Diagnosis of bovine group A rotavirus, bovine coronavirus, Escherichia coli K99, Cryptosporidium spp., Giardia spp. in calf diarrhea with one-step duplex RT-PCR: histopathological and immunochromotographic methods

Document Type : Original Article

Authors

1 Department of Parasitology, Faculty of Veterinary Medicine, University of Harran, Şanliurfa, Türkiye

2 Department of Pathology, Faculty of Veterinary Medicine, University of Sivas Cumhuriyet, Sivas, Türkiye

3 Pendik Veterinary Control Institute, Virology Diagnostic Laboratory, Istanbul, Türkiye

4 Elaziğ Veterinary Control Institute, Elaziğ, Türkiye

Abstract
Infections causing diarrhea in cattle breeding pose a substantial threat to calf breeding and health, causing considerable economic losses worldwide, including in Türkiye. The primary causative agents of calf diarrhea include bovine group A rotavirus, bovine coronavirus (BCoV), Escherichia coli K99, Cryptosporidium spp., and Giardia spp. The objective of this study was to diagnose the pathogens present in samples taken from calves exhibiting diarrhea symptoms in the Elaziğ province of Türkiye. The study material consisted of 85 fecal samples from calves up to 30 days of age with diarrhea symptoms and 42 small intestine samples from calves died as a result of diarrhea symptoms after necropsy. Following total RNA isolation from the samples, one-step duplex reverse transcription polymerase chain reaction (RT-PCR) analysis was conducted to identify the bovine rotavirus (BRV) and BCoV. Our results indicated that 24 of the samples were positive for BRV, 13 were positive for BCoV, and 25 were positive for BRV and BCoV in the form of mixed infection. The immunochromatographic analysis revealed that three samples were positive for E. coli K99 antigens, 12 for Cryptosporidium spp. antigens, and 28 for Giardia spp. antigens. Our results indicate that one-step duplex RT-PCR, immunohistochemistry, and immunochromatography methods can be valuable tests to be used in routine diagnostic laboratories for the detection of pathogens associated with calf diarrhea.

Keywords

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  1. Cho YI, Yoon KJ. An overview of calf diarrhea - infectious etiology, diagnosis, and intervention. J Vet Sci 2014; 15(1): 1-17.
  2. Windeyer MC, Leslie KE, Godden SM, et al. Factors associated with morbidity, mortality, and growth of dairy heifer calves up to 3 months of age. Prev Vet Med 2014; 113(2): 231-240.
  3. Svensson C, Linder A, Olsson SO. Mortality in Swedish dairy calves and replacement heifers. J Dairy Sci 2006; 89(12): 4769-4777.
  4. Foster DM, Smith GW. Pathophysiology of diarrhea in calves. Vet Clin North Am Food Anim Pract 2009; 25(1): 13-36, xi.
  5. Gillhuber J, Pallant L, Ash A, et al. Molecular identification of zoonotic and livestock-specific Giardia-species in faecal samples of calves in Southern Germany. Parasit Vectors 2013; 6(1): 346. doi: 10.1186/1756-3305-6-346.
  6. Gillhuber J, Rügamer D, Pfister K, et al. Giardiosis and other enteropathogenic infections: a study on diarrhoeic calves in Southern Germany. BMC Res Notes 2014; 7: 112. doi: 10.1186/1756-0500-7-112.
  7. Acres SD. Enterotoxigenic Escherichia coli infections in newborn calves: a review. J Dairy Sci 1985; 68(1): 229-256.
  8. Tzipori S. The relative importance of enteric pathogens affecting neonates of domestic animals. Adv Vet Sci Comp Med 1985; 29: 103-206.
  9. Snodgrass DR, Terzolo HR, Sherwood D, et al. Aetiology of diarrhoea in young calves. Vet Rec 1986; 119(2): 31-34.
  10. Krogh HV, Henriksen SA. Bovine cryptosporidiosis in Denmark. 2. Cryptosporidia associated with neonatal calf diarrhea. Nord Vet Med 1985; 37(1): 42-47.
  11. Torres-Medina A, Schlafer DH, Mebus CA. Rotaviral and coronaviral diarrhea. Vet Clin North Am Food Anim Pract 1985; 1(3): 471-493.
  12. Santín M, Trout JM, Fayer R. A longitudinal study of cryptosporidiosis in dairy cattle from birth to 2 years of age. Vet Parasitol 2008; 155(1-2): 15-23.
  13. Sherwood D, Snodgrass DR, Lawson GH. Prevalence of enterotoxigenic Escherichia coli in calves in Scotland and northern England. Vet Rec 1983; 113(10): 208-212.
  14. Reynolds DJ, Morgan JH, Chanter N, et al. Microbiology of calf diarrhoea in southern Britain. Vet Rec 1986; 119(2): 34-39.
  15. Fayer Cryptosporidium: a water-borne zoonotic parasite. Vet Parasitol 2004; 126(1-2): 37-56.
  16. Thompson RC, Palmer CS, O'Handley R. The public health and clinical significance of Giardia and Cryptosporidium in domestic animals. Vet J 2008; 177(1): 18-25.
  17. Xiao L, Ryan UM. Cryptosporidiosis: an update in molecular epidemiology. Curr Opin Infect Dis 2004; 17(5): 483-490.
  18. Feng Y, Xiao L. Zoonotic potential and molecular epidemiology of Giardia species and giardiasis. Clin Microbiol Rev 2011; 24(1): 110-140.
  19. Cacciò SM, Thompson RC, McLauchlin J, et al. Unravelling Cryptosporidium and Giardia epidemiology. Trends Parasitol 2005; 21(9): 430-437.
  20. Smith HV, Cacciò SM, Tait A, et al. Tools for investigating the environmental transmission of Cryptosporidium and Giardia infections in humans. Trends Parasitol 2006; 22(4): 160-167.
  21. Díaz P, Quílez J, Chalmers RM, et al. Genotype and subtype analysis of Cryptosporidium isolates from calves and lambs in Galicia (NW Spain). Parasitology 2010; 137(8): 1187-1193.
  22. Imre K, Lobo LM, Matos O, et al. Molecular characterisation of Cryptosporidium isolates from pre-weaned calves in Romania: is there an actual risk of zoonotic infections? Vet Parasitol 2011; 181(2-4): 321-324.
  23. Zhu W, Dong J, Haga T, et al. Rapid and sensitive detection of bovine coronavirus and group a bovine rotavirus from fecal samples by using one-step duplex RT-PCR assay. J Vet Med Sci 2011; 73(4): 531-534.
  24. Min HG, Lim YH, Kang SY. Determination of porcine rotavirus serotypes by RT-PCR and RFLP analysis. J Bacteriol Virol 2006; 36(4): 255-261.
  25. Cho KO, Hasoksuz M, Nielsen PR, et al. Cross-protection studies between respiratory and calf diarrhea and winter dysentery coronavirus strains in calves and RT-PCR and nested PCR for their detection. Arch Virol 2001; 146(12): 2401-2419.
  26. Hasoksuz M, Hoet AE, Loerch SC, et al. Detection of respiratory and enteric shedding of bovine coronaviruses in cattle in an Ohio feedlot. J Vet Diagn Invest 2002; 14(4): 308-313.
  27. Izzo MM, Kirkland PD, Mohler VL, et al. Prevalence of major enteric pathogens in Australian dairy calves with diarrhoea. Aust Vet J 2011; 89(5): 167-173.
  28. Bartels CJ, Holzhauer M, Jorritsma R, et al. Prevalence, prediction and risk factors of enteropathogens in normal and non-normal faeces of young Dutch dairy calves. Prev Vet Med 2010; 93(2-3): 162-169.
  29. De Verdier K. Infection panorama in diarrhea in Swedish calves [Swedish]. Svensk Veterinärtidning 2006; 8(9): 29-32.
  30. Al M, Balikci E. Detection of Rotavirus, Coronavirus, E. coli K99, Cryptosporidium parvumin neonatal calves with diarrhoea by rapid diagnostic test kits and relationship between these enteropathogens and maternal immunity. [Turkish]. Fırat Univ Sağ Bil Vet Derg 2012; 26(2): 73-78.
  31. Erdoğan HM, Ünver A, Güneş V, et al. Frequency of rotavirus and coronavirus in neonatal calves in Kars region [Turkish]. Kafkas Univ Vet Fak Derg 2003; 9(1): 65-68.
  32. Scott PR, Hall GA, Jones PW, et al. Calf diarrhoea. In: Andrews AH, Blowey RW, Boyd H, et al. (Eds). Bovine medicine diseases and husbuandry of cattle. 2 Oxford, UK: Blackwell Science 2004; 185-214.
  33. Blowey RW. A veterinary book for dairy farmers. 2nd London, UK: Farming Press Ltd. 1993; 15-77.
  34. Bachmann P. Pathogenetic and immunologic features of viral gastroenteritis in calves. Rev Sci Tech 1984; 3(4): 819-842.
  35. Markowska-Daniel I, Winiarczyk S, Gradzki Z, et al. Evaluation of different methods (ELISA, IF, EM, PAGE) for the diagnosis of rotavirus infection in piglets. Comp Immunol Microbiol Infect Dis 1996; 19(3): 219-232.
Volume 16, Issue 4
April 2025
Pages 221-227

  • Receive Date 21 March 2024
  • Revise Date 04 May 2024
  • Accept Date 05 June 2024