Bogoch, I.

Marti, H.

N'Goran, E.

Raso, G.

Utzinger, J.

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Eur J Clin Microbiol Infect Dis (2006) 25: 344–347DOI 10.1007/s10096-006-0135-xBRIEF REPORTI. I. Bogoch . G. Raso . E. K. N’Goran .H. P. Marti . J. UtzingerDifferences in microscopic diagnosis of helminths and intestinalprotozoa among diagnostic centresPublished online: 25 April 2006# Springer-Verlag 2006Sub-specialized tropical medicine laboratories are essen-tial for the accurate diagnosis of many tropical infectionsthat may not be correctly diagnosed in conventionallaboratories. However, processing and examination of‘identical’ biofluid specimens such as blood, stool andurine at different laboratories often results in considerablediagnostic discrepancies. For example, interlaboratorydifferences have been documented in the microscopicexamination of malaria slides [1], and discrepancies havebeen observed when comparing different rapid tests formalaria diagnosis [2]. The aim of this study was tocompare the results obtained at sub-specialized tropicalmedicine laboratories for the microscopic diagnosis ofhelminths and intestinal protozoa in faecal samplesobtained from a rural community in an endemic area ofsub-Saharan Africa. The study compared two Europeantropical disease diagnostic centres with each other and oneof the two European centres with a West African centre.Our study was designed as a cross-sectional community-based survey and was carried out in May 2002 in thevillage of Zouatta II, western Côte d’Ivoire. This village isendemic for Schistosoma mansoni, soil-transmitted hel-minths (Ascaris lumbricoides, hookworms and Trichuristrichiura) and intestinal protozoa [3]. Polyparasitism isextremely common [3, 4]. Details of the study area, as wellas field and laboratory procedures, have been presentedelsewhere [3]. In brief, approval of the study was obtainedfrom the institutional review boards of the Centre Suisse deRecherches Scientifiques (Abidjan, Côte d’Ivoire) and theSwiss Tropical Institute (Basel, Switzerland). The studyreceived ethical clearance from the Ministry of PublicHealth in Côte d’Ivoire and permission was also granted tous by local village authorities.Three consecutive morning stool specimens werecollected over a 3-day period from 561 randomly selectedcommunity members aged 5 days to 91 years. Specimenswere transferred to a laboratory in the nearby district townand processed the same day. From each specimen, a 42 mgKato–Katz thick smear was prepared according to astandard procedure [5]. The slides were allowed to clearfor 30–45 min prior to examination under a light micro-scope by experienced laboratory technicians. Specimenswere analysed for S. mansoni and soil-transmittedhelminths, with the numbers of eggs counted and recordedfor each species separately. Individuals who tested positivefor S. mansoni and/or soil-transmitted helminths weretreated with a single oral dose of 40 mg/kg praziquanteland/or 400 mg albendazole, respectively. In addition, 1–2 gof each stool specimen was conserved in a sodium acetate-acetic acid-formalin (SAF) solution on the same day ofcollection for future processing 6–12 months later at theother diagnostic centres.I. I. BogochFaculty of Medicine, University of Toronto,Toronto, M5S 1A8, Canadae-mail: ibogoch@hotmail.comG. Raso . J. Utzinger (*)Department of Public Health and Epidemiology,Swiss Tropical Institute,P.O. Box, 4002 Basel, Switzerlande-mail: juerg.utzinger@unibas.chTel.: +41-61-2848129Fax: +41-61-2848105H. P. MartiDiagnostic Centre, Swiss Tropical Institute,P.O. Box, 4002 Basel, Switzerlande-mail: hanspeter.marti@unibas.chG. RasoMolecular Parasitology Laboratory,Queensland Institute of Medical Research,Brisbane, Queensland, 4006, Australiae-mail: giovanna.raso@qimr.edu.auE. K. N’GoranCentre Suisse de Recherches Scientifiques,01 BP 1303, Abidjan 01, Côte d’Ivoiree-mail: eliezerngoran@yahoo.frE. K. N’GoranUFR Biosciences, Université d’Abidjan-Cocody,22 PB 770, Abidjan 22, Côte d’IvoireTwo interlaboratory comparisons were performed. Thefirst comparison was made between two European diag-nostic centres sub-specialized in tropical diseases (A and B).Fifty randomly selected SAF-conserved stool specimensfrom collection day 2 were shaken and then split into twoequal parts; one half was examined by European centre Aand the other half was examined by European centre B. Twospecimens were discarded because the participants hadprovided less than three stool samples for Kato–Katzprocessing, resulting in a sample size of 48.The second comparison was made between Europeancentre A and a West African diagnostic centre. A total of 78randomly selected SAF-preserved samples were compared;centre A analysed samples obtained from participants oncollection day 1 or 2, while the West African diagnosticcentre analysed corresponding stool samples obtained fromthe same participants on collection day 3. In these threelaboratories, stool specimens were processed using aformol-ether concentrate, according to the procedureoutlined by Allen and Ridley [6], and analysed for thepresence of helminth eggs and intestinal protozoa under alight microscope by experienced laboratory technicians.Data were first entered into Microsoft Excel and thentransferred to EpiInfo version 6.04 (Centre for DiseaseControl and Prevention, Atlanta, GA, USA) for statis-tical analysis. The results obtained for the individualSAF-preserved stool specimens examined at Europeancentre A and European centre B were compared againstthe laboratory gold standard, which was the results of thethree locally performed Kato–Katz thick smears for thecorresponding participants plus the collective resultsobtained by European centers A and B for the 48 SAF-preserved specimens. This laboratory gold standard wasdevised to maximize sensitivity values, with the under-lying rationale being that an egg detected at any level ofprocessing (e.g. ‘on the spot’ by the Kato–Katz tech-nique or in specialized European laboratories) shouldcount as a true-positive result. Negative predictive values(NPV) were also calculated; however, specificity andTable 1 Interlaboratory differences in microscopic examination of faecal samples for helminths and intestinal protozoa between twoEuropean tropical disease diagnostic centres A and B (n=48) and between European centre A and a West African centre (n=78)Parasite Number (%) Kappa coefficient Number (%) Kappa coefficientEuropeancentre AEuropeancentre BDifference κ (SE) pvalueEuropeancentre AWest AfricancentreDifference κ (SE) pvalueSchistosoma mansoni 19 (38) 5 (10) 14 (28) 0.21(0.10)0.02 24 (31) 9 (12) 15 (19) 0.16(0.09)0.04Soil-transmittedhelminthsHookworm 18 (36) 12 (24) 6 (12) 0.43(0.14)<0.01 31 (40) 15 (19) 16 (21) 0.18(1.10)0.03Ascaris lumbricoides 0 0 0 NA 0 0 0 NATrichuris trichiura 0 0 0 NA 0 1 (1) 1 (1) 0.00(0.07)0.50Intestinal protozoaBlastocystis hominis 29 (58) 38 (76) 9 (18) −0.00(0.13)0.51 38 (49) 7 (9) 31 (40) 0.08(0.07)0.10Chilomastix mesnili 7 (14) 0 7 (14) NA 9 (12) 7 (9) 2 (3) 0.44(0.11)<0.01Endolimax nana 10 (20) 2 (4) 8 (16) 0.29(0.10)<0.01 12 (15) 15 (19) 3 (4) −0.03(0.11)0.60Entamoeba coli 35 (70) 34 (68) 1 (2) 0.76(0.14)<0.01 50 (64) 34 (44) 16 (20) 0.21(0.10)0.02Entamoebahartmanni16 (32) 0 16 (32) NA 20 (26) 2 (3) 18 (23) −0.05(0.06)0.80Entamoeba histolytica/E. dispar18 (36) 2 (4) 16 (32) 0.13(0.72)0.03 25 (33) 10 (13) 15 (20) 0.33(0.10)<0.01Giardia duodenalis 6 (12) 5 (10) 1 (2) 0.69(0.14)<0.01 7 (9) 7 (9) 0 0.37(0.11)<0.01Iodamoeba bütschlii 9 (18) 8 (16) 1 (2) 0.36(0.14)<0.01 16 (21) 2 (3) 14 (18) −0.05(0.07)0.77SE standard error, NA not applicable345positive predictive values (PPV) could not be determinedwith this approach. Sensitivity and NPV were alsoestimated using the mathematical model of Marti andKoella [7]. This model relates the number of stoolsamples found to be positive for ova and parasites tofalse-negative results and is an accurate method ofpredicting sensitivity based on the number of stoolspecimens provided. Positivity rates were calculated asthe number of positive diagnoses per number of samplesanalysed and compared between the two Europeancentres and between European centre A and the WestAfrican centre. A kappa coefficient was calculated todetermine the degree of agreement between diagnoses atdifferent centres.Frank diagnostic discrepancies became apparent whencomparing the results from the three Kato–Katz thicksmears plus the SAF-preserved specimens analysed atEuropean centres A and B (our laboratory gold standard)with those obtained for each individual SAF-preservedspecimen processed at centre A and at centre B. Forexample, the diagnostic sensitivity values for S. mansoni atcentres A and B were 71% (95% confidence interval [CI]49–87%) and 17% (95% CI 6–38%), respectively. Forhookworm, the respective sensitivity values for centres Aand B were 75% (95% CI 53–98%) and 50% (95% CI 30–70%). Using a mathematical model to evaluate the resultsof microscopic examination of the three Kato–Katz thicksmears [7], the overall diagnostic sensitivity and NPV forhookworms was 44 and 43%, respectively, while thediagnostic sensitivity and NPV for S. mansoni was 89 and91%, respectively.Major interlaboratory differences became apparent whencomparing the rates of detection of S. mansoni, soil-transmitted helminths and intestinal protozoa in SAF-preserved specimens between European centres A and Band between European centre A and the West Africancentre. Table 1 summarizes the positivity rates and kappaagreement coefficients for the different helminths andintestinal protozoa investigated.This study revealed considerable diagnostic discrepan-cies among laboratories that specialize in the diagnosis oftropical infections. In particular, European diagnosticcentre B performed significantly worse than centre A,which warrants urgent attention. However, the testsemployed in our study do not reflect the entire spectrumof diagnostic approaches that are used for returningtravelers. Many travelers returning from endemic areaswill be screened with serologic tests such as the enzyme-linked immunosorbent assay (ELISA) or the Falcon assayscreening test (FAST) [8, 9], the Kato–Katz thick smear offresh stool [5], or a combination of different stool-examination methods [10]. Such combinations of testsenhance the diagnostic performance and result in a highersensitivity and specificity. Newer diagnostic tests, such asthe polymerase chain reaction (PCR) for S. mansoni, alsohold promise for improving future diagnoses [11].Although many laboratories use SAF-preserved stoolspecimens, they will often examine multiple specimensfrom each patient in order to increase the diagnosticsensitivity. Microscopic examination of one SAF-pre-served stool specimen is not the optimal diagnosticapproach due to the irregular excretion pattern of mostintestinal parasites and the difficulty of detecting largeeggs, such as S. mansoni, which are frequently missed [12].This approach consequently leads to false-negative resultsand an underestimation of the true prevalence. Still,microscopic examination of SAF-preserved stool samplesis more sensitive than direct examination of fresh faecalsmears in the diagnosis of intestinal protozoa [13].Moreover, SAF concentration is used by many laboratoriesas a screening method for intestinal parasites. In case ofclinical suspicion of a specific helminth infection, moresensitive methods should be employed (e.g. culture forStrongyloides stercoralis) [14].In this study, we were surprised to see such a largedegree of variance in the positivity rates reported bylaboratories analysing the same SAF-preserved stoolspecimens. However, the discrepancy in results obtainedby the West African diagnostic centre and European centreA could be explained by the fact that these centres testedstool samples collected from the same individuals ondifferent days, since day-to-day sample variation has beendocumented for S. mansoni and hookworms [15]. Ingeneral, diagnostic discrepancies among laboratoriesspecialized in tropical parasitology is likely multifactorial,involving such aspects as the level of training of laboratorypersonnel, malfunctioning or dated laboratory equipment,and human error. A multifaceted approach is thus necessaryto decrease the incidence of interlaboratory discrepanciesand it typically involves three general initiatives. (a)Institutions that train laboratory personnel must be up-to-date and adhere to the highest standards of care, althoughthis may be difficult in resource-poor settings, such as sub-Saharan Africa. (b) Laboratories should utilize a series ofstandardized methodologies designed to reduce errors andto facilitate communication and collaboration amongdiagnostic centres. (c) Systems that reduce human errorshould be introduced to the laboratory environment [16].Such error-reducing systems are often technologicallyoriented, but they can also be as simple as having multiplepeople confirm that transcribed information is correct, orusing a series of standardized laboratory procedures toreduce the number of false-positive and false-negativeresults. Such a combined approach will decrease the highlevel of discrepancies in laboratory analyses.Acknowledgements This investigation received financial supportfrom the University of Toronto Medical Alumni Association for anInternational Summer Research Scholarship to I.I.B, the RocheResearch Foundation for partial funding of a PhD fellowship to G.R,and the Swiss National Science Foundation supporting G.R (projectno. PBBSB-109011) and J.U. (project no. PPOOB-102883).References1. Hemme F, Gay F (1998) Internal quality control of the malariamicroscopy diagnosis for 10 laboratories on the Thai-Myanmarborder. Southeast Asian J Trop Med Public Health 29:529–5363462. De Monbrison F, Gerome P, Chaulet JF, Wallon M, Picot S,Peyron F (2004) Comparative diagnostic performance of twocommercial rapid tests for malaria in a non-endemic area. Eur JClin Microbiol Infect Dis 23:784–7863. Raso G, Luginbühl A, Adjoua CA, Tian-Bi NT, Silué KD,Matthys B, Vounatsou P, Wang Y, Dumas ME, Holmes E,Singer BH, Tanner M, N’Goran EK, Utzinger J (2004) Multipleparasite infections and their relationship to self-reportedmorbidity in a community of rural Côte d’Ivoire. Int JEpidemiol 33:1092–11024. Keiser J, N’Goran EK, Traoré M, Lohourignon KL, Singer BH,Lengeler C, Tanner M, Utzinger J (2002) Polyparasitism withSchistosoma mansoni, geohelminths, and intestinal protozoa inrural Côte d’Ivoire. J Parasitol 88:461–4665. Katz N, Chaves A, Pellegrino J (1972) A simple device forquantitative stool thick-smear technique in schistosomiasismansoni. Rev Inst Med Trop Sao Paulo 14:397–4006. Allen AVH, Ridley DS (1970) Further observations on theformol-ether concentration technique for faecal parasites. J ClinPathol 23:545–5467. Marti H, Koella JC (1993) Multiple stool examinations for ovaand parasites and rate of false-negative results. J Clin Microbiol31:3044–30458. Tsang VC, Wilkins PP (1997) Immunodiagnosis of schistoso-miasis. Immunol Invest 26:175–1889. Al-Sherbiny MM, Osman AM, Hancock K, Deelder AM,Tsang VC (1999) Application of immunodiagnostic assays:detection of antibodies and circulating antigens in humanschistosomiasis and correlation with clinical findings. Am JTrop Med Hyg 60:960–96610. van Gool T, Weijts R, Lommerse E, Mank TG (2003) TripleFaeces Test: an effective tool for detection of intestinal parasitesin routine clinical practice. Eur J Clin Microbiol Infect Dis22:284–29011. Pontes LA, Oliveira MC, Katz N, Dias-Neto E, Rabello A(2003) Comparison of a polymerase chain reaction and theKato–Katz technique for diagnosing infection with Schistoso-ma mansoni. Am J Trop Med Hyg 68:652–65612. Yang J, Scholten T (1977) A fixative for intestinal parasitespermitting the use of concentration and permanent stainingprocedures. Am J Clin Pathol 67:300–30413. Mank TG, Zaat JO, Blotkamp J, Polderman AM (1995)Comparison of fresh versus sodium acetate acetic acid formalinpreserved stool specimens for diagnosis of intestinal protozoalinfections. Eur J Clin Microbiol Infect Dis 14:1076–108114. Koga K, Kasuya S, Khamboonruang C, Sukhavat K, Ieda M,Takatsuka N, Kita K, Ohtono H (1991) A modified agar platemethod for detection of Strongyloides stercoralis. Am J TropMed Hyg 45:518–52115. Booth M, Vounatsou P, N’Goran EK, Tanner M, Utzinger J(2003) The influence of sampling effort and the performanceof the Kato–Katz technique in diagnosing Schistosomamansoni and hookworm co-infections in rural Côte d’Ivoire.Parasitology 127:525–53116. Mutter M (2003) One hospital’s journey toward reducingmedication errors. Jt Comm J Qual Saf 29:279–288347

Differences in microscopic diagnosis of helminths and intestinal protozoa among diagnostic centres

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Differences in microscopic diagnosis of helminths and intestinal protozoa among diagnostic centres

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