Evaluation of Semi-Nested PCR Compared with Indirect-ELISA to Diagnose Human Fasciolosis
-
Gita Alizadeh
,
Mojgan Aryaeipour
,
Mehdi Mohebali
,
Gholam Reza Mowlavi
,
Vahid Raissi
,
Mohammad Bagher Rokni
Abstract
Background: We aimed to compare semi-nested PCR with indirect ELISA to diagnose human fasciolosis.
Methods: Overall, 70 serum samples were collected from different areas in Iran suspected for fascioliasis. Individuals were classified based on diagnostic of fascioliasis and habitat in endemic areas. Finally, all serum samples were tested by indirect ELISA (using secretory excretory antigen) and semi-nested PCR (using ITS1 gene). The study was conducted in the School of Publish Health, Tehran University of Medical Sciences, Iran in 2021.
Results: Significant differences were found between agreement and similarity of patients' results of indirect ELISA and semi-nested PCR 94.46% and 98.4% respectively (Cohen's kappa ≥0.6; P-value≤0.05). No cross-reactions were observed with other parasitic diseases (toxocariasis, hydatidosis, strongyloidiasis, toxoplasmosis, cutaneous leishmaniasis, taeniasis and trichinosis). 69.84% of samples were positive by both techniques. In addition, the percentage of agreement and similarity between the results of the two techniques based on habitat in endemic areas was 88.9-100% and 97.7-100%, respectively (Cohen's kappa ≥0.6; P-value≤0.05).
Conclusion: Semi-nested PCR could be a suitable method for following up on patients' treatment and a confirmatory method for ELISA as for diagnosis of human fascioliasis.
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2. Siles-Lucas M, Becerro-Recio D, Serrat J, et al (2021). Fascioliasis and fasciolopsiasis: Current knowledge and future trends. Res Vet Sci, 134(1):27-35.
3. Bargues MD, Artigas P, Angles R, et al (2020). Genetic uniformity, geographical spread and anthropogenic habitat modi-fications of lymnaeid vectors found in a one health initiative in the highest human fascioliasis hyperendemic of the Bolivian Altiplano. Parasites Vectors,13(1):1-19.
4. Aguayo V, Valdes B, Espino AM (2018). Assessment of Fasciola hepatica glutathione S-transferase as an antigen for serodiag-nosis of human chronic fascioliasis. Acta Trop, 186(1):41-9.
5. Moghaddam AS, Massoud J, Mahmoodi M, et al (2004). Human and animal fascio-liasis in Mazandaran province, northern Iran. Parasitol Res, 94(1): 61–9.
6. Sarkari B, Khabisi SA (2017). Immunodiag-nosis of human fascioliasis: an update of concepts and performances of the sero-logical assays. J Clin Diagn Res, 11(6): OE05–OE10.
7. Rehman ZU, Martin K, Zahid O, et al (2021). High-throughput sequencing of Fasciola spp. shows co-infection and in-termediate forms in Balochistan, but only F. gigantica in the Punjab province of Pa-kistan. Infect Genet Evol, 94(1):105-12.
8. Mirzadeh A, Jafarihaghighi F, Kazemirad E, et al (2021). Recent Developments in Re-combinant Proteins for Diagnosis of Human Fascioliasis. Acta Parasitol, 66(1):13-25.
9. Carnevale S, Rodrı́guez MI, Santillán G, et al (2001). Immunodiagnosis of human fascioliasis by an enzyme-linked im-munosorbent assay (ELISA) and a mi-cro-ELISA. Clin Diagn Lab Immunol, 8(1):174-77.
10. Alda P, Lounnas M, Vázquez AA, et al (2018). A new multiplex PCR assay to distinguish among three cryptic Galba species, intermediate hosts of Fasciola he-patica. Vete Parasitol, 251(1):101-05.
11. Carnevale S, Malandrini JB, Pantano ML, et al (2021). Use of the PCR in a Combined Methodological Approach for the Study of Human Fascioliasis in an Endemic Ar-ea. Acta Parasit, 66(1):455–60.
12. Arifin MI, Höglund J, Novobilský A (2016). Comparison of molecular and conven-tional methods for the diagnosis of Fasci-ola hepatica infection in the field. Vet Para-sitol, 232(1):8-11.
13. Aryaeipour M, Kazemi B, Bozorgomid A, et al (2020). Detection Cell-Free DNA (cfDNA) Using nested PCR as a Diag-nosis Tool for Human Fascioliasis Infec-tion. Iran J Public Health, 49(6):1148-56.
14. Aryaeipour M, Rouhani S, Bandehpour M, et al (2014). Genotyping and phylogenetic analysis of Fasciola spp. isolated from sheep and cattle using PCR-RFLP in Ar-dabil province, northwestern Iran. Iran J Public Health. 43(10):1364-70.
15. Itagaki T, Kikawa M, Terasaki K, et al (2005). Molecular characterization of partheno-genic Fasciola spp. in Korea on the basis of DNA sequences of ribosomal ITS1 and mitochondrial NDI gene. J Vet Med Sci, 67(11):1115-18.
16. Rokni MB, Bozorgomid A, Heydarian P, et al (2018). Molecular evidence of human Fasciolosis due to Fasciola gigantica in Iran: a case report. Iran J Public Health, 7(5):750-57.
17. Aghayan S, Gevorgian H, Ebi D, et al (2019). Fasciola spp. in Armenia: genetic diversity in a global context. Vet Parasitol, 268(1):21-31.
18. Cabada MM, White Jr AC (2012). New de-velopments in epidemiology, diagnosis, and treatment of fascioliasis. Curr Opin In-fect Dis, 25(5):518-22.
19. Caravedo MA, Cabada MM (2020). Human fascioliasis: Current epidemiological status and strategies for diagnosis, treatment, and control. Res Rep Trop Med, 1(1): 149–58.
20. Sah R, Khadka S, Khadka M, et al (2017). Human fascioliasis by Fasciola hepatica: the first case report in Nepal. BMC Res Notes,10(1):1-4.
21. Sarkari B, Ghobakhloo N, Moshfea A, et al (2012). Seroprevalence of human fasci-olosis in a new-emerging focus of fasci-olosis in Yasuj district, southwest of Iran. Iran J Parasitol, 7(2):15-22.
22. Abdolahi Khabisi S, Sarkari B, Moshfe A, et al (2017). Production of monoclonal an-tibody against excretory-secretory antigen of Fasciola hepatica and evaluation of its ef-ficacy in the diagnosis of fascioliasis. Monoclon Antibodies Immunodiagn Immunother, 36(1):8-14.
23. Mas-Coma S, Bargues MD, Valero MA (2014). Diagnosis of human fascioliasis by stool and blood techniques: update for the present global scenario. Parasitolo-gy, 141(14):1918-46.
24. Mirahmadi H, Rahmati-Balaghaleh M, Afzalaghaee M, et al (2021). Detection of malaria using blood smear by light mi-croscopy, RDT and nested PCR for sus-pected patients in south-eastern Iran. Gene Reports,13(4):1013-19.
25. Raissi V, Mohebali M, Eshrat Bigom KI, et al (2020). Expression of Mir-21 and Mir-103a in toxocara canis: potential for diag-nosis of human toxocariasis. Iran J Parasi-tol, 15(4):559-66.
26. Tang ZL, Huang Y, Yu XB (2016). Current status and perspectives of Clonorchis sinensis and clonorchiasis: epidemiology, patho-genesis, omics, prevention and control. Infect Dis Poverty, 5(1):1-2.
27. Khalifa MM, Abdel-Rahman SM, Bakir HY, et al (2020). Comparison of the diagnos-tic performance of microscopic examina-tion, Copro-ELISA, and Copro-PCR in the diagnosis of Capillaria philippinensis in-fections. Plos one, 15(6): e0234746.
28. Saki J, Mowla K, Arjmand R, et al (2021). Prevalence of Toxoplasma gondii and Toxo-cara canis Among Myositis Patients in the Southwest of Iran. Infect Disord Drug Tar-gets, 21(1):43-8.
29. Wichmann D, Panning M, Quack T, et al (2009). Diagnosing schistosomiasis by detection of cell-free parasite DNA in human plasma. PLOS Negl Trop Dis, 3(4):1-9.
30. Moghaddassani H, Mirhendi H, Hosseini M, et al (2011). Molecular diagnosis of Strongyloides stercoralis infection by PCR de-tection of specific DNA in human stool samples. Iran J Parasitol, 6(2): 23-30.
31. Ramahefarisoa RM, Rakotondrazaka M, Jambou R, et al (2010). Comparison of ELISA and PCR assays for the diagnosis of porcine cysticercosis. Vete Parasitol, 173(3-4):336-39.
32. Garcia JL, Burrells A, Bartley PM, et al (2017). The use of ELISA, nPCR and qPCR for diagnosis of ocular toxoplas-mosis in experimentally infected pigs. Res Vet Sci, 115(1):490-95.
33. Le TH, Nguyen KT, Nguyen NT, et al (2012). Development and evaluation of a single-step duplex PCR for simultaneous detection of Fasciola hepatica and Fasciola gi-gantica (family Fasciolidae, class Trema-toda, phylum Platyhelminthes). J Clin Mi-crobiol, 50(8):2720-26.
34. Martínez-Pérez JM, Robles-Pérez D, Rojo-Vázquez FA, et al (2012). Comparison of three different techniques to diagnose Fasciola hepatica infection in experimentally and naturally infected sheep. Vet Parasitol, 190(1-2):80-6.
35. Valero MA, Periago MV, Pérez-Crespo I, et al (2012). Field evaluation of a coproanti-gen detection test for fascioliasis diagno-sis and surveillance in human hyperen-demic areas of Andean countries. PLOS Negl Trop Dis, 6(9):1-11.
36. Cabada MM, Malaga JL, Castellanos-Gonzalez A, et al (2017). Recombinase polymerase amplification compared to real-time polymerase chain reaction test for the detection of Fasciola hepatica in human stool. Am J Trop Med Hyg, 96(2):341–46.
37. Amiri S, Shemshadi B, Shirali S, et al (2021). Accurate and rapid detection of Fasciola hepatica copro‐DNA in sheep using loop‐mediated isothermal amplification (LAMP) technique. Vet Med Sci, 7(4):1316-24.
38. Hosseini-Safa A, Rokni MB, Mosawi SH, et al (2019). High-resolution melting analysis as an appropriate method to differentiate between F. hepatica and F. gigantica. Iran J Public Health, 48(3):501-07.
2. Siles-Lucas M, Becerro-Recio D, Serrat J, et al (2021). Fascioliasis and fasciolopsiasis: Current knowledge and future trends. Res Vet Sci, 134(1):27-35.
3. Bargues MD, Artigas P, Angles R, et al (2020). Genetic uniformity, geographical spread and anthropogenic habitat modi-fications of lymnaeid vectors found in a one health initiative in the highest human fascioliasis hyperendemic of the Bolivian Altiplano. Parasites Vectors,13(1):1-19.
4. Aguayo V, Valdes B, Espino AM (2018). Assessment of Fasciola hepatica glutathione S-transferase as an antigen for serodiag-nosis of human chronic fascioliasis. Acta Trop, 186(1):41-9.
5. Moghaddam AS, Massoud J, Mahmoodi M, et al (2004). Human and animal fascio-liasis in Mazandaran province, northern Iran. Parasitol Res, 94(1): 61–9.
6. Sarkari B, Khabisi SA (2017). Immunodiag-nosis of human fascioliasis: an update of concepts and performances of the sero-logical assays. J Clin Diagn Res, 11(6): OE05–OE10.
7. Rehman ZU, Martin K, Zahid O, et al (2021). High-throughput sequencing of Fasciola spp. shows co-infection and in-termediate forms in Balochistan, but only F. gigantica in the Punjab province of Pa-kistan. Infect Genet Evol, 94(1):105-12.
8. Mirzadeh A, Jafarihaghighi F, Kazemirad E, et al (2021). Recent Developments in Re-combinant Proteins for Diagnosis of Human Fascioliasis. Acta Parasitol, 66(1):13-25.
9. Carnevale S, Rodrı́guez MI, Santillán G, et al (2001). Immunodiagnosis of human fascioliasis by an enzyme-linked im-munosorbent assay (ELISA) and a mi-cro-ELISA. Clin Diagn Lab Immunol, 8(1):174-77.
10. Alda P, Lounnas M, Vázquez AA, et al (2018). A new multiplex PCR assay to distinguish among three cryptic Galba species, intermediate hosts of Fasciola he-patica. Vete Parasitol, 251(1):101-05.
11. Carnevale S, Malandrini JB, Pantano ML, et al (2021). Use of the PCR in a Combined Methodological Approach for the Study of Human Fascioliasis in an Endemic Ar-ea. Acta Parasit, 66(1):455–60.
12. Arifin MI, Höglund J, Novobilský A (2016). Comparison of molecular and conven-tional methods for the diagnosis of Fasci-ola hepatica infection in the field. Vet Para-sitol, 232(1):8-11.
13. Aryaeipour M, Kazemi B, Bozorgomid A, et al (2020). Detection Cell-Free DNA (cfDNA) Using nested PCR as a Diag-nosis Tool for Human Fascioliasis Infec-tion. Iran J Public Health, 49(6):1148-56.
14. Aryaeipour M, Rouhani S, Bandehpour M, et al (2014). Genotyping and phylogenetic analysis of Fasciola spp. isolated from sheep and cattle using PCR-RFLP in Ar-dabil province, northwestern Iran. Iran J Public Health. 43(10):1364-70.
15. Itagaki T, Kikawa M, Terasaki K, et al (2005). Molecular characterization of partheno-genic Fasciola spp. in Korea on the basis of DNA sequences of ribosomal ITS1 and mitochondrial NDI gene. J Vet Med Sci, 67(11):1115-18.
16. Rokni MB, Bozorgomid A, Heydarian P, et al (2018). Molecular evidence of human Fasciolosis due to Fasciola gigantica in Iran: a case report. Iran J Public Health, 7(5):750-57.
17. Aghayan S, Gevorgian H, Ebi D, et al (2019). Fasciola spp. in Armenia: genetic diversity in a global context. Vet Parasitol, 268(1):21-31.
18. Cabada MM, White Jr AC (2012). New de-velopments in epidemiology, diagnosis, and treatment of fascioliasis. Curr Opin In-fect Dis, 25(5):518-22.
19. Caravedo MA, Cabada MM (2020). Human fascioliasis: Current epidemiological status and strategies for diagnosis, treatment, and control. Res Rep Trop Med, 1(1): 149–58.
20. Sah R, Khadka S, Khadka M, et al (2017). Human fascioliasis by Fasciola hepatica: the first case report in Nepal. BMC Res Notes,10(1):1-4.
21. Sarkari B, Ghobakhloo N, Moshfea A, et al (2012). Seroprevalence of human fasci-olosis in a new-emerging focus of fasci-olosis in Yasuj district, southwest of Iran. Iran J Parasitol, 7(2):15-22.
22. Abdolahi Khabisi S, Sarkari B, Moshfe A, et al (2017). Production of monoclonal an-tibody against excretory-secretory antigen of Fasciola hepatica and evaluation of its ef-ficacy in the diagnosis of fascioliasis. Monoclon Antibodies Immunodiagn Immunother, 36(1):8-14.
23. Mas-Coma S, Bargues MD, Valero MA (2014). Diagnosis of human fascioliasis by stool and blood techniques: update for the present global scenario. Parasitolo-gy, 141(14):1918-46.
24. Mirahmadi H, Rahmati-Balaghaleh M, Afzalaghaee M, et al (2021). Detection of malaria using blood smear by light mi-croscopy, RDT and nested PCR for sus-pected patients in south-eastern Iran. Gene Reports,13(4):1013-19.
25. Raissi V, Mohebali M, Eshrat Bigom KI, et al (2020). Expression of Mir-21 and Mir-103a in toxocara canis: potential for diag-nosis of human toxocariasis. Iran J Parasi-tol, 15(4):559-66.
26. Tang ZL, Huang Y, Yu XB (2016). Current status and perspectives of Clonorchis sinensis and clonorchiasis: epidemiology, patho-genesis, omics, prevention and control. Infect Dis Poverty, 5(1):1-2.
27. Khalifa MM, Abdel-Rahman SM, Bakir HY, et al (2020). Comparison of the diagnos-tic performance of microscopic examina-tion, Copro-ELISA, and Copro-PCR in the diagnosis of Capillaria philippinensis in-fections. Plos one, 15(6): e0234746.
28. Saki J, Mowla K, Arjmand R, et al (2021). Prevalence of Toxoplasma gondii and Toxo-cara canis Among Myositis Patients in the Southwest of Iran. Infect Disord Drug Tar-gets, 21(1):43-8.
29. Wichmann D, Panning M, Quack T, et al (2009). Diagnosing schistosomiasis by detection of cell-free parasite DNA in human plasma. PLOS Negl Trop Dis, 3(4):1-9.
30. Moghaddassani H, Mirhendi H, Hosseini M, et al (2011). Molecular diagnosis of Strongyloides stercoralis infection by PCR de-tection of specific DNA in human stool samples. Iran J Parasitol, 6(2): 23-30.
31. Ramahefarisoa RM, Rakotondrazaka M, Jambou R, et al (2010). Comparison of ELISA and PCR assays for the diagnosis of porcine cysticercosis. Vete Parasitol, 173(3-4):336-39.
32. Garcia JL, Burrells A, Bartley PM, et al (2017). The use of ELISA, nPCR and qPCR for diagnosis of ocular toxoplas-mosis in experimentally infected pigs. Res Vet Sci, 115(1):490-95.
33. Le TH, Nguyen KT, Nguyen NT, et al (2012). Development and evaluation of a single-step duplex PCR for simultaneous detection of Fasciola hepatica and Fasciola gi-gantica (family Fasciolidae, class Trema-toda, phylum Platyhelminthes). J Clin Mi-crobiol, 50(8):2720-26.
34. Martínez-Pérez JM, Robles-Pérez D, Rojo-Vázquez FA, et al (2012). Comparison of three different techniques to diagnose Fasciola hepatica infection in experimentally and naturally infected sheep. Vet Parasitol, 190(1-2):80-6.
35. Valero MA, Periago MV, Pérez-Crespo I, et al (2012). Field evaluation of a coproanti-gen detection test for fascioliasis diagno-sis and surveillance in human hyperen-demic areas of Andean countries. PLOS Negl Trop Dis, 6(9):1-11.
36. Cabada MM, Malaga JL, Castellanos-Gonzalez A, et al (2017). Recombinase polymerase amplification compared to real-time polymerase chain reaction test for the detection of Fasciola hepatica in human stool. Am J Trop Med Hyg, 96(2):341–46.
37. Amiri S, Shemshadi B, Shirali S, et al (2021). Accurate and rapid detection of Fasciola hepatica copro‐DNA in sheep using loop‐mediated isothermal amplification (LAMP) technique. Vet Med Sci, 7(4):1316-24.
38. Hosseini-Safa A, Rokni MB, Mosawi SH, et al (2019). High-resolution melting analysis as an appropriate method to differentiate between F. hepatica and F. gigantica. Iran J Public Health, 48(3):501-07.
| Files | ||
| Issue | Vol 51 No 3 (2022) | |
| Section | Original Article(s) | |
| DOI | https://doi.org/10.18502/ijph.v51i3.8947 | |
| Keywords | ||
| Human fascioliasis Semi-nested Polymerase Chain Reaction Parasitology | ||
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How to Cite
1.
Alizadeh G, Aryaeipour M, Mohebali M, Mowlavi GR, Raissi V, Rokni MB. Evaluation of Semi-Nested PCR Compared with Indirect-ELISA to Diagnose Human Fasciolosis. Iran J Public Health. 2022;51(3):686-694.
