Efficiency of Direct Microscopy of Stool Samples Using an Antigen-Specific Adhesin Test for Entamoeba Histolytica

Abstract

Background: E. histolytica is among the common causes of acute gastroenteritis. The pathogenic species E. histolytica and the nonpathogenic species E. dispar cannot be morphologically differentiated, although correct identification of these protozoans is important for treatment and public health. In many laboratories, the screening of leukocytes, erythrocytes, amoebic cysts, trophozoites and parasite eggs is performed using Native-Lugol’s iodine for pre-diagnosis. Aims: In this study, we aimed to investigate the frequency of E. histolytica in stool samples collected from 788 patients residing in the Anatolian region of İstanbul who presented with gastrointestinal complaints. We used the information obtained to evaluate the effectiveness of microscopic examinations when used in combination with the E. histolytica adhesin antigen test. Study Design: Retrospective cross-sectional study Methods: Preparations of stool samples stained with Native-Lugol’s iodine were evaluated using the E. histolytica adhesin test and examined using standard light microscopy at ×40 magnification. Pearson’s Chi-square and Fisher’s exact tests were used for statistical analysis. Logistic regression analysis was used for multivariate analysis. Results: Of 788 samples, 38 (4.8%) were positive for E. histolytica adhesin antigens. When evaluated together with the presences of erythrocytes, leukocytes, cysts, and trophozoites, respectively, using logistic regression analysis, leukocyte positivity was significantly higher. The odds ratio of leukocyte positivity increased adhesin test-positivity by 2,530-fold (95% CI=1.01–6.330). Adhesin test-positivity was significant (p=0.047). Conclusion: In line with these findings, the consistency between the presence of cysts and erythrocytes and adhesin test-positivity was found to be highly significant, but that of higher levels of leukocytes was found to be discordant. It was concluded that leukocytes and trophozoites were easily misjudged using direct microscopy. Although microscopic examination of samples stained with Native-Lugol’s iodine is a cheap and simple method, the confusion of trophozoites with leukocytes may direct the clinician toward an incorrect pre-diagnosis. Because trichrome staining is difficult and time consuming, and results may vary depending on the technician, this method is not preferred in most laboratories. Therefore, an enzyme-linked immunosorbent assay method, which is a more advanced method than polymerase chain reaction, should be used to distinguish between E. histolytica and E. dispar in order to achieve an accurate diagnosis.

Keywords

• Adhesin test, Microscopic examination, Entamoeba histolytica

References

1. 1. Ximenez C, Moran P, Rojas L, Valadez A, Gomez A. Reassessment of the epidemiology of amebiasis: State of the art. Infect Genet Evol 2009;9:1023-32. [CrossRef]
2. 2. WHO. Amoebiasis. Wkly Epidemiol Rec 1997;72:97-100
3. 3. el-hamshary EM, el-Shewy KA, Hegazy MM, Zakaria H. Diagnostic potentials of copro antijen detection based ELISA, compored to microscopy in intestinal amobiasis. J Egypt Soc Parasitol 2004;34:601-10.
4. 4. Fotedar R, Stark D, Beebe N, Marriott D, Ellis J, Harkness J. Laboratory diagnostic techniques for Entamoeba species. Clin Microbiol Rev 2007;20:511-32. [CrossRef]
5. 5. Tuncay S, İnceboz T, Over L, Yalçın G, Usluca S, Şahin S et al. The evaluation of the techniques used for of Entamoeba histolytica in stool specimens Turkiye Parazitol Derg 2007;31:188-93.
6. 6. Yüksel P, Çelik DG, Güngördü Z, Ziver T, İzmirli S, Yakar H et al. Entamoeba histolytica Lectin Antigen from stool specimens by ELISA method: 3 Year Data. Klimik Dergisi 2011;24:150-3.
7. 7. Nath J, Ghosh SK, Singha B, Paul J. Molecular epidemiology of Amoebiasis: A cross-sectional study among North East Indian population. PLoS Negl Trop Dis 2015 Dec 3;9:e0004225. [CrossRef]
8. 8. Anuar TS, Al-Mekhlafi HM, Abdul Ghani MK, Abu Bakar E, Azreen SN, Salleh FM et al. Molecular epidemiology of amoebiasis in Malaysia: Highlighting the different risk factors of Entamoeba histolytica and Entamoeba dispar infections among Orang Asli communities. Int J Parasitol 2012;42:1165-75. [CrossRef]

9. 9. Haque R, Faruque AS, Hahn P, Lyerly D, Petri WA Jr. Entamoeba histolytica and Entamoeba dispar infection in children in Bangladesh. J Infect Dis 1997;175:734-6. [CrossRef]
10. 10. Hegazi MA, Patel TA, El-Deek BS. Prevalance and characters of Entamoeba histolytica infection in Saudi infants and children admitted with diarrhea at 2 main hospitals at South Jeddah:a re-emerging serious infection with unusual presentation. Braz J Infect Dis 2013;17:32-40. [CrossRef]
11. 11. Tasawar Z, Kausar S, Lashari MH. Prevalance of Entamoeba histolytica in Humans. Pak J Pharm Sci 2010;23:344-8.
12. 12. Yıldırım D, Hasbek M, Nur N. Analysis of intestinal ameobiasis in patients with dairrhea by adhesin antigen test and direct microscopy. Turkiye Parazitol Derg 2014;38:155-8. [CrossRef]
13. 13. Alver O, Topaç T, Töre O. Evaluation of two methods (Nativelugol preparation and enzyme -linked immunosorbent assay) for detection of Entamoeba histolytica in stool Ssamples. Turkiye Parazitol Derg 2015;39:185-9. [CrossRef]
14. 14. Braga LL, Mendonca Y, Paiva CA, Sales A, Cavalcante AL, Mann BJ. Seropositivity for and intestinal colonization with Entamoeba histolytica and Entamoeba dispar in individuals in northeasten Brazil. J Clin Microbiol 1998;36:3044-5.
15. 15. Taylan Özkan 2005. Rutin Dışkı Bakısına Alternatif: Antijen Tarama Yöntemleri.14.Ulusal Parazitoloji Kongresi.
16. 16. Delialioglu N, Aslan G, Ozturk C, Ozturhan H, Sen S, Emekdas G. Detection of Entamoeba histolytica antigen in stool samples in Mersin, Turkey. J Parasitol 2008;94:530-2. [CrossRef]