The Differential detection of some Entamoeba spp. in fresh stool specimens by PCR
Abstract
The current study aimed to collect stool samples from patients visiting Sharqat General Hospital and primary healthcare centers in Salah ad-Din Governorate to diagnose some the species of Entamoeba parasite using the molecular PCR technique, and by designing specialized primers to detect parasite DNA. The study spanned a period of time extending from the beginning of November 2023 to the end of March 2024. Stool samples were collected from individuals infected with amoebic parasites after they were clinically diagnosed by doctors based on their symptoms. Patient data was obtained and classified to determine if they were infected with parasites. Microscopic analyses were carried out on the stool in order to determine if individuals were infected with Entamoeba. PCR was utilized in molecular studies and every case identified using microscopy was confirmed by the molecular assay. We conclude the PCR technique is very accurate in diagnosis species of organisms, and this was observed through what was obtained from the current study.
Downloads
References
Abuseir, S. A. (2023). Systematic review of frequency and geographic distribution of water-borne parasites in the Middle East and North Africa. Eastern Mediterranean Health Journal, 29(2), 151–161. https://doi.org/10.26719/emhj.23.016
Al-Agha, R., & Teodorescu, I. (2000). Intestinal parasitic infestation and anemia in primary school children in Gaza Governorat Palestine. Roumanian Archives of Microbiology and Immunology, 53, 131–143.
Alelign, A., Mulualem, N., & Tekeste, Z. (2024). Prevalence of intestinal parasitic infections and associated risk factors among patients attending Debarq Primary Hospital, northwest Ethiopia. PLOS ONE, 19(3), e0298767. https://doi.org/10.1371/journal.pone.0298767
Amar, C. F., Dear, P. H., Pedraza-Diaz, S., Looker, N., Linnane, E., & McLauchlin, J. (2002). Sensitive PCR-restriction fragment length polymorphism assay for detection and genotyping of Giardia duodenalis in human feces. Journal of Clinical Microbiology, 40(2), 446–452. https://doi.org/10.1128/jcm.40.2.446-452.2002
Beavogui, A. H., Cherif, M. S., Camara, B. S., Delamou, A., Kolie, D., Cissé, A., & Djimdé, A. (2021). Prevalence of parasitic infections in children of Boke, Guinea. The Journal of Parasitology, 107(5), 783–789. https://doi.org/10.1645/19-198
Bruijns, B., Hoekema, T., Oomens, L., Tiggelaar, R., & Gardeniers, H. (2022). Performance of spectrophotometric and fluorometric DNA quantification methods. Analytica, 3(3), 371–384. https://doi.org/10.3390/analytica3030025
Calegar, D. A., Nunes, B. C., Monteiro, K. J. L., dos Santos, J. P., Toma, H. K., Gomes, T. F., & Carvalho-Costa, F. A. (2016). Frequency and molecular characterisation of Entamoeba histolytica, Entamoeba dispar, Entamoeba moshkovskii, and Entamoeba hartmanni in the context of water scarcity in northeastern Brazil. Memórias do Instituto Oswaldo Cruz, 111(2), 114–119. https://doi.org/10.1590/0074-02760150383
Calle-Pacheco, G. L., Jiménez-Chunga, J. A., & Vivas-Ruiz, D. E. (2022). Molecular diagnosis of amoebiasis. Boletín Médico del Hospital Infantil de México, 79(1), 3–16. https://doi.org/10.24875/bmhim.21000044
Costa, J. D., Resende, J. A., Gil, F. F., Santos, J. F. G., & Gomes, M. A. (2018). Prevalence of Entamoeba histolytica and other enteral parasitic diseases in the metropolitan region of Belo Horizonte, Brazil: A cross-sectional study. Sao Paulo Medical Journal, 136(4), 319–323. https://doi.org/10.1590/1516-3180.2018.0036170418
Cuellar-Guevara, F. L., Barron-Gonzalez, M. P., & Menchaca-Arredondo, J. L. (2019). Effect of Lactobacillus postbiotic on Entamoeba histolytica trophozoites. Revista de Investigación Clínica, 71(6), 402–407. https://doi.org/10.24875/ric.19003134
David, E. B., Coradi, S. T., Oliveira-Sequeira, T. C. G., Ribolla, P. E. M., Katagiri, S., & Guimarães, S. (2011). Diagnosis of Giardia infections by PCR-based methods in children of an endemic area. The Journal of Venomous Animals and Toxins Including Tropical Diseases, 17(2), 209–215. https://doi.org/10.1590/S1678-91992011000200012
Farhan, S. M., Abdulrazzaq, S. A., & Mohammed, A. J. (2024). Differential diagnosis of Entamoeba spp. using the 18SrRNA gene in gastroenteritis patients. Iraqi Journal of Science, 65(1), 119–125. https://doi.org/10.24996/ijs.2024.65.1.11
Flaih, M. H., Khazaal, R. M., Kadhim, M. K., Hussein, K. R., & Alhamadani, F. A. B. (2021). The epidemiology of amoebiasis in Thi-Qar Province, Iraq (2015-2020): Differentiation of Entamoeba histolytica and Entamoeba dispar using nested and real-time polymerase chain reaction. Epidemiology and Health, 43, e2021034. https://doi.org/10.4178/epih.e2021034
Furrows, S. J., Moody, A. H., & Chiodini, P. L. (2004). Comparison of PCR and antigen detection methods for diagnosis of Entamoeba histolytica infection. Journal of Clinical Pathology, 57(12), 1264–1266. https://doi.org/10.1136/jcp.2004.017822
Haghighi, A., Salimi Khorashad, A., Nazemalhosseini Mojarad, E., Kazemi, B., Rostami Nejad, M., & Rasti, S. (2009). Frequency of enteric protozoan parasites among patients with gastrointestinal complaints in medical centers of Zahedan, Iran. Transactions of the Royal Society of Tropical Medicine and Hygiene, 103(5), 452–454. https://doi.org/10.1016/j.trstmh.2008.11.004
Hooshyar, H., Rostamkhani, P., & Rezaian, M. (2012). Molecular epidemiology of human intestinal amoebas in Iran. Iranian Journal of Public Health, 41(9), 10–17.
Iannacone, J., Osorio-Chumpitaz, M., Utia-Yataco, R., Alvariño-Flores, L., Ayala-Sulca, Y., Del Águila-Pérez, C. A., & Wetzel, E. J. (2021). Enteroparasitosis in Peru and its relation to the Human Development Index. Revista Médica del Instituto Mexicano del Seguro Social, 59(5), 368–376.
Kadri, K. (2019). Polymerase Chain Reaction (PCR): Principle and application. In M. L. Nagpal, O. Boldura, C. Balta, & S. Enany (Eds.), Synthetic biology: New interdisciplinary science. IntechOpen. https://doi.org/10.5772/intechopen.86491
Karim, A., Zartashia, B., Khwaja, S., Akhter, A., Raza, A. A., & Parveen, S. (2024). Prevalence and risk factors associated with human intestinal parasitic infections (IPIs) in rural and urban areas of Quetta, Pakistan. Brazilian Journal of Biology, 84, e266898. https://doi.org/10.1590/1519-6984.266898
Kathryn, M. M., & Charles, R. S. (2007). Sensitivity of nested PCR in the detection of low numbers of Giardia lamblia cysts. Applied and Environmental Microbiology, 73(18), 5949–5950. https://doi.org/10.1128/AEM.00668-07
Lalle, M., Bruschi, F., Castagna, B., Campa, M., Pozio, E., & Cacciò, S. M. (2009). High genetic polymorphism among Giardia duodenalis isolates from Sahrawi children. Transactions of the Royal Society of Tropical Medicine and Hygiene, 103(8), 834–838. https://doi.org/10.1016/j.trstmh.2009.04.017
López-López, P., Martínez-López, M. C., Boldo-León, X. M., Hernández-Díaz, Y., González-Castro, T. B., Tovilla-Zárate, C. A., & Luna-Arias, J. P. (2017). Detection and differentiation of Entamoeba histolytica and Entamoeba dispar in clinical samples through PCR-denaturing gradient gel electrophoresis. Brazilian Journal of Medical and Biological Research, 50(4), e5997. https://doi.org/10.1590/1414-431X20175997
Madden, G. R., Shirley, D. A., Townsend, G., & Moonah, S. (2019). Lower gastrointestinal bleeding due to Entamoeba histolytica detected early by multiplex PCR: Case report and review of the laboratory diagnosis of amebiasis. The American Journal of Tropical Medicine and Hygiene, 101(6), 1380–1383. https://doi.org/10.4269/ajtmh.19-0237
Neto, W. S. P., Novais, L. C. A., Siqueira, L. R. L., Machado, M. N., Mousinho, A. A. M., Marques, M. F., & Rocha, T. J. M. (2025). Prevalence of enteroparasitosis in users of the clinical analysis laboratory of a municipality of the agreste region of Alagoas. Brazilian Journal of Biology, 85, e290948. https://doi.org/10.1590/1519-6984.290948
Ngui, R., Angal, L., Fakhrurrazi, S. A., Lian, Y. L. A., Ling, L. Y., Ibrahim, J., & Mahmud, R. (2012). Differentiating Entamoeba histolytica, Entamoeba dispar and Entamoeba moshkovskii using nested polymerase chain reaction (PCR) in rural communities in Malaysia. Parasites & Vectors, 5, 187. https://doi.org/10.1186/1756-3305-5-187
Ögren, J., Dienus, O., Beser, J., Henningsson, A. J., & Matussek, A. (2020). Protozoan infections are under-recognized in Swedish patients with gastrointestinal symptoms. European Journal of Clinical Microbiology & Infectious Diseases, 39, 2153–2160. https://doi.org/10.1007/s10096-020-03974-w
Oliveira, Y. L. C., Oliveira, L. M., Oliveira, Y. L. M., Nascimento, A. M. D., Corte, R. L., Geraldi, R. M., & Dolabella, S. S. (2020). Changes in the epidemiological profile of intestinal parasites after a school-based large-scale treatment for soil-transmitted helminths in a community in northeastern Brazil. Acta Tropica, 202, 105279. https://doi.org/10.1016/j.actatropica.2019.105279
Parija, S. C., & Khairnar, K. (2007). Detection of excretory Entamoeba histolytica DNA in the urine, and detection of E. histolytica DNA and lectin antigen in the liver abscess pus for the diagnosis of amoebic liver abscess. BMC Microbiology, 7, 41. https://doi.org/10.1186/1471-2180-7-41
Rahman, H. U., Khatoon, N., Arshad, S., Masood, Z., Ahmad, B., Khan, W., & Garedaghi, Y. (2022). Prevalence of intestinal nematodes infection in school children of urban areas of district Lower Dir, Pakistan. Brazilian Journal of Biology, 82, e244158. https://doi.org/10.1590/1519-6984.244158
Ramos, A. L. A., Cavalcante, N. T. P., Carvalho Neto, A. P. M., Silva, L. E. O., Silva, A. R., Messias, H. B. G., & Rocha, T. J. M. (2023). Socioeconomic profile and knowledge about helminths among students of a public school. Contribuciones a las Ciencias Sociales, 16(4), 1575–1592. https://doi.org/10.55905/revconv.16n.4-007
Rivero, Z., Villarreal, L., Bracho, Á., Prieto, C., & Villalobos, R. (2021). Identificación molecular de Entamoeba histolytica, Entamoeba dispar y Entamoeba moshkovskii en niños con diarrea en Maracaibo, Venezuela. Biomédica, 41(Supl. 1), 23–34. https://doi.org/10.7705/biomedica.5584
Rosa, W. D., Acuña, A. M., Giachetto, G., Durán, E., Cancel, M. J., Guttérrez, S., & Picheli, E. (2020). Intestinal parasites in schoolchildren, public health problem: Intervention from the National Integrated Health System of Uruguay. Revista de Salud Pública, 22(1), 82–87. https://doi.org/10.15446/rsap.V22n1.77311
Santos, P., Demacedo, B., & Peralto, M. (2007). Comparison of multiplex-PCR and antigen detection for differential diagnosis of E. histolytica. The Brazilian Journal of Infectious Diseases, 11(3), 365–370. https://doi.org/10.1590/S1413-86702007000300013
Sastry, A. S., & Bhat, S. K. (2018). Essentials of medical parasitology (2nd ed.). Jaypee Brothers Medical Publishers.
Servián, A., Zonta, M. L., & Navone, G. T. (2023). Differential diagnosis of human Entamoeba infections: Morphological and molecular characterization of new isolates in Argentina. Revista Argentina de Microbiología, 56(1), 16–24. https://doi.org/10.1016/j.ram.2023.05.003
Sharif, B. O., Ali, Z. R., & Mohammed, H. M. (2022). Impact of Entamoeba histolytica on the human body. International Journal of Medical Sciences and Nursing Research, 2(4), 5–8. https://doi.org/10.55349/ijmsnr.20222458
Silva, B. R., Martins, M. E. L., Silva Dias, T., Costa Martins, J. I., Neves, E. B., Melo, E. S. S., & Monteiro, E. L. (2021). Assistência de enfermagem a crianças ribeirinhas com parasitoses na Amazônia: Revisão integrativa de literatura. Research, Society and Development, 10(5), e34010515010. https://doi.org/10.33448/rsd-v10i5.15010
Silva, S. R. G., Azevedo, P. V. M., Júnior, C. J. D., da Costa, J. G., Pavão, J. M. S. J., dos Santos, A. F., & Rocha, T. J. M. (2021). Environmental characteristics, nutritional and executive functions in children of 6 to 7 years. Brazilian Journal of Biology, 83, e248778. https://doi.org/10.1590/1519-6984.248778
Utaminingsih, S., & Sophian, A. (2022). Analysis of purity and concentration of DNA isolation results on chondroitin samples. BiosciED: Journal of Biological Science and Education, 3(2), 56–61. https://doi.org/10.37304/bed.v3i2.5425
Vasquez-Rios, G., Machicado, J. D., Terashima, A., & Marcos, L. A. (2016). Irritable bowel syndrome and intestinal parasites: A view from South America. Revista de Gastroenterología del Perú, 36(2), 153–158.
Wesel, J., Shuman, J., Bastuzel, I., Dickerson, J., & Ingram-Smith, C. (2021). Encystation of Entamoeba histolytica in axenic culture. Microorganisms, 9(4), 873. https://doi.org/10.3390/microorganisms9040873
Ysea, M. A. V., Umana, M. C., Fuentes, S. P., Campos, I. V., & Carmona, M. C. (2022). Standardization of molecular techniques for the detection and characterization of intestinal protozoa and other pathogens in humans. Journal of Venomous Animals and Toxins Including Tropical Diseases, 28, e20210099. https://doi.org/10.1590/1678-9199-JVATITD-2021-0099
Copyright (c) 2026 Sadiq Omar Khattab Badr, Maroof Sabti Juma Al-Ammash (Autor)
This work is licensed under a Creative Commons Attribution 4.0 International License.
DECLARATION OF ORIGINALITY AND COPYRIGHTS
I Declare that current article is original and has not been submitted for publication, in part or in whole, to any other national or international journal.
The copyrights belong exclusively to the authors. Published content is licensed under Creative Commons Attribution 4.0 (CC BY 4.0) guidelines, which allows sharing (copy and distribution of the material in any medium or format) and adaptation (remix, transform, and build upon the material) for any purpose, even commercially, under the terms of attribution.
Read this link for further information on how to use CC BY 4.0 properly.
