstrated high performance in DRC. Thus, the test can be employed to assess the efficacy of ACT in pediatric malaria patients and prioritize areas that require the deployment of advanced testing like polymerase chain reaction (PCR). Key words : Malaria, AgPf(HRP-2/pLDH) RDT, artemisinin combination therapy, Democratic Republic of the Congo

The in vitro antiplasmodial activity and cytotoxicity of methanolic and dichloromethane extracts from five Congolese plants were evaluated. The plants were selected following an ethnobotanical survey conducted in D.R. Congo and focusing on plants used traditionally to treat malaria. The in vivo antimalarial activity of aqueous and methanolic extracts active in vitro was also determined in mice infected by Plasmodium berghei berghei.The growth inhibition of Plasmodium falciparum strains was evaluated using the measurement of lactate dehydrogenase activity. The extracts (aqueous, CH(3)OH, EtOH and CH(2)Cl(2)) were prepared by maceration and tested in vitro against the 3D7 (chloroquine sensitive) and W2 (chloroquine resistant) strains of Plasmodium falciparum and against the human normal fetal lung fibroblasts WI-38 to determine the selectivity index. Some extracts were also used at the dose of 300 mg/kg to evaluate their activity in mice infected since 4 days by Plasmodium berghei.Two plants presented a very high activity (IC(50)3 microg/ml). These plants were Strychnos icaja roots bark (MeOH and CH(2)Cl(2)) and Physalis angulata leaves (MeOH and CH(2)Cl(2)). One plant (Anisopappus chinensis whole plant, MeOH and CH(2)Cl(2)) presented a high activity (IC5015 microg/ml). The extracts of Anisopappus chinensis and Physalis angulata showed also a good inhibition of parasitemia in vivo. Flavonoids, phenolic acids and terpenes were identified in these plants by a general phytochemical screening method.Three plants showed a very interesting antiplasmodial activity (Anisopappus chinensis, Physalis angulata and Strychnos icaja) and one of them showed a good selectivity index (10, Anisopappus chinensis). Anisopappus chinensis and Physalis angulata were also active in vivo.

The present External Quality Assessment (EQA) assessed microscopy of blood parasites among diagnostic laboratories in the Democratic Republic of the Congo. The EQA addressed 445 participants in 10/11 provinces (October 2013-April 2014). Participants were sent a panel of five slides and asked to return a routinely stained slide which was assessed for quality of preparation and staining. Response rate was 89.9% (400/445). For slide 1 (no parasites), 30.6% participants reported malaria, mostly Plasmodium falciparum. Only 11.0% participants reported slide 2 (Plasmodium malariae) correctly, 71.0% reported "malaria" or "Plasmodium falciparum" (considered acceptable). Slide 3 contained Plasmodium falciparum (109/μl) and Trypanosoma brucei brucei trypomastigotes: they were each reported by 32.5% and 16.5% participants respectively, 6.0% reported both. Slide 4 (Trypanosoma) was recognised by 44.9% participants. Slide 5 (Plasmodium ovale) was correctly reported by 6.2% participants, another 68.8% replied "malaria" or "Plasmodium falciparum" (considered acceptable). Only 13.6% of routine slides returned were correctly prepared and stained. The proportion of correct/acceptable scores for at least 4/5 slides was higher among EQA-experienced participants compared to first time participants (40.9% versus 22.4%, p = 0.001) and higher among those being trained < 2 years ago compared to those who were not (42.9% versus 26.3%, p = 0.01). Among diagnostic laboratories in Democratic Republic of the Congo, performance of blood parasite microscopy including non-falciparum species and Trypanosoma was poor. Recent training and previous EQA participation were associated with a better performance. ispartof: PLoS One vol:11 issue:1 pages:1-15 ispartof: location:United States status: published

Background: The present External Quality Assessment (EQA) assessed reading and interpretation of malaria rapid diagnostic tests (RDTs) in the Democratic Republic of the Congo (DRC). Methods: The EQA consisted of (i) 10 high-resolution printed photographs displaying cassettes with real-life results and multiple choice questions (MCQ) addressing individual health workers (HW), and (ii) a questionnaire on RDT use addressing the laboratory of health facilities (HF). Answers were transmitted through short message services (SMS). Results: The EQA comprised 2344 HW and 1028 HF covering 10/11 provinces in DRC. Overall, median HW score (sum of correct answers on 10 MCQ photographs for each HW) was 9.0 (interquartile range 7.5 – 10); MCQ scores (the % of correct answers for a particular photograph) ranged from 54.8% to 91.6%. Most common errors were (i) reading or interpreting faint or weak line intensities as negative (3.3%, 7.2%, 24.3% and 29.1% for 4 MCQ photographs), (ii) failure to distinguish the correct Plasmodium species (3.4% to 7.0%), (iii) missing invalid test results (8.4% and 23.6%) and (iv) missing negative test results (10.0% and 12.4%). HW who were trained less than 12 months ago had best MCQ scores for 7/10 photographs as well as a significantly higher proportion of 10/10 scores, but absolute differences in MCQ scores were small. HW who had participated in a previous EQA performed significantly better for 4/10 photographs compared to those who had not. Except for two photographs, MCQ scores were comparable for all levels of the HF hierarchy and non-laboratory staff (HW from health posts) had similar performance as to laboratory staff. Main findings of the questionnaire were (i) use of other RDT products than recommended by the national malaria control programme (nearly 20% of participating HF), (ii) lack of training for a third (33.6%) of HF, (iii) high proportions (two-thirds, 66.5%) of HF reporting stock-outs. Conclusions: The present EQA revealed common errors in RDT reading and interpretation by HW in DRC. Performances of non-laboratory and laboratory staff were similar and dedicated training was shown to improve HW competence although to a moderate extent. Problems in supply, distribution and training of RDTs were detected.

Background : For the past decades, developing countries have received considerable support to fight infectious illnesses in their homelands. This global effort has tremendously reduced case fatality rates associated with illnesses such as HIV/AIDS, tuberculosis and malaria in many countries. However, this information is still missing in some developing countries, hindering international effort for control programs; we designed this study in effort to close this gap. Methods : Data on 23,487 inpatients from Kinshasa hospitals were gathered and analyzed using EpiData and SPSS. Major illnesses affecting inpatients were identified; mortality and case fatality rates associated with each such illness were estimated. Case fatality rates associated with each illness were compared between consecutive years. Socio demographic and economic factors associated with mortality due to HIV/AIDS, TB and malaria were investigated using logistic regression. Results : The outstanding findings were that case fatality rates associated with major illnesses were relatively higher in 2008 than in the previous year; inpatients hospitalized for HIV/AIDS, TB and malaria in 2008 were more likely to die than those hospitalized in the previous year. Low socioeconomic status inpatients hospitalized for malaria, HIV/AIDS or TB were more likely to die than high socioeconomic status inpatients (AOR 0.29, 95% CI 0.22–0.40; AOR 0.20, 95%CI0.12–0.33; AOR 0.33, 95%CI 0.21–0.53), even though both groups presumably had access to free life-saving treatment and care. Conclusion : These results indicate that while improvement in health indicators greatly depends on funds availability and sustainability, these alone might not be enough in resource poor developing countries. Other factors, i.e., population SES also need to be addressed before needed changes may occur. Keywords : Case fatality rates, Mortality, Socioeconomic status, Funding policies

To report the findings of a second external quality assessment of Giemsa-stained blood film microscopy in the Democratic Republic of the Congo, performed one year after the first.A panel of four slides was delivered to diagnostic laboratories in all provinces of the country. The slides contained: (i) Plasmodium falciparum gametocytes; (ii) P. falciparum trophozoites (reference density: 113,530 per µl); (iii) Trypanosoma brucei subspecies; and (iv) no parasites.Of 356 laboratories contacted, 277 (77.8%) responded. Overall, 35.0% of the laboratories reported all four slides correctly but 14.1% reported correct results for 1 or 0 slides. Major errors included not diagnosing trypanosomiasis (50.4%), not recognizing P. falciparum gametocytes (17.5%) and diagnosing malaria from the slide with no parasites (19.0%). The frequency of serious errors in assessing parasite density and in reporting false-positive results was lower than in the previous external quality assessment: 17.2% and 52.3%, respectively, (P0.001) for parasite density and 19.0% and 33.3%, respectively, (P0.001) for false-positive results. Laboratories that participated in the previous quality assessment performed better than first-time participants and laboratories in provinces with a high number of sleeping sickness cases recognized trypanosomes more frequently (57.0% versus 31.2%, P0.001). Malaria rapid diagnostic tests were used by 44.3% of laboratories, almost double the proportion observed in the previous quality assessment.The overall quality of blood film microscopy was poor but was improved by participation in external quality assessments. The failure to recognize trypanosomes in a country where sleeping sickness is endemic is a concern.Présenter les résultats d'une deuxième évaluation externe de la qualité de la microscopie de frottis sanguins colorés au Giemsa en République démocratique du Congo, réalisée un an après la première.Un ensemble de quatre lames a été livré aux laboratoires de diagnostic dans toutes les provinces du pays. Les lames contenaient: (i) des gamétocytes de Plasmodium falciparum, (ii) des trophozoïtes de PSur les 356 laboratoires contactés, 277 (77,8%) ont répondu. Dans l'ensemble, 35,0% des laboratoires ont correctement étudié les quatre lames mais 14,1% d'entre eux ont obtenu les bons résultats pour 1 ou 0 lame. Parmi les erreurs majeures figuraient le non-diagnostic de la trypanosomiase (50,4%), la non-reconnaissance des gamétocytes de P. falciparum (17,5%) et le diagnostic du paludisme sur la lame dépourvue de parasite (19,0%). La fréquence de ces erreurs graves dans l'évaluation de la densité parasitaire et de l'obtention de résultats faussement positifs est inférieure à celle de la précédente évaluation externe: 17,2% et 52,3%, respectivement, (P 0,001) pour la densité parasitaire et 19% et 33,3%, respectivement, (P 0,001) pour les résultats faussement positifs. Les laboratoires ayant participé à la précédente évaluation de la qualité ont obtenu de meilleurs résultats que les nouveaux participants, et les laboratoires situés dans les provinces présentant un nombre élevé de cas de maladie du sommeil ont mieux reconnu les trypanosomes (57,0% contre 31,2%, P 0,001). Les tests de diagnostic rapide du paludisme ont été utilisés par 44,3% des laboratoires, presque le double de la proportion observée dans la précédente évaluation de la qualité.La qualité globale de la microscopie de frottis sanguin était faible, mais a été améliorée par la participation à l'évaluation externe de la qualité. L'incapacité à reconnaître les trypanosomes dans un pays où la maladie du sommeil est endémique est une préoccupation majeure.Informar de los resultados de una segunda evaluación de calidad externa de la microscopía en frotis de sangre con tinción de Giemsa en la República Democrática del Congo, llevada a cabo un año después de la primera.Se entregó un panel de cuatro muestras a laboratorios de diagnóstico de todas las provincias del país. Las muestras contenían: (i) gametocitosDe los 356 laboratorios contactados, respondieron 277 (77,8%). En total, el 35,0% de los laboratorios informó de las cuatro muestras correctamente, pero el 14,1% informó de resultados correctos en una o ninguna de las muestras. Los principales errores consistieron en no diagnosticar la tripanosomiasis (50,4%), no reconocer los gametocitosLa calidad general de la microscopía en frotis de sangre se reveló escasa, pero experimentó mejoras gracias a la participación en evaluaciones de calidad externas. La incapacidad para reconocer los tripanosomas en un país en que la enfermedad del sueño es endémica es un problema.الإبلاغ عن نتائج تقييم ثان للجودة الخارجية للفحص المجهري للغشاء الدموي المصبوغ بصبغة غيمزا في جمهورية الكونغو الديمقراطية، الذي تم إجراؤه بعد التقييم الأول بسنة واحدة.تم إيصال مجموعة من أربع شرائح إلى المختبرات التشخيصية في جميع مقاطعات البلد. واحتوت الشرائح على ما يلي: (1) شريحة بها عرسياتتم تلقي ردود من 277 (77.8 %) مختبراً من بين 356 مختبراً تم الاتصال بها. وبشكل عام، أبلغت 35.0 % من المختبرات عن جميع الشرائح الأربعة بشكل صحيح غير أن 14.1 % أبلغت عن نتائج صحيحة لشريحة واحدة أو لم تبلغ عن أي شريحة. واشتملت الأخطاء الرئيسية على عدم تشخيص داء المثقبيات (50.4 %)، وعدم التعرف على عرسياتكانت جودة الفحص المجهري للغشاء الدموي بشكل عام ضعيفة ولكنها تحسنت بالمشاركة في تقييمات الجودة الخارجية. ويمثل الفشل في التعرف على داء المثقبيات في بلد يتوطن بها مرض النوم أمراً مثيراً للقلق.报告刚果民主共和国吉氏液染色血片镜检第二次外部质量评估报告的结果（在首次评估的一年后执行）。将四张载片组交给该国所有省份的诊断实验室。载片包含：(i)在联系的356 家实验室中，有277（77.8%）家响应。整体而言，35.0%的实验室正确报告所有四个载片，但是14.1%的实验室正确报告1 个或0 个载片结果。主要错误包括：未诊断出锥虫病（50.4%），未识别出血片镜检整体质量不良，但是通过参与外部质量评估得以改善。在有地方性昏睡病的国家无法识别锥体虫是一个需要关注的问题。Представить отчет о результатах второй внешней оценки качества микроскопии мазков крови с окрашиванием по Гимзе в Демократической Республике Конго, проведенной через год после первой.Набор из четырех микроскопических препаратов был доставлен в диагностические лаборатории во всех провинциях страны. В число микроскопических препаратов входили: (i) ГаметоцитыОтвет был получен из 277 (77,8%) лабораторий из 356 лабораторий, в которые был направлен запрос. В общем, 35,0% лабораторий представили все четыре правильных микроскопических препарата, но только 14,1% сообщили правильные результаты по 1 или 0 препаратам. К числу основных ошибок диагностики относится недиагностирование трипаносомоза (50,4%), невыявлениеВ целом, качество микроскопии мазков крови было низким, но оно улучшилось благодаря участию во внешних оценках качества. Неспособность выявить трипаносомы в стране, в которой сонная болезнь является эндемической, вызывает беспокойство.

Background In endemic settings, diagnosis of malaria increasingly relies on the use of rapid diagnostic tests (RDTs). False positivity of such RDTs is poorly documented, although it is especially relevant in those infections that resemble malaria, such as human African trypanosomiasis (HAT). We therefore examined specificity of malaria RDT products among patients infected with Trypanosoma brucei gambiense. Methodology/Principal Findings Blood samples of 117 HAT patients and 117 matched non-HAT controls were prospectively collected in the Democratic Republic of the Congo. Reference malaria diagnosis was based on real-time PCR. Ten commonly used malaria RDT products were assessed including three two-band and seven three-band products, targeting HRP-2, Pf-pLDH and/or pan-pLDH antigens. Rheumatoid factor was determined in PCR negative subjects. Specificity of the 10 malaria RDT products varied between 79.5 and 100% in HAT-negative controls and between 11.3 and 98.8% in HAT patients. For seven RDT products, specificity was significantly lower in HAT patients compared to controls. False positive reactions in HAT were mainly observed for pan-pLDH test lines (specificities between 13.8 and 97.5%), but also occurred frequently for the HRP-2 test line (specificities between 67.9 and 98.8%). The Pf-pLDH test line was not affected by false-positive lines in HAT patients (specificities between 97.5 and 100%). False positivity was not associated to rheumatoid factor, detected in 7.6% of controls and 1.2% of HAT patients. Conclusions/Significance Specificity of some malaria RDT products in HAT was surprisingly low, and constitutes a risk for misdiagnosis of a fatal but treatable infection. Our results show the importance to assess RDT specificity in non-targeted infections when evaluating diagnostic tests., Author Summary Rapid diagnostic tests (RDT) for malaria are currently rolled-out as the backbone of parasite-based diagnosis, and their diagnostic accuracy is sufficiently high to substitute microscopy. One decade ago, attention has been given to occurrence of limited false positivity in a number of malaria RDTs, but false positivity of RDTs has remained poorly documented since then. In the last years, the number of available RDT products has dramatically increased and test performance has improved. False positivity may therefore not be perceived as a problem anymore. In this manuscript, we demonstrate that specificities of malaria rapid diagnostic tests detecting parasite antigens are seriously affected by human African trypanosomiasis (sleeping sickness), with values down to 11%. Malaria constitutes the main differential diagnosis of human African trypanosomiasis, and the false-positive results for malaria RDTs increase the risk of misdiagnosis or delayed diagnosis of human African trypanosomiasis which is a fatal but treatable infection.

Background Rapid diagnostic tests (RDT) are increasingly replacing microscopy for diagnosis of malaria in endemic settings. Although RDTs are simple and robust, errors in the post analytical phase i.e. in reading and interpretation of the RDT result, are not uncommon. In the Democratic Republic of the Congo (DRC) malaria is endemic, and malaria RDTs have been introduced since 2010. In JuneJuly 2012, an external quality assessment (EQA) addressing correct reading and interpretation of the three band malaria RDT recommended by the National Malaria Control Programme was organized among end-users in DRC.

In areas with stable transmission, malaria is an alarming threat both for mothers (anemia) and fetus (abortion, premature birth, a birth ponderal deficit, death in utero). Our study aims at estimating the malaria prevalence among parturients and their newborn babies in Kinshasa, Democratic Republic of Congo, in order to conduct the national programme of control. Between September and November 2004, 196 pregnant women aged of 14 to 45 years old (average: 25.8 years) were recruited consecutively from four maternity hospitals in Kinshasa; those who received antimalarial drugs 2 weeks before delivery were not selected. The socio-demographic information and clinical symptoms/signs were obtained by questionnaire. Blood smears were performed on the mother's capillary blood, by placental apposition and with the newborn baby's blood. Smears were stained with Giemsa. 42 out of the 196 parturients (21%) were infected by Plasmodium falciparum. Parasites were found both in capillary blood and placenta of the 37 parturients; in 5 cases, only the placental appositions were positive. Prevalence was higher among primiparae (26.5%) than among multiparae (18.8%) (p = 0.20). 19.7% of the parturients who received an Intermittent Preventive Treatment (IPT) with sulfadoxine-pyrimethamin (SP) were positive. 13 out of the 196 newborn babies had a positive malaria smear. Malaria at delivery is thus a reality in Kinshasa, despite the use of SP as an IPT. The weak protection conferred on the IPT could be explained by the inefficacy of the SP a failing in prenatal record andlor by the low compliance of the mothers with this strategy This is the reason why we strongly recommend a large-scale evaluation of this strategy.

Objective: (i) Highlight field realities on proportions of parturient mothers who use during pregnancy intermittent preventive treatment-sulfadoxine-pyrimethamin (IPT-SP) without insecticide-treated net (ITN), IPT-SP in combination with ITN or ITN without IPT-SP; (ii) investigate associations existing between these preventive approaches with low prevalence of peripheral parasitemia, placental malaria, low birth weight (LBW) and anemia. Materials and Methods: Proportions of parturient mothers who utilize any of these approaches during pregnancy were estimated as well as associated rates of peripheral parasitemia, placental malaria, anemia and LBW; associations were investigated by comparing each group with participants who never utilized IPT-SP or ITN during pregnancy. Results and Conclusions: Of the 705 participants, 121 (17.2%) never used IPT-SP or ITN during pregnancy; 83 (11.8%) utilized ITN without IPT-SP and 501 (71.0%) utilized IPT-SP of those, 97% used IPT-SP1 and 3% used IPT-SP2/SP3. 275 (39%) used IPT-SP without ITN and 226 (32%) used IPT-SP in combination with ITN. While significant associations were found between: (i) Combined utilization of IPT-SP with ITN and low prevalence of peripheral parasitemia, placental malaria and LBW, (ii) utilization of IPT-SP without ITN and low prevalence of LBW and (iii) utilization of ITN without IPT-SP and low prevalence of placental malaria, no associations were seen between any of these approaches and low prevalence of anemia. Neither IPT nor ITN alone reduced as much adverse outcomes as when used together in combination, suggesting that in areas of moderate or high transmission of malaria, combined utilization of IPT-SP1/SP2 with ITN was the most effective approach for malaria prevention in pregnancy.

Background In endemic settings, diagnosis of malaria increasingly relies on the use of rapid diagnostic tests (RDTs) instead of microscopic examinations. False positivity of such RDTs is poorly documented, although it may be particularly relevant in infections for which the differential diagnosis includes malaria, such as sleeping sickness, a fatal but treatable disease caused by Trypanosoma brucei parasite subspecies. We therefore examined the effect of Trypanosoma brucei gambiense sleeping sickness on the specificity of malaria RDTs. Materials and methods Blood samples of 117 sleeping sickness patients and 117 matched non-sleeping sickness controls were prospectively collected in the Democratic Republic of the Congo. Reference malaria diagnosis was based on microscopy corrected by a four primer real-time PCR. Ten commonly used rapid diagnostic tests for malaria were evaluated including three two-band tests and seven three-band tests, based on the detection of Pf-HRP-2, Pf-pLDH and/or pan-pLDH antigens of Plasmodium. Results Specificity of RDTs for diagnosis of malaria in controls was between 97.5 and 100% and was between 11.3 and 98.8% in sleeping sickness patients. For seven out of 10 RDTs, specificity was significantly lower in sleeping sickness patients compared to controls. Decreased specificity of malaria RDTs in sleeping sickness was mainly caused by false positivity of the pan-pLDH test lines, but also occurred frequently for the HRP-2 test lines.The Pf-pLDH test lines were not affected. Conclusions