Development of an ex vivo model to study adherence of Mannheimia haemolytica serovar 1 to mucosal tissues of the respiratory tract of cattle

Jean M. Clarke Department of Veterinary Pathobiology, College of Veterinary Medicine, Oklahoma State University, Stillwater, OK 74078.

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Rebecca J. Morton Department of Veterinary Pathobiology, College of Veterinary Medicine, Oklahoma State University, Stillwater, OK 74078.

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Cyril R. Clarke Department of Physiological Sciences, College of Veterinary Medicine, Oklahoma State University, Stillwater, OK 74078.
resent address is Nomadics Inc, 1024, S Innovation Way, Stillwater, OK 74074.

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Robert W. Fulton Department of Veterinary Pathobiology, College of Veterinary Medicine, Oklahoma State University, Stillwater, OK 74078.

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Jeremiah T. Saliki Department of Veterinary Pathobiology, College of Veterinary Medicine, Oklahoma State University, Stillwater, OK 74078.

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Abstract

Objective—To develop and validate an ex vivo model for study of adherence of Mannheimia haemolytica (formerly Pasteurella haemolytica) to respiratory tract mucosa of cattle and to use this model to confirm adherence of M haemolytica serovar 1 (Mh1) to several relevant respiratory mucosal surfaces.

Sample Population—Excised nasal, nasopharyngeal, turbinate, and tonsillar mucosal tissue from the bovine upper respiratory tract.

Procedure—Mh1 was radiolabeled by use of tritiated leucine. Various concentrations of labeled bacteria were incubated with bovine upper respiratory tract tissues for various times. Tissue was washed to remove nonadherent bacteria, and percentage of bacteria adhered (percentage of adherence) was estimated using radioactivity. Using an optimal inoculum concentration and incubation time, percentage of Mh1 adherence was compared on nasal, nasopharyngeal, turbinate, and tonsillar mucosal tissue, and adherence to nasopharyngeal tissue was confirmed by scanning and transmission electron microscopy.

Results—The optimal Mh1 inoculum concentration was 1 × 107 colony forming units/ml and incubation time was 3 hours. Percentage of adherence of Mh1 to nasopharyngeal tissue was greater than adherence to other tissue types.

Conclusions and Clinical Relevance—The ex vivo model maintained the functional and structural integrity of bovine upper respiratory tract mucosa, as confirmed by light and electron microscopy. Electron microscopy revealed participation of epithelial cell cilia and surface mucus in adherence of Mh1 to nasopharyngeal tissue. Adherence of Mh1 was confirmed in repeated assays, indicating that this organism adheres to upper respiratory tract mucosa of cattle. (Am J Vet Res 2001;62:805–811)

Abstract

Objective—To develop and validate an ex vivo model for study of adherence of Mannheimia haemolytica (formerly Pasteurella haemolytica) to respiratory tract mucosa of cattle and to use this model to confirm adherence of M haemolytica serovar 1 (Mh1) to several relevant respiratory mucosal surfaces.

Sample Population—Excised nasal, nasopharyngeal, turbinate, and tonsillar mucosal tissue from the bovine upper respiratory tract.

Procedure—Mh1 was radiolabeled by use of tritiated leucine. Various concentrations of labeled bacteria were incubated with bovine upper respiratory tract tissues for various times. Tissue was washed to remove nonadherent bacteria, and percentage of bacteria adhered (percentage of adherence) was estimated using radioactivity. Using an optimal inoculum concentration and incubation time, percentage of Mh1 adherence was compared on nasal, nasopharyngeal, turbinate, and tonsillar mucosal tissue, and adherence to nasopharyngeal tissue was confirmed by scanning and transmission electron microscopy.

Results—The optimal Mh1 inoculum concentration was 1 × 107 colony forming units/ml and incubation time was 3 hours. Percentage of adherence of Mh1 to nasopharyngeal tissue was greater than adherence to other tissue types.

Conclusions and Clinical Relevance—The ex vivo model maintained the functional and structural integrity of bovine upper respiratory tract mucosa, as confirmed by light and electron microscopy. Electron microscopy revealed participation of epithelial cell cilia and surface mucus in adherence of Mh1 to nasopharyngeal tissue. Adherence of Mh1 was confirmed in repeated assays, indicating that this organism adheres to upper respiratory tract mucosa of cattle. (Am J Vet Res 2001;62:805–811)

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