Prevalence and risk factors analysis of fasciolid trematodes with phylogenetic analysis of Fasciola gigantica in small ruminants of Malakand Division, Pakistan

A. U. Khan; W. Khan; M. I. Khan; A. Akbar; A. Ullah; M. Siraj; J. Jamil; A. B. Ismael; A. A. Swelum

doi:10.2478/helm-2026-0012

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Prevalence and risk factors analysis of fasciolid trematodes with phylogenetic analysis of Fasciola gigantica in small ruminants of Malakand Division, Pakistan

Helminthologia

Volume 63 (2026): Issue 2 (June 2026)

By: A. U. Khan, W. Khan , M. I. Khan, A. Akbar, A. Ullah, M. Siraj, J. Jamil, A. B. Ismael and A. A. Swelum

Open Access

|Sep 2026

Abstract

Fasciolosis infects small ruminants, reduces productivity, and causes considerable economic losses to livestock producers. Despite its economic and veterinary importance, there is a lack of comprehensive studies on fasciolosis in the Malakand Division, Pakistan. Therefore, the study aimed to investigate the prevalence and risk factors associated with fasciolosis in small ruminants in Malakand Division. A total of 1307 rectal fecal samples of sheep and 1274 of goats were collected randomly. The collected fecal samples were examined for Fasciolid eggs using the standard sedimentation technique, followed by microscopic identification based on egg morphology and operculum characteristics. The overall prevalence of fasciolosis in sheep and goats was 3.9 % and 2.04 %, respectively. The present study showed significant differences based on the Chi-square test (P < 0.05) in the prevalence of fasciolosis among animals of different ages, health conditions, drinking water sources, treatment status, and across the four seasons of the study year. Fasciolosis was observed to be significantly higher in young sheep and goats (7.08 %, 2.88 %), in untreated (4.45 %, 2.27 %), emaciated (5.88 %, 5.15 %), stream water drinking (5.41 %, 2.51 %), and during the summer season (6.48 %, 2.98 %), respectively. Microscopic examination revealed the presence of two Fasciola species, Fasciola hepatica and Fasciola gigantica, in fecal samples. However, the internal transcribed spacer (ITS) gene amplification and sequencing were performed for Fasciola gigantica to confirm the species identity and explore genetic relationships. Phylogenetic analysis based on the ITS gene, using a neighbor-joining method implemented in MEGA 11 software, revealed a close genetic relationship between isolates from the Malakand Division and those from Egypt, Iran, Nigeria, and Saudi Arabia. The present study highlights the occurrence of Fasciola species in the area, underscoring age, health status, and treatment practices as key risk factors. These findings provide a foundation for the development of effective control strategies against fasciolosis in small ruminants in the Malakand Division.

References

Abdullah, S.H. (2023): Fascioliasis. In: Aguilar-Marcelino, L., Younus, M., Khan, A., Saeed, N.M., Abbas, R.Z. (Eds) One Health Triad. Volume 3. Faisalabad, Pakistan, Unique Scientific Publishers, pp. 78 – 85. DOI: 10.47278/book.oht/2023.80
Open DOI Search in Google Scholar Back to article
Afshan, K., Valero, M.A., Qayyum, M., Peixoto, R.V., Magraner, A., Mas-Coma, S. (2014a): Phenotypes of intermediate forms of Fasciola hepatica and F. gigantica in buffaloes from Central Punjab, Pakistan. J Helminthol, 88: 417. DOI: 10.1017/S0022149X13000369
Open DOI Search in Google Scholar Back to article
Afshan, K., Fortes-Lima, C.A., Artigas, P., Valero, M.A., Qayyum, M., Mas-Coma, S. (2014b): Impact of climate change and man-made irrigation systems on the transmission risk, long-term trend and seasonality of human and animal fascioliasis in Pakistan. Geospat Health, 8: 317 – 334. DOI: 10.4081/gh.2014.22
Open DOI Search in Google Scholar Back to article
Afshan, K., Sajid, M., Komal, M., ul Hassan, H.S.Z., Narjis, G., Firasat, S. (2022): Diagnostic potential of 36–55 kDa somatic antigens of F. gigantica for bovine fasciolosis. Buffalo Bull, 41(1): 49 – 61. DOI: 10.56825/bufbu.2022.4113170
Open DOI Search in Google Scholar Back to article
Aghayan, H., Gevorgian, D., Ebi, H., Atoyan, F., Addy, U., Mackenstedt, U., Rehmann-Sutter, C. (2019): Fasciola spp. in Armenia: genetic diversity in a global context. Vet Parasitol, 268: 21 – 31. DOI: 10.1016/j.vetpar.2019.02.009
Open DOI Search in Google Scholar Back to article
Ahmed, S., Nawaz, M., Gul, R., Zakir, M., Razzaq, A. (2005): Diversity and prevalence of trematodes in livers of sheep and goat in Quetta, Pakistan. Pak J Zool, 37: 205 – 210
Search in Google Scholar Back to article
Akhtar, A., Arshad, M., Shakeebullah, Habibullah, Hidayatullah, Umer, Ameer, M. (2012): Prevalence of Fasciola hepatica in Sheep and Goat in District Dera Ismail Khan. Pakistan J Sci, 64(1): 31 – 34
Search in Google Scholar Back to article
Alkahtani, N.S., Almutairi, M., Aljasham, A., Alouffi, A., Alshammari, M., Al-Rashoodh, S., Al-Olayan, E. (2024): Molecular identification of Fasciola species based on ribosomal and mitochondrial DNA sequences in Riyadh, Saudi Arabia. Arq Bras Med Vet Zootec, 76(1): 55 – 66. DOI: 10.1590/1678-4162-13133
Open DOI Search in Google Scholar Back to article
Amor, N., Farjallah, S., Salem, M., Lamine, D.M., Merella, P., Said, K., Slimane, B.B. (2011): Molecular characterization of F. gigantica from Mauritania based on mitochondrial and nuclear ribosomal DNA sequences. Exp Parasitol, 129(2): 127 – 136. DOI: 10.1016/j.exppara.2011.07.002
Open DOI Search in Google Scholar Back to article
Ashoor, S.J., Wakid, M.H. (2023): Prevalence and hepatic histopathological findings of fascioliasis in sheep slaughtered in Jeddah, Saudi Arabia. Sci Rep, 13(1): 6609. DOI: 10.1038/s41598-023-33927-0
Open DOI Search in Google Scholar Back to article
Ayele, Y., Wondmnew, F., Yeshiwas, T. (2018): The Prevalence of Bovine and Ovine fasciolosis and the Associated Economic Loss Due to Liver Condemnation in and around Debire Birhan, Ethiopia. Symbiosis Online Publ Immunol, 6(3): 1 – 11. DOI: 10.15226/2372-0948/6/3/00177
Open DOI Search in Google Scholar Back to article
Bilal, M.Q., Hameed, A., Ahmad, T. (2009): Prevalence of Gastrointestinal Parasites in buffalo and cow in Rural Areas of Toba Tek Singh, Pakistan. J Anim Plant Sci, 19(2): 67 – 70
Search in Google Scholar Back to article
Bulla-Castaneda, D.M., Lancheros-Buitrago, D.J., García-Corredor, D.J., C-Giraldo-Forero, J., Pulido-Medellin, M.O. (2023): Seroprevalence and risk factors of bovine fasciolosis in the municipalities of Colombia. Vet World, 16(6): 1293 – 1300. DOI: 10.14202/vetworld.2023.1293-1300
Open DOI Search in Google Scholar Back to article
Caravedo, M.A., Cabada, M.M. (2020): Human fascioliasis: current epidemiological status and strategies for diagnosis, treatment, and control. Res Rep Trop Med, 26(11): 149 – 158. DOI: 10.2147/RRTM.S237461
Open DOI Search in Google Scholar Back to article
Dabasa, G., Shanko, T., Zewdei, W., Jilo, K., Gurmesa, G., Abdela, N. (2017): Prevalence of small ruminant gastrointestinal parasites infections and associated risk factors in selected districts of Bale zone, south eastern Ethiopia. J Parasitol Vector Biol, 9(6): 81 – 88. DOI: 10.5897/JPVB2017.0286
Open DOI Search in Google Scholar Back to article
Durrani, A.Z., Khan, M.S., Ashraf, K., Avais, M., Ijaz, M., Saleem, M.H., Hameed, K. (2017): Molecular epidemiology of small ruminant fasciolosis in selected regions of Azad Jammu and Kashmir. J Anim Plant Sci, 27(5): 1637 – 1642
Search in Google Scholar Back to article
Haleem, S., Shadab, F., Niaz, S., Rehman, H.U., Sajad, S., Qureshi, N.A., Kabir, M. (2016): Prevalence of fascioliasis in cows and sheep in district Mardan (KPK), Pakistan. J Entomol Zool Stud, 4(3): 330 – 334
Search in Google Scholar Back to article
Hussain Shah, M.Z., Khan, M.A., Jan, A., Haseeb, A., Ullah, R. (2015): Detection of gastrointestinal helminthiasis through Coprological examination in small ruminants in and around Agriculture University Peshawar, Pakistan. World J Zool, 10(3): 211 – 215. DOI: 10.5829/idosi.wjz.2015.10.3.9549
Open DOI Search in Google Scholar Back to article
Hussein, A.N.A., Hassan, I.M., Khalifa, R.M.A. (2010): Description of eggs and larval stages of Fasciola, light and scanning electron microscopic studies. Res J Parasitol, 5(1): 1 – 12. DOI: 10.3923/jp.2010.1.12
Open DOI Search in Google Scholar Back to article
Ibrahim, N. (2017): Fascioliasis: systematic review. Adv Biol Res, 11(5): 278 – 285. DOI: 10.5829/idosi.abr.2017.278.285
Open DOI Search in Google Scholar Back to article
Irshad, A., Ullah, A., Khan, M.F., Khan, W., Yasmin, S., Aksar, N., Shah, S.S.H. (2023): Identification and Prevalence of Gastrointestinal Parasites in Goats and Sheep of District Lower Dir, Pakistan. J Xi'an Shiyou Univ Nat Sci Ed, 19(8): 251 – 256. Retrieved February 26, 2025, from Journal of Xi'an Shiyou University, Natural Science Edition database http://www.xisdxjxsu.asia/V19I08-21.pdf
Search in Google Scholar Back to article
Khan, M.I., Khan, W., Akbar, A., Ullah, A., Ali, A., Khan, A.U., Ríos-Escalante, P.R.L. (2025a): Epidemiology, morphometry, and phylogenetic analysis of Haemonchus contortus in small ruminants of Malakand division, Pakistan. Helminthologia, 62(3): 197 – 208. DOI: 10.2478/helm-2025-0023
Open DOI Search in Google Scholar Back to article
Khan, N., Khan, W., Shoaib, M., Althubyani, S.A., Yousaf, M. (2025b): Risk factors, pathologies and control of gut-dwelling roundworms in chickens. Helminthologia, 62(4): 331 – 339. DOI: 10.2478/helm-2025-0032
Open DOI Search in Google Scholar Back to article
Kruchynenko, O.V., Mykhailiutenko, S.M., Petrenko, M.O. (2022): Prevalence of Fascioliasis in Ruminants of the World—meta-analysis. Zoodiversity, 56(5): 419 – 428. DOI: 10.15407/zoo2022.05.419
Open DOI Search in Google Scholar Back to article
Kurnianto, H., Ramanoon, S.Z., Aziz, N.A.A., Indarjulianto, S. (2022): Prevalence, risk factors, and infection intensity of fasciolosis in dairy cattle in Boyolali, Indonesia. Vet World, 15(6): 1438 – 1448. DOI: 10.14202/vetworld.2022.1438-1448
Open DOI Search in Google Scholar Back to article
Mas-Coma, S., Valero, M.A., Bargues, M.D. (2005): Fascioliasis and other plant-borne trematode zoonoses. Int J Parasitol, 35(11–12): 1255 – 1278. DOI: 10.1016/j.ijpara.2005.07.010
Open DOI Search in Google Scholar Back to article
Mas-Coma, S., Valero, M.A., Bargues, M.D. (2009): Fasciola, lymnaeids and human fascioliasis, with a global overview on disease transmission, epidemiology, evolutionary genetics, molecular epidemiology and control. Adv Parasitol, 69: 41 – 146. DOI: 10.1016/S0065-308X(09)69002-3
Open DOI Search in Google Scholar Back to article
Mazid, M.A., Bhattacharjee, J., Begum, N., Rahman, M.H. (2006): Helminth parasites of the digestive system of sheep in Mymensingh, Bangladesh. Bangladesh J Vet Med, 4(2): 117 – 122. DOI: 10.3329/bjvm.v4i2.1294
Open DOI Search in Google Scholar Back to article
Mehmood, N., Zulfiqar, S., Muqaddas, H., Hasnain, M., Zheng, Y., Bisetegn, H., Iqbal, F. (2024): Molecular Characterization and Phylogenetic Analysis of F. gigantica Based on ITS-1 Genetic Marker and copro-epidemiological Survey from Ruminants of Pakistan. Acta Parasitol, 69(4): 1866 – 1875. DOI: 10.1007/s11686-024-00911-1
Open DOI Search in Google Scholar Back to article
Mohammed, U.A., Omar, A.A., Malann, Y.D. (2023): Studies on the prevalence of fascioliasis in slaughtered goats in Bauchi North Senatorial Zone abattoirs Bauchi State, Nigeria. FUDMA J Sci, 7(2): 42 – 44. DOI: 10.33003/fjs-2023-0702-1704
Open DOI Search in Google Scholar Back to article
Mumtaz, I., Khalid, B., Mirafzal, A. (2024): Assessing the Prevalence's of Fascioliasis in Goats in the Central Areas of Khost Province Afghanistan. J Res Appl Sci Biotechnol, 3(1): 209 – 213. DOI: 10.55544/jrasb.3.1.32
Open DOI Search in Google Scholar Back to article
Najib, M., Izani, N., Amilah, W., Faez, A., Shafzol, Z. (2020): A scoping review of the prevalence of fascioliasis in Malaysia and risk factors for infection. Malaysian J Med Sci, 27(1): 22 – 36. DOI: 10.21315/mjms2020.27.1.3
Open DOI Search in Google Scholar Back to article
Ndosi, B.A., Lee, D., Bia, M.M., Yang, H., Hong, M.J., Seo, S., Eom, K.S. (2023): Morphometry and molecular identification of Haemonchus Cobb, 1898 (Trichostrongylidae: Nematoda) isolates from small ruminants in Tanzania based on mitochondrial cox1 and rRNA-ITS genes. J Parasitol Res, 2023: 1923804, 10 pp. DOI: 10.1155/2023/1923804
Open DOI Search in Google Scholar Back to article
Nkurunziza, S., Nishemezwe, G., Ntirandekura, J.B., Niyokwizera, P., Nyabongo, L., Omore, A., Odhiambo, R. (2024): Prevalence and associated risk factors assessment of bovine fasciolosis in the Imbo Region, Burundi. Parasitol Res, 123(1): 33. DOI: 10.1007/s00436-023-08040-w
Open DOI Search in Google Scholar Back to article
Phalee, A., Wongsawad, C., Rojanapaibul, A., Chai, J.Y. (2015): Experimental life history and biological characteristics of F. gigantica (Digenea: Fasciolidae). Korean J Parasitol, 53(1): 59 – 64. DOI: 10.3347/kjp.2015.53.1.59
Open DOI Search in Google Scholar Back to article
Radostits, O.M., Gay, C.C., Hinchcliff, K.W., Constable, P.D. (2007): Veterinary Medicine. A textbook of the diseases of cattle, horses, sheep, pigs and goats. 10th Edition, London, UK, Saunders Elsevier, pp. 2045 – 2050
Search in Google Scholar Back to article
Raza, M.A., Younas, M., Schlecht, E. (2014): Prevalence of gastrointestinal helminths in pastoral sheep and goat flocks in the Cholistan desert of Pakistan. J Anim Plant Sci, 24(1): 127 – 134.
Search in Google Scholar Back to article
Rashid, A., Khattak, M.N.K., Khan, M.F., Ayaz, S., Rehman, A.U. (2016): Gastrointestinal helminthoses: prevalence and associated risk factors in small ruminants of district Kohat, Pakistan. J Anim Plant Sci, 26(4): 956 – 962
Search in Google Scholar Back to article
Rizwan, M., Khan, M.R., Afzal, M.S., Manahil, H., Yasmeen, S., Jabbar, M., Cao, J. (2022): Prevalence of fascioliasis in livestock and humans in Pakistan: a systematic review and meta-analysis. Trop Med Infect Dis, 7(7): 126. DOI: 10.3390/tropicalmed7070126
Open DOI Search in Google Scholar Back to article
Ruhoollah, Khan, W., Al-Jabr, O.A., Khan, T., Khan, A., El-Ghareeb, W.R., Swelum, A.A. (2021): Prevalence of gastrointestinal parasite in small ruminants of District Dir Upper Khyber Pakhtunkhwa Province of Pakistan. Braz J Biol, 83: e248978. DOI: 10.1590/1519-6984.248978
Open DOI Search in Google Scholar Back to article
Ruhoollah, Rafiq, N., Kamal, M., Khan, W., Khan, T., Niaz, S., De Los Ríos-Escalante, P.R., Ben Said, M. (2025): Prevalence and molecular characterization of ascarid nematodes in cattle population from Malakand Division, Pakistan. Helminthologia, 62(4): 321. DOI: 10.2478/helm-2025-0033
Open DOI Search in Google Scholar Back to article
Sarkari, B., Mansouri, M., Noorpisheh Ghadimi, S., Abdolahi Khabisi, S., Doshmanziari, A. (2018): Molecular Evaluation of a Case of Fasciola hepatica in Wild Boar in Southwestern Iran: A Case Report. Iran J Parasitol, 13(1): 149 – 155
Search in Google Scholar Back to article
Sarwar, M.M., Butt, A.A., Ashfaq, K., Chughtai, S.A., Javed, S., Zulfiqar, S., Alvi, M.A. (2025): Survey on Prevalence, Risk Factors and Molecular Detection of Fasciola in Pakistan. Indus J Biosci Res, 3(4): 60 – 72. DOI: 10.70749/ijbr.v3i4.956
Open DOI Search in Google Scholar Back to article
Seldemir, O.S. (2000): Differentiation of cattle and sheep originated Fasciola hepatica by RAPD-PCR. Méd Vét, 157(6): 65 – 67.
Search in Google Scholar Back to article
Soulsby, E.J.L. (1982): Helminths, Arthropods and Protozoa of Domestic Animals. 7th Edition, Philadelphia, USA, Lea and Febiger, pp. 212 – 502
Search in Google Scholar Back to article
Tang, M., Zhou, Y., Liu, Y. (2021): Molecular identification and genetic-polymorphism analysis of Fasciola flukes in Dali Prefecture, Yunnan Province, China. Parasitol Int, 85: 102416. DOI: 10.1016/j.parint.2021.102416
Open DOI Search in Google Scholar Back to article
Tasleem, S., Saddozai, S., Ullah, A., Rahat, A., Aziz, T., Azeem, T., Saeed, K. (2023): Prevalence of Gastrointestinal Tract Parasites in Dairy Cattle in Tehsil Gagra District Buner Khyber Pakhtunkhwa, Pakistan. J Health Rehabil Res, 3(2): 982 – 987. DOI: 10.61919/jhrr.v3i2.265
Open DOI Search in Google Scholar Back to article
Ulhaq, Z., Khan, W., Khan, M.F., Kabir, M., Ujjan, A.A., Ullah, W. (2022): Prevalence of intestinal parasitic diseases in school children of rural areas of district Lower Dir, Pakistan. Braz J Biol, 82: e243150. DOI: 10.1590/1519-6984.243150
Open DOI Search in Google Scholar Back to article
Valino, L.S.I., Venturina, V.M., Mingala, C.N. (2017): Gross and molecular comparison of Fasciola hepatica and F. gigantica from the field in the Philippines. Int J Vet Sci, 6(4): 216 – 222
Search in Google Scholar Back to article
World Health Organization (2021): Foodborne trematode infections. Retrieved February 26, 2025, from World Health Organization https://www.who.int/news-room/fact-sheets/detail/food-borne-trematode-infections
Search in Google Scholar Back to article
Zajac, A., Conboy, G.A. (2012): Veterinary clinical parasitology. 8th Edition, Oxford, UK, Wiley-Blackwell, 354 pp.
Search in Google Scholar Back to article

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DOI: https://doi.org/10.2478/helm-2026-0012 | Journal eISSN: 1336-9083 | Journal ISSN: 0440-6605

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Language: English

Page range: 109 - 117

Submitted on: Jul 5, 2025

Accepted on: Mar 19, 2026

Published on: Sep 7, 2026

Published by: Slovak Academy of Sciences, Institute of Parasitology

In partnership with: Paradigm Publishing Services

Publication frequency: Volume open

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Related subjects:

Zoology,

Ecology,

Life sciences, other,

Medicine,

Clinical medicine,

Microbiology, virology and infection epidemiology

© 2026 A. U. Khan, W. Khan, M. I. Khan, A. Akbar, A. Ullah, M. Siraj, J. Jamil, A. B. Ismael, A. A. Swelum, published by Slovak Academy of Sciences, Institute of Parasitology
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Volume 63 (2026): Issue 2 (June 2026)