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- Hindawi Publishing Corporation
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- Copyright © 2018 Pembe Issamou Mayengue et al. This is an open access article distributed under the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.
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- 10.1155/2018/4914358
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Abstract
Hindawi Malaria Research and Treatment Volume 2018, Article ID 4914358, 6 pages https://doi.org/10.1155/2018/4914358 Research Article Evaluation of Routine Microscopy Performance for Malaria Diagnosis at Three Different Health Centers in Brazzaville, Republic of Congo 1,2 1 Pembe Issamou Mayengue , Dezi Kouhounina Batsimba, 2,3 1,2 2 Louis Régis Dossou-Yovo, Roch Fabien Niama, Lucette Macosso, 1 2 2 Brice Pembet Singana, Igor Louzolo, Nadia Claricelle Bongolo Loukabou, 4 1 2 Géril Sekangue Obili, Simon Charles Kobawila, and Henri Joseph Parra Facult´e des Sciences et Techniques, Universit´e Marien Ngouabi, BP 69 Brazzaville, Congo Laboratoire National de Sant´e Publique, BP 120 Brazzaville, Congo Ecole Normale Sup´erieure, Universit´e Marien Ngouabi, BP 69 Brazzaville, Congo Centre Hospitalier Universitaire de Brazzaville, BP 1846, Congo Correspondence should be addressed to Pembe Issamou Mayengue; pmayengue@yahoo.fr Received 10 March 2018; Accepted 25 July 2018; Published 2 September 2018 Academic Editor: Liwang Cui Copyright © 2018 PembeIssamou Mayengueetal. iTh sisan openaccess articledistributed underthe Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited. Background. In Republic of Congo, malaria diagnosis still widely relies on microscopy. We aimed to evaluate the performance of routine microscopy for malaria diagnosis at three different health centers in Brazzaville. Methods. A total of 259, 416, and 131 patients with clinical signs of uncomplicated malaria were enrolled at the Hop ˆ ital de Mfilou, Centre de Sant´eIntegr ´ e“ ´ MamanMbouale,” ´ and Laboratoire National de Sante´ Publique, respectively. Two thick blood smears were prepared for each patient, the first being examined by routine microscopists and the second by expert. Results.At the Hoˆpital de Mfilou, sensitivity was 62.1% and specificity was 67.3%. Positive and negative predictive values were 55.6% and 72.9%, respectively. At the Centre de SanteI ´ ntegr ´ e“ ´ Maman Mbouale,” ´ sensitivity was 94.2% and specificity was 33.6%. Positive and negative predictive values were 50% and 89.1%, respectively. At the Laboratoire National de Sante´ Publique, sensitivity and specificity were high with 91.7% and 94.9%, respectively. Positive and negative predictive values were 64.7% and 99.1%, respectively. Conclusion. eTh performance of routine malaria microscopy in Brazzaville remains inaccurate with large variations among different health centers. er Th efore, repeated training including supervision and evaluation would improve routine malaria diagnosis for better management of malaria in Brazzaville, the Republic of Congo. 1. Introduction Thirteen countries, mainly in sub-Saharan Africa, accounted for 76% of malaria cases and 75% deaths globally [1]. Malaria remains a public health problem in the majority In the Republic of Congo, where malaria is still the of countries in sub-Saharan Africa. Global morbidity and leading cause of attendance in health facilities, the high level mortality are estimated at 212 million malaria cases and of resistance of Plasmodium falciparum to chloroquine and 429,000 deaths attributable to malaria, respectively, in 2015, the inefficacy of sulphadoxine-pyrimethamine and amodi- with 70% of all malaria deaths occurring among children aquine either singly or in combination for the treatment under the age of ve fi . Approximately 90% of malaria cases of uncomplicated malaria have been well documented [2– and 92% of deaths occurred in the WHO African Region. 5].Thus, in 2006,the Republic of Congo has changed its 2 Malaria Research and Treatment antimalarial drug policy for treating uncomplicated malaria endemicity. Malaria infection is primarily due to P. falci- to artemisinin-combination therapies [6]. To avoid the threat parum. Two rainy seasons are observed each year with the of potential emergence of drug resistance, accurate diagnosis main one during the months of February to May and a short strategies for malaria, as well as, prompt case management one from October to December [12]. Instead of their location, are warranted. Therefore, the World Health Organization the malaria transmission variation, and their ability to receive (WHO) has recommended a parasitological test to all patients many patients from all socioeconomic status requiring a with suspected uncomplicated malaria before initiating anti- large number of microscopits, the CSI “Maman Mbouale” malarial treatment [7]. Laboratory conrfi mation of malaria and Hoˆpital de Mlfi ou have been also selected based on the with microscopy or a rapid diagnostic test (RDT) will help participation of 4 microscopists technicians from each center prevent unnecessary treatment with expensive artemisinin- in PARAMED training, who were also present in their centers combination therapies (ACTs) and the development of drug during the study. However, since the training was done at the LNSP, all miscroscopists from this laboratory were trained. resistance and will also allow rapid and correct management of other febrile illnesses [8–11]. The quality control of microscopy reading was performed at the laboratory of the Centre Hospitalier Universitaire de In the Republic of Congo, diagnosis of malaria is still Brazzaville (CHU), which is the big referral hospital with widely based on microscopy in the majority of health facil- a reference laboratory in Brazzaville. Expert reading was ities, and confirmatory testing rate with RDTs remains low. A performed by the head of the laboratory, who was also trainer study conducted in Brazzaville has shown that utility of RDTs in PARAMED, with the urge experience on parasitology and as a diagnostic tool was poor, due to lack of experience of blinded from the results obtained in all three study health staff and their lacking availability [12]. This observation has centers. been also supported by the WHO showing no use of RDT tests in 2013 and 2015 in the Republic of Congo [1], although 2.2. Ethical Clearance and Site Preparation. The study was microscopy has been the primary method and remains the approved by the institutional “Comited’Ethique de la gold standard for malaria diagnosis in clinical routine, with Recherche en Sciences de la Sante” ´ (N 032/CERSSA-2015). rather poor quality mainly at primary healthcare level, thus In the early project stage, rfi st, a meeting with the site actors requiring supervision and training activities. including the head of each laboratory and microscopists Over the last vfi e years the Ministry of Health has was organized for the purpose of presenting the project initiated the “PARAMED” project which aims at capacity objectives, methodology, and expected results, as well as to building of laboratory technicians was involved in routine obtain microscopists consent. Second, a meeting was held in malaria microscopy. First, individual microscopists from the various study sites, assuring the availability of material for various healthcare sites were selected and trained in malaria thick blood smears preparation during the study execution. light microsocpy. Subsequently, after their returning to their Also, training was provided ensuring the standardization of original health-facility laboratories they would know how thick blood preparation and parasite density assessments in to perform microscopy and spread relevant knowledge to all sites. their peers. However, the impacts of these trainings are not well documented. Consequently, little is known about 2.3. Study Population, Blood Samples, and Data Collection. the actual performance of malaria routine microscopy in From May 2015 to May 2016, patients with clinical signs of different healthcare level facilities in Brazzaville. us Th , the uncomplicated malaria, presenting at the laboratory of one of present study aimed at evaluating the performance of routine the three study sites, were invited to participate in this study. microscopy for malaria diagnosis at three different health Exclusion criteria comprised pregnancy, severe malaria, or centers in Brazzaville, the Republic of Congo. other severe illness as judged by the attending physician. A number of representative patients to be included each 2. Methods month, per week, and per day have been estimated by the statistician taking into account the proportion of malaria 2.1. Study Areas. Brazzaville, the political capital, hosts 38% reported in each health center, one year before starting (1 642 105 inhabitants) of the total population of the Republic thestudy. In sample sizecalculations, using a power of of Congo, estimated at 4 312 715 inhabitants. With the 80% and a significance level of 5%, the SCHWARZ method population expansion due to urbanization, Brazzaville is now yielded a minimum number of 310, 200, and 100 patients divided into nine districts: Bacongo, Makelekele, Poto-Poto, to be recruited at the CSI “Maman Mbouale,” ´ Hopi ˆ tal de Moungali, Ouenze, Talanga¨ı, Mfilou, Madibou, and Djiri. The Mlo fi u, and the LNSP, respectively. Recruited patients were present study was conducted in three centers: Centre de Sante´ randomly selected from Monday to Friday. At the CSI Integ ´ re´ (CSI) “Maman Mbouale” ´ located in the district of “Maman Mbouale,” ´ a minimum of 7 patients were recruited Talanga¨ı, in the north part of city, Hopi ˆ tal de Mlo fi u located per week, with at least one patient per day, whereas at the in the district of Mfilou, in the south part of the city, and the Hopi ˆ tal de Mlfi ou the minimum number was 5 patients per Laboratoire National de SanteP ´ ublique (LNSP), the national week, with one per day. For the LNSP, 3 patients were enough, reference laboratory located, in the center part of city, in the with one included on Monday, on Wednesday, and on Friday. district of Poto-Poto. After informed consent was obtained, records were made on Malaria transmission in the study areas varies from patient demographics, fever or history of fever in the last low and moderate to intense with meso-hyper-to perennial 48 hours, other signs of malaria, provenance, and previous Malaria Research and Treatment 3 Table 1: Characteristics of patients. Characteristics Hoˆpital de Mfilou CSI “Maman Mbouale” ´ LNSP Total numbers 259 416 131 Gender (F/M) 131/126 207/203 68/63 Median age (years) 26 (0.5-84) 10 (0.5-75) 42 (11 - 74) Groups of age (n, %) < 5 years 38 (14.8) 99 (23.8) 0 (0.0) ≥ 5 years 219 (85.2) 317 (76.2) 131(100) antimalarial drugs intake used of bed net treated. The axillary 203 being female and male, respectively, and 99 patients were temperature was taken for fever confirmation. children under 5 years of age (Table 1). At each study site, two thick blood smears were prepared Among 131 patients recruited at the LNSP, 68 and 63 were for each patient, with one being read immediately to inform female and male, respectively, and all of them had an age the patient of the respective result. Before reading, thick above 5 years. blood smears were dried and stained with 10% Giemsa 3.2. Slide Positivity and Quality Control of Three Health Cen- solution (Sigma Chemical, Sigma Aldrich ChemieGmbh, ters in Brazzaville. Of the 259 slides collected at the Hopi ˆ tal Taufkirchen, Germany) in pH 7.2, for approximately 10 min. de Mlo fi u, 115 (44.4%) were reported positive by routine The stain was gently washed away by adding drops of clean microscopists, whereas 103 (39.8%) slides were positive by the water and the slide was completely dried before examination. quality control expert. Of the 144 slides that were negative Thick blood smears were assessed by micrsocopists until 200 by the routine microscopy, 39 were identified positive by leucocytes had been counted. Parasite density was calculated for each patient assuming an average of 8000 leucocytes the quality control expert. False positive proportions (51/115; 44.3%) and false negative proportions (39/144; 27.1%) were per𝜇 l of blood using the proposed method of the WHO not statistically significant ( p = 0.240). u Th s, the performance [13]. Individual diagnostic result was given to each patient at the Hopital de Mfilou in terms of sensitivity and specicfi ity and advised to meet the prescribers for possible antimalarial was 62.1% (CI: 52.5% -70.9%) and 67.3% (CI: 59.6% -74.2%), chemotherapy. The second uncolored thick blood smear was respectively. Positive and negative predictive values were transferred to the CHU for microscopy quality control. At the 55.6% (CI 46.5% - 64.4%) and 72.9% (CI 65.1% - 79.5%), CHU, also the Giemsa for sample staining was provided. respectively (Table 2). With regard to CSI “Maman Mbouale,” ´ 324 (77.9%) slides 2.4. Data Analysis. Data were entered and verified in Microsoft Excel (Microsoft Corp., Seattle, USA) and validated were reported positive by routine microscopists, while only 172 (41.3%) slides were positive by the quality control expert. in EpiInfo for Windows version 3.5.1. Data were analyzed using the SPSS 16.0 for Windows (Inc., Chicago, USA). Age Out of the 92 slides that were rated negative by the routine was expressed as median (range), while categorical variables microscopists, 10 were revealed positive by the quality control expert (Table 2). At this site, routine microscopists reported were expressed as percentages. Correlation coefficients were used to compare parasites densities from health centers and signicfi antly more false positive (162/324; 50%) than false quality control expert. The Bayesian theory was used to negative (10/92; 10.8%) with p< 0.001. Routine microscopists had a performance of high sensitivity at 94.2% (CI: 89. 6%- estimate the sensitivity, specificity, positive predictive value, and negative predictive value as described by Joseph et al. 96.8%), but of low specificity at 33.6% (CI: 29.0%- 38.7%). [14]. 95% confidences intervals (95% CI) were generated Positive and negative predictive values were 50% (CI: 44. 6%- 55.4%) and 89.1 (81.1%- 93.9%), respectively. for parameters mentioned above. Statistical significance of performance differences between health center microscopists At the LNSP, 17 (12.98%) out of 131 slides were reported positive by the routine microscopists, whereas 12 (9.2%) and quality control expert was evaluated using McNemar’s test. Statistical significance level was set at p <0.05. were rated positive by the quality control expert. Out of the 114 slides that were rated negative by routine microscopists, only one was revealed positive by the quality control expert 3. Results (Table2).However,themicroscopistsatthissitereportedalso 3.1. Characteristics of Febrile Patients with Symptoms of more false positives (6/17; 35.3%) than false negatives (1/114; Malaria. A total of 259, 416, and 131 patients with suspected 0.9%) with p< 0.001. malaria were enrolled at the Hoˆpital de Mfilou, CSI “Maman Both sensitivity and specificity were high at this study site Mbouale,” ´ and the LNSP, respectively. with 91.7% (64.6% -98.5%) and 94.9% (89.4%-97.7%), respec- Out of 259 patients enrolled at the Hopital de Mlo fi u, tively. Positive and negative predictive values were 64.7% (CI: gender and age were recorded for 257 of them, with 131 being 41.3%- 82.7%) and 99.1 (95.2%- 99.8%), respectively (Table 2). female and 126 male. Thirty-eight patients had an age below 5 ´ 3.3. Identification of Plasmodium Species. P. falciparum was years. With regards to CSI “Maman Mbouale,” out of the 416 recruited patients, 410 had records on gender with 207 and the only species identified in all positive slides confirmed by 4 Malaria Research and Treatment Table 2: Slide positivity and quality control of three health centers in Brazzaville. Site Quality control Sensitivity Specificity PPV NPV Positive Negative % (95%CI) %(95%CI) %(95%CI) %(95% CI) Positive 64 51 Hˆopital de Mfilou 62.1 (52.5-70.9) 67.3 (59.6-74.2) 55.6 (46.5-64.4) 72.9 (65.1-79.5) Negative 39 105 Positive 162 162 CSI Maman Mbouale´ 94.2 (89.6-96.8) 33.6 (29.0-38.7) 50.0 (44.6-55.4) 89.1 (81.1-93.9) Negative 10 82 Positive 11 6 LNSP 91.7 (64.6-98.5) 94.9 (89.4 -97.7) 64.7 (41.3-82.7) 99.1 (95.2-99.8) Negative 1 113 Health Center Mfilou Maman Mboualé LNSP 6,00 r= 0.44;p-value= 0.001 r= 0.55;p-value= 0.048 r= 0.39;p-value= 0.009 5,00 4,00 3,00 2,00 1,00 1,00 2,00 3,00 4,00 5,00 6,00 1,00 2,00 3,00 4,00 5,00 6,00 1,00 2,00 3,00 4,00 5,00 6,00 Log10 parasite density from Health center Figure 1: Comparison of parasites densities between dieff rent sites and control. the quality control expert. However, P. malariae was detected level. This performance may be explained by the fact that the by routine microscopists in one and two slides of 64 and LNSP is the national site for microscopist training and as 162 P. falciparum confirmed positives slides from H opi ˆ tal de such having qualified trainers and technicians who are also Mfilou and CSI “Maman Mboual e,” respectively. At the LNSP, working in parasitological diagnostic routine. Other factors out of 11 conrfi med positive slides, one slide was positive for that contribute to this performance were favorable quality of Plasmodium ovale by the routine microscopists. equipment and the relative low number of slides prepared ranging from one to ten, related to low malaria transmission 3.4. Evaluation of Parasite Densities at Different Sites. All as it was observed in Kenya [15]. However, the relative differ- ence on estimation of parasite density as well as the trend of three study sites assessed parasite density different compar- atively to the quality control expert (Figure 1). However, species misclassification requires training and improvement knowledge on the parasite for better management of malaria the differences from the quality control expert were more pronounced at the CSI “Maman Mbouale” ´ (p= 0.001) and cases at this site. Hopi ˆ tal de Mlo fi u ( p= 0.009) compared to that with the LNSP In contrast to LNSP, different performances have been found at the Hoˆpital de Mlfi ou and at the CSI “Maman (p= 0.048). Mbouale.” The lower sensitivity and specificity of routine microscopy were observed at the Hop ˆ ital de Mlo fi u and the 4. Discussion CSI “Maman Mbouale.” ´ Both over- and underdiagnoses of This study demonstrates the inaccurate performance of malaria are common in malaria-endemic African countries routine microscopy in the three health centers compared such as Tanzania, Uganda, Burkina Faso, Kenya, and Cote ˆ d’Ivoire [10, 16–19]. These lower performances have potential to expert microscopy, indicating poor performance, where large quantities of blood smear examination and permanent clinical implications, with low sensitivity implying many involvement of several microscopists were necessary. underdiagnosed malaria cases with no safe and appropri- Both sensitivity and specicfi ity were signicfi antly higher ate treatment. Low specificity leads to overdiagnosis with at the LNSP, which is the reference laboratory at the national overtreatment and unnecessary prescription of antimalarials, Log10 parasite density from Control Malaria Research and Treatment 5 as well as underdiagnosis and undertreatment of real causes seasons. Additionally, the lack of distribution of malaria of actual symptoms [10, 15, 20, 21]. Plasmodial species bed nets treated since 2013 may have an impact on the were also confused in these two sites and parasite density increasing incidence of the disease. The different situation observed at the LNSP is probably due to a small number estimation was largely different from the expert’s assessment. The observed phenomena are a worrying situation in these of suspected malaria patients presenting at the laboratory two health centers, receiving a high amount of suspected with the majority of patients being adults. Extensive repeated studies in different sites including other departments are malaria patients of all age groups, including children under 5 years of age, the population most vulnerable to severe and needed to provide the actual picture of malaria prevalence in potentially fatal malaria complications. Additionally, due to the Republic of Congo. the lack of recent entomological data in these study sites, malaria burdenismostly estimated based onthe proportion 5. Conclusion of diagnosed cases. Therefore, misdiagnosis and resulting biased interpretations of in reality-different epidemiological This study showed that the performance of routine malaria patterns on malaria may lead to biased reports on the local microscopy in Brazzaville remains substandard between dif- burden of malaria and to biased evaluations of various ferent health centers. Therefore, repeated training including malaria control intervention programs. Several factors may supervision and evaluation at the national level as well as justify the poor performance of routine microscopy in these implementation and use of RDTs would improve routine sites including the large number of thick blood smears to read malaria diagnosis for better management and control of in daily routine, lack of skilled microscopists, supervision, malaria in Brazzaville. and maintenance of infrastructure mainly at the CSI “Maman Mbouale,” ´ and lack of regular quality control. Although Data Availability four selected microscopists from each of these two sites had recently participated in the PARAMED training, the The data used to support the findings of this study are daily work in these two centers was also done by several available from the corresponding author upon request. nonselected microscopists. However, the obtained results are an indicator of the lack of supervision after training as well as Conflicts of Interest low impactofspreadofacquired knowledge to others micro- scopists working permanently at these sites. Training can The authors declare that they have no conflicts of interest. improve the capacity of individual microscopists; but, when they return to respective laboratories, they often face many Authors’ Contributions challenges such as equipment and heavy workloads. These challenges can contribute to the marginal improvements in Pembe Issamou Mayengue designed and coordinated eld fi the performance observed after training [15]. study, analyzed the data, and wrote the draft of the article. The PARAMED training lasted only 1 month with many Dezi Kouhounina Batsimba, Roch Fabien Niama, Lucette modules. The short duration of training of microscopists Macosso, Brice Pembet Singana, Igor Louzolo, and Nadia may also impact acquired knowledge. Therefore, perma- Claricelle Bongolo supervised eld fi samples and data col- nent training including laboratory supervision as well as lection; Louis Re´gis Dossou-Yovo participated in patients improvement of infrastructure quality and implementation recruitment; Ger ´ il Sekangue Obili supervised quality control. of internal and/or external quality control are needed for All authors read and approved the final version and the final achieving improvement of malaria diagnosis by microscopy manuscript. as observed in Angola [22]. The other alternative should be the introduction of RDTs in malaria diagnosis procedure and Acknowledgments their systematic use in addition to microscopy in these sites. However study conducted by Ntoumi et al. [12] has shown The authors are grateful to all patients and all microscopists that the RDTs are not largely use mainly in Brazzaville. working in their three sites, who participated in this study. The responsibility of the National Malaria Control Pro- They thank also Professor Michael Ramharter and Doctor gram in the Republic of Congo may be in increasing training Johannes Mischlinger for carefully reading of manuscript. on the use of RDTs and implementing their systematic use in The study was supported by the “Laboratoire National de the algorithm of malaria diagnosis in the country. Sante´ Publique” and the World Academy of Sciences (RGA By considering the proportion of positive slides deter- no.16-040 RG/BIO/AF/AC I- FR3240293321). mined by the expert reader, the prevalence of malaria was estimated at 39.77%, 41.34%, and 9.16% at the Hopi ˆ tal de References Mlfi ou, the CSI “Maman Mboual e,” ´ and the LNSP, respec- tively. 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