Isolation of Mycobacterium paratuberculosis from mononuclear cells in tissues, blood, and mammary glands of cows with advanced paratuberculosis

Gerard J. Koenig From the Departments of Veterinary Clinical Sciences (Koenig, Hoffsis, Rings, St-Jean) and Veterinary Preventive Medicine (Shulaw, Bech-Nielsen), College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210.

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Glen F Hoffsis From the Departments of Veterinary Clinical Sciences (Koenig, Hoffsis, Rings, St-Jean) and Veterinary Preventive Medicine (Shulaw, Bech-Nielsen), College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210.

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William P. Shulaw From the Departments of Veterinary Clinical Sciences (Koenig, Hoffsis, Rings, St-Jean) and Veterinary Preventive Medicine (Shulaw, Bech-Nielsen), College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210.

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Steen Bech-Nielsen From the Departments of Veterinary Clinical Sciences (Koenig, Hoffsis, Rings, St-Jean) and Veterinary Preventive Medicine (Shulaw, Bech-Nielsen), College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210.

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D. Michael Rings From the Departments of Veterinary Clinical Sciences (Koenig, Hoffsis, Rings, St-Jean) and Veterinary Preventive Medicine (Shulaw, Bech-Nielsen), College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210.

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Guy St-Jean From the Departments of Veterinary Clinical Sciences (Koenig, Hoffsis, Rings, St-Jean) and Veterinary Preventive Medicine (Shulaw, Bech-Nielsen), College of Veterinary Medicine, The Ohio State University, Columbus, OH 43210.

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Summary

Seven mature dairy cows from 6 herds were obtained with history, clinical signs of disease, and laboratory findings suggestive of advanced paratuberculosis. A surgically implanted collection chamber was used to obtain peripheral tissue fluid. Blood, mammary gland flush fluid, and collection chamber flush fluid (ccff) samples were obtained 6 times over a 2-week period from each cow. Mononuclear cell-rich portions of these fluids obtained by gradient centrifugation were submitted for bacteriologic culture of Mycobacterium paratuberculosis and for total and differential cell counts.

Bacteriologic culture of feces for M paratuberculosis and complete necropsy performed on each cow at the conclusion of the study confirmed the diagnosis of paratuberculosis. Numbers of tissue macrophages obtained from ccff samples were lower than expected. Mean (± SD) differential count of tissue macrophages collected from ccff was 65.57 (± 23.39). Mean calculated tissue macrophages (total cell count × differential count) collected from ccff samples was 623.1 (±784.55) cells/μl. Mycobacterium paratuberculosis was isolated from 1 of 42 (2.4%) collections of mononuclear cell-rich portions of plasma and from 2 of 42 (4.8%) ccff samples. Mycobacterium paratuberculosis was not isolated from any collections of mammary gland flush fluid. The collection and processing techniques used in this study did not enhance detection of M paratuberculosis infection in cows with advanced paratuberculosis, beyond that of ileocecal lymph node biopsy or fecal culture.

Summary

Seven mature dairy cows from 6 herds were obtained with history, clinical signs of disease, and laboratory findings suggestive of advanced paratuberculosis. A surgically implanted collection chamber was used to obtain peripheral tissue fluid. Blood, mammary gland flush fluid, and collection chamber flush fluid (ccff) samples were obtained 6 times over a 2-week period from each cow. Mononuclear cell-rich portions of these fluids obtained by gradient centrifugation were submitted for bacteriologic culture of Mycobacterium paratuberculosis and for total and differential cell counts.

Bacteriologic culture of feces for M paratuberculosis and complete necropsy performed on each cow at the conclusion of the study confirmed the diagnosis of paratuberculosis. Numbers of tissue macrophages obtained from ccff samples were lower than expected. Mean (± SD) differential count of tissue macrophages collected from ccff was 65.57 (± 23.39). Mean calculated tissue macrophages (total cell count × differential count) collected from ccff samples was 623.1 (±784.55) cells/μl. Mycobacterium paratuberculosis was isolated from 1 of 42 (2.4%) collections of mononuclear cell-rich portions of plasma and from 2 of 42 (4.8%) ccff samples. Mycobacterium paratuberculosis was not isolated from any collections of mammary gland flush fluid. The collection and processing techniques used in this study did not enhance detection of M paratuberculosis infection in cows with advanced paratuberculosis, beyond that of ileocecal lymph node biopsy or fecal culture.

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