Original Article

Molecular Characterization of Liver Fluke Isolated from Sheep, Goat and Cattle in Sulaymaniyah, Iraq

Abstract

Background: We aimed to determine species of liver fluke that predominately cause fascioliasis in sheep, goats, and cattle in the Sulaymaniyah Province, Iraq using the molecular technique of DNA sequencing and restriction fragment length polymorphism (RFLP).
Methods: The samples were collected from November 2021 to May 2022. The flukes were collected from infected livers of livestock at the slaughterhouse of Sulaymaniyah Governorate, Iraq. A total of 205 flukes were collected from 56 hosts, cattle (n=22), sheep (n=28), and goats (n=6). The specific primers for FCOX1 and 28S rDNA gene amplification were used. The PCR products were subjected to restriction fragment polymorphism (RFLP) assay using Hpy188III and Dra II restriction enzymes, besides DNA sequencing.
Results: The results showed the genetic polymorphisms among the flukes. Three patterns of RFLP were observed Fasciola hepatica, F. gigantica, and F. intermediate, where 28 of them displayed F. hepatica (sheep, n=14, goat, n=3 and cattle, n= 11), whereas 24 samples displayed the F. gigantica (sheep, n=12, goat, n=3 and cattle, n= 9), and only four samples belonged to F. intermediate (sheep n=3 and cattle, n=1). In addition, the result of the ribosomal DNA (28S rDNA) sequencing confirmed that the isolated flukes belonged to F. hepatica, F. gigantica and F. intermediate.
Conclusion: All three main species are present in the study area and F. hepatica predominated among the animal species in this area also, our results concluded that PCR-RFLP is a rapid and reliable method for liver fluke species identification.

1. Da Costa RA, Corbellini LG, Castro-Janer E, Riet-Correa F. Evaluation of losses in carcasses of cattle naturally infected with Fasciola hepatica: effects on weight by age range and on carcass quality parameters. Int J Parasitol. 2019; 1;49(11):867-72.
2. Çelik Ö Y, CELiK BA. Investigation of the prevalence of Fasciola hepatica in small ruminants in the Siirt region, Turkey. Iran J Parasitol. 2018; 13(4):627-31.
3. Kobra PI, Saidijam M, Maghsood A, Matini M, Fallah M. Prevalence of animal fasciolosis and specification of Fasciola spp. isolated from sheep, goats and cattle by molecular method: Hamadan province, west of Iran. Iran J Parasitol. 2018; 13(4):524-31.
4. Pinilla JC, Muñoz AA, Delgado NU. Prevalence and risk factors associated with liver fluke Fasciola hepatica in cattle and sheep in three municipalities in the Colombian Northeastern Mountains. Vet Parasitol Reg Stud Reports. 2020; 19:100364.
5. Muhammad MJ, Hassan ZI. Molecular Diagnosis of Fasciola hepatica in livestock using cox1 gene in Erbil Pro-vence-Kurdistan Region/Iraq. Zanco J Pure Appl Sci. 2021; 33(4):36-42.
6. Ouchene-Khelifi NA, Ouchene N, Dahmani H, Dahmani A, Sadi M, Douifi M. Fasciolosis due to Fasciola hepatica in ruminants in abattoirs and its economic impact in two regions in Algeria. Trop Biomed. 2018; 35(1):181-7.
7. Farjallah S, Slimane BB, Piras CM, Amor N, Garippa G, Merella P. Molecular characterization of Fasciola hepatica from Sardinia based on sequence analysis of genomic and mitochondrial gene markers. Exp Parasitol. 2013; 135(3):471-8.
8. Mikaeel FB. Prevalence of Fasciola hepatica in goats and sheep by using ELISA in sera and milk in Duhok, Iraq. The Iraqi Journal of Veterinary Medicine. 2020; 44(2):113-9.
9. Muhammad MJ, Hassan ZI. Molecular Diagnosis of Fasciola hepatica in livestock using cox1 gene in Erbil Provence-Kurdistan Region/Iraq. Zanco Journal of Pure and Applied Sciences. 2021;33(4):36-42.
10. Othman VS, Hama AA, Garib DT, Zorab RH. Liver Fluke Species Identification Isolated From Humans and Animals Using PCR-RFLP and DNA Sequencing. Uhd J Sci Tech. 2023, 7(1): 66-70.
11. Siribat P, Dekumyoy P, Komalamisra C, Sumruayphol S, Thaenkham U. Molecular identification of Fasciola spp. representative samples from Thailand based on PCR-RFLP. J Trop Med Parasitol. 2018;41:1-7.
12. Raoof HS, Marif HF, Rahman HS, Omar M, Sheikh B, San Ahmed AM. Molecular characterization and phylogenetic analysis of Fasciola species in sheep and goats in Sulaymaniyah Province, Northern Iraq. J Z S, 2020; 22(1):297-305.
13. Teofanova D, Kantzoura V, Walker S, et al. Genetic diversity of liver flukes (Fasciola hepatica) from Eastern Europe. Infect Genet Evol. 2011; 11(1):109-15.
14. Anh DN, Anh LT, Tuan LQ, et al. Identification of Fasciola species isolates from Nghe An Province, Vietnam, based on ITS1 sequence of ribosomal DNA using a simple PCR-RFLP method. J Parasitol Res. 2018; 2018:2958026.
15. Marcilla A, Bargues MD, Mas-Coma SA. PCR-RFLP assay for the distinction between Fasciola hepatica and Fasciola gigantica. Mol Cell Probes. 2002; 16(5):327-33.
16. Ichikawa-Seki M, Ortiz P, Cabrera M, Hobán C, Itagaki T. Molecular characterization and phylogenetic analysis of Fasciola hepatica from Peru. Parasitol Int. 2016; 65(3):171-4.
17. Kasahara S, Ohari Y, Jin S, Calvopina M, Takagi H, Sugiyama H, Itagaki, T. Molecular characterization revealed Fasciola specimens in Ecuador are all Fasciola hepatica, none at all of Fasciola gigantica or parthenogenic Fasciola species. Parasitol Int. 2021;80:102215.
18. Alsulami MN, Mohamed K, Wakid MH, et al. Molecular Characterization of Fasciola hepatica in Sheep Based on DNA Sequences of Ribosomal ITS-1. Infect Drug Resist. 2023; 16:6661-6671.
19. Ashrafi K, Valero M A, Panova M, Periago M V, Massoud J, Mas-Coma S. Phenotypic analysis of adults of Fasciola hepatica, Fasciola gigantica and intermediate forms from the endemic region of Gilan, Iran. Parasitol Int. 2006; 55(4):249-60.
20. Mas-Coma S, Valero MA, Bargues MD. Human and Animal Fascioliasis: Origins and Worldwide Evolving Scenario. Clin Microbiol Rev. 2022 ;35(4):e0008819.
21. Meerkhan AA, Razak AH, Younis BM. The differences between Direct examination and enzyme linked immunosorbent assay (ELISA) test, during the diagnosis of fasciolosis in Jaundiced slaughtered sheep in Duhok Abattoir, Kurdistan Region of Iraq. Inter J Chem Envir Biol Sci. 2013; 1(5):707-9.
22. Aryaeipour M, Rouhani S, Bandehpour M, Mirahmadi H, Kazemi B, Rokni MB. Genotyping and phylogenetic analysis of Fasciola spp. isolated from sheep and cattle using PCR-RFLP in Ardabil province, northwestern Iran. Iran J Public Health. 2014; 43(10):1364-71.
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IssueVol 18 No 4 (2023) QRcode
SectionOriginal Article(s)
DOI https://doi.org/10.18502/ijpa.v18i4.14264
Keywords
Fasciola hepatica Fasciola gigantica Ruminants Iraq

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How to Cite
1.
Othman VS, Hama AA, Zorab RH, Dalimi A. Molecular Characterization of Liver Fluke Isolated from Sheep, Goat and Cattle in Sulaymaniyah, Iraq. Iran J Parasitol. 2023;18(4):554-562.