Comarison of direct microscopic examination, Enzyme Linked Immunosorbent Assay (ELISA), culture and Nested-PCR for diagnosis of herds bulk tank milk infection with Mycobacterium avium subspecies paratuberculosis

Subject Areas : Veterinary Clinical Pathology

آریا Badiei ¹ ( Department of Clinical Scienece, Faculity of Veterinary Medicine, Karaj Branch, Islamic Azad University, Karaj, Iran )
فرهاد Mousakhani ² ( Department of Pathobiology, Faculity of Veterinary Medicine, Karaj Branch, Islamic Azad University, Karaj, Iran )
عباس Barin ³ ( Department of Basic Science, Faculity of Veterinary Medicine, University of Tehran, Tehran, Iran )
امیر Hamidi ⁴ ( Graduate of Veterinary Medicine, Faculty of Veterinary Medicine, Karaj Branch, Islamic Azad University, Karaj, Iran )
محسن Zafari ⁵ ( Graduate of Veterinary Medicine, Faculty of Veterinary Medicine, Karaj Branch, Islamic Azad University, Karaj, Iran )

Received: 2011-02-10 Accepted : 2011-06-08 Published : 2012-02-20

Keywords: cattle, Elisa, Nested-PCR, Johne’s diseases, Mycobacterium avium, Subspecies paratuberculosis, Milk calture, Direct microscopic examination,

Abstract :

Causative agent of Johne’s diseases is Mycobacterium avium subspecies paratuberculosis (MAP) and each year, it hurts dairy cattle industry worldwide by depression of production and reproductive indexes and culling of affected animals. This bacteria is known as a zoonotic pathogen and recent researches explain its probable role in Crohn’s disease in humans. This study compared four laboratory diagnostic tests; including Enzyme Linked ImmunoSorbent Assay (ELISA), direct microscopic examination, milk culture and Nested-PCR for diagnosis of herds bulk tank milk infection with MAP bacteria. 100 bulk tank milk samples were obtained from 100 industrial dairy herds of Tehran province. Samples were screened by four tests and 82 samples (82%) detected as positive in culture media, 94 milk samples (94%) were positive by Nested-PCR test and 98 samples were positive in ELISA test results (98%). But direct microscopic examination identified just 33 milk samples as positive. Four positive samples of ELISA test were detected as negative samples by Nested-PCR. These 4 samples in addition to 12 other samples which diagnosed as positive by ELISA, had no growth in culture media (16 samples). These results indicated superiority of Nested-PCR among these four tests for diagnosis of bulk tank milk infection with MAP bacteria. Furthermore, the results represent high prevalence of MAP bacteria in bulk tank milk of dairy herds in Tehran province and it needs more serious eradication efforts.  

References:

1. حسنی طباطبایی، ع. و فیروزی، ر. 1380. بیماری­های باکتریایی دام. چاپ اول، انتشارات دانشگاه تهران، صفحات: 425-398.


2. شاه حسینی، م.ح. و سید رضای تهرانی، م . 1384. واکنش زنجیره­ای پلیمراز. چاپ اول، انتشارات  دانشگاه آزاد اسلامی واحد اسلامشهر- شهریار/ شهرقدس،  صفحه: 165.


3. کریمی، م. و زینلی، س. 1383. PCR مبانی و کاربردهای آزمایشگاهی. (ترجمه)، تالیف: مکفرسون، ام. ج. مولر، اس. جی. چاپ اول، انتشارات اندیشه ظهور، صفحات: 118- 1.


4. Abbas, B. and Riemann, H.P. 1983. Diagnosis of Johne’s disease (paratuberculosis) in northern California cattle and a note on its economic significance. Calif. Vet. 37: 16.


5. Abbas, B., Riemann, H.P. and Lonnerdal, B. 1983. Isolation of specific peptides from Mycobacterium paratuberculosis protoplasm and their use in enzyme-linked immunosorbent assay for the detection of paratuberculosis (Johne’s disease) in cattle. Am. J. Vet. Res. 44: 2229-2236.


6. Benedictus, G., Dijkhuizen, A.A. and Stelwagen, J. 1987. Economic losses due to paratuberculosis in dairy cattle. Vet. Rec. 121: 142-146.


7. Buergelt, C.D., Williams, J.E. 2004.  Nested PCR on blood and milk for the detection of Mycobacterium avium subsp paratuberculosis DNA in clinical and subclinical bovine paratuberculosis. Australian Veterinary Journal. 82(8): 497-503.


8. Chiodini, R.J., Van Kruiningen, H.J. and Merkal, R.S. 1984. Ruminant paratuberculosis (Johne’s disease): The current status and future prospects. Cornell Vet. 74: 218-262.


9. Collins, M.T., and Sockett, D.C. 1993. Accuracy and economics of the USDA-licensed enzyme-linked immunosorbent assay for bovine paratuberculosis. JAVMA. 203: 1456-1463.


10. Corti, S. and Stephan, R. 2002. Detection of Mycobacterium avium subspecies paratuberculosis specific IS900 insertion sequences in bulk-tank milk samples obtained from different regions throughout Switzerland. BMC Microbiology. 2: 2-7.


11. Dundee, L., Grant, I.R., Ball, H.J. and Rowe, M.T. 2001. Comparative evaluation of four decontamination protocols for the isolation of Mycobacterium avium subsp. paratuberculosis from milk. Letters in Applied Microbiology. 33:173-177.


12. Eamens, G.J., Whittington, R.J., Marsh, I.B., Turner, M.J., Saunders, V., et al. 2000. Comparative sensitivity of various faecal culture methods and ELISA in dairy cattle herds with endemic Johne's disease. Veterinary Microbiology. 77: 357-367.


13. Haghkhah, M., Ansari-Lari, M., Novin-Baheran, A.M. and Bahramy, A. 2008. Herd-level prevalence of Mycobacterium avium subspecies paratuberculosis by bulk-tank milk PCR in Fars province (southern Iran) dairy herds. Preventive Veterinary Medicine. 86:8-13.


14. Johnson-Ifearulundu, Y.J. and Kaneene, J.B. 1997. Epidemiology and economic impact of subclinical Johne’s disease: A review. Vet. Bull. 67: 437-447.


15. Johnson-Ifearulundu, Kaneene, Y.J., Sprecher, J.B., Gardiner, D.J. and Lloyd, J.W. 2000. The effect of subclinical Mycobacterium paratuberculosis infection on days opens in Michigan, USA, Dairy Cows. Preventive Veterinary Medicine. 46: 171-181.


16. Nielsen, S.S., Thamsborg, S.M., Houe, H. and Bitsch, V. 2000. Bulk-tank milk ELISA antibodies for estimating the prevalence of paratuberculosis in Danish dairy herds. Preventive Veterinary Medicine. 44: 1-7.


17. Ott, S.L., Wells, S.J. and Wagner, B.A. 1999. Herd-level economic losses associated with Johne’s disease on US dairy operations. Prev. Vet. Med. 40:179-192.


18. Pillai, S.R. and Jayarao, B.M. 2002. Application of IS900 PCR for detection of Mycobacterium avium subsp. paratuberculosis directly from raw milk. American Dairy Science Association. 85:1052-1057.


19. Quinn, P.J., Markey, B.K., Carter, M.E. and Donnelly, W.J.C. 2002. Veterinary microbiology and microbial disease. Blackwell Sience Ltd. U.S.A., First Edition. p: 97-105. 


20. Radostits, O.M., Gay, C.C., Hinchcliff, K.W. and Constable, P.D. 2007. Vetrinary medicine, a textbook of the disease of cattle, horses, sheep, pigs and goats. Saunders, Philadelphia, Tenth Edition. p: 1017-1044.


21. Ristow, P., Silva, M.G., Fonseca, L.S. and Lilenbaum, W. 2006. Evaluation of Mycobacterium avium subsp. paratuberculosis faecal culture protocols and media. Pesq. Vet. Bras. 26(1): 1-4.


22. Sharma, G., Singh, S.V., Sevilla, I., Singh, A.V., Whittington, R.J. and Juste, R.A. 2008. Evaluation of indigenous milk ELISA with m-culture and m-PCR for the diagnosis of Bovine Johne’s disease (BJD) in lactating Indian dairy cattle. Research in Veterinary Science. 84: 30-37.


23. Smith, B.P. 2009. Large animal internal medicine. Mosby Elsevier, USA, Forth Edition. p: 881-887.


24. Sockett, D.C., Conrad, T.A., Thomas, C.B. and Collins, M.T. 1992. Evaluation of four serological tests for bovine paratuberculosis. J. Clin. Microbiol. 30: 1134-1139.


25. Stabel, J.R. 1996. An improved method for cultivation of Mycobacterium paratuberculosis from bovine fecal samples and comparison to three other methods. J. Vet. Diagn. Invest. 9: 375-380.


26. Stabel, J.R., Wells, S.J. and Wagner, B.A. 2002. Relationships between fecal culture, ELISA, and bulk tank milk test results for Johne’s disease in US dairy herds. American Dairy Science Association. 85: 525-531.


27. Sweeney, R.W., Whitlock, R.H., Buckley, C.L. and Spencer, P.A. 1995. Evaluation of a commercial enzyme-linked immunosorbent assay for the diagnosis of paratuberculosis in dairy cattle. J. Vet. Diagn. Invest. 7: 488-493.


28. Sweeney, R.W., Whitlock, R.H. and Rosenberger, A.E. 1992. Mycobacterium paratuberculosis cultured from milk and supramammary lymph nodes of infected asymptomatic cows. J. Clin. Microbiol. 30: 166-171.


29. Taylor, J.H. 2000. Mycobacterium avium subspecies paratuberculosis in the causation of Crohn’s disease. World Journal of Gastroenterology. 6(5): 630-632.


30. Taylor, J.H. and Bull, T. 2002. Crohn's disease caused by Mycobacterium avium subspecies paratuberculosis- a public health tragedy whose resoution is long overdue. J. Med. Microbial. 51: 3-6.


31. Taylor, J.H. and El-Zaatari, F.A.K. 2004. The Mycobacterium avium subspecies paratuberculosis problem and its relation to the causation of Crohn disease. IWA Publishing, London, UK. p: 74-95.