|Klima, Cassidy - Agriculture And Agri-Food Canada|
|Zaheer, Rahat - Agriculture And Agri-Food Canada|
|Briggs, Robert - Bob|
|Mcallister, Tim - Agriculture And Agri-Food Canada|
Submitted to: Journal of Microbiological Methods
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/17/2017
Publication Date: 5/24/2017
Publication URL: http://handle.nal.usda.gov/10113/5706134
Citation: Klima, C.L., Zaheer, R., Briggs, R.E., McAllister, T.A. 2017. A multiplex PCR assay for molecular capsular serotyping of Mannheimia haemolytica serotypes 1, 2, and 6. Journal of Microbiological Methods. 139(2017):155-160. doi. 10.1016/j.mimet.2017.05.010.
Interpretive Summary: The bacterium Mannheimia haemolytica is a major cause of respiratory disease losses in beef cattle production. Serotypes 1, 2, and 6 are commonly carried by cattle in health or disease but each differ in their ability to elicit disease or response to immunity. Identification of serotype has previously been dependent upon cumbersome serologic methods which require specific reagents not commercially available. In this experiment a simple method was developed to identify serotype of isolated Mannheimia which utilizes unique DNA sequences of serotype-specific biosynthetic genes. Utilizing equipment and reagents commonly available in diagnostic and research laboratories, the new assay will enable routine diagnostic testing and thereby improve application of disease control measures.
Technical Abstract: Mannheimia haemolytica is an important respiratory pathogen of ruminants. Of the 12 capsular serovars identified, 1 and 6 are most frequently associated with disease in cattle, while 2 is largely a commensal. Comparative analysis of 24 M. haemolytica genomes was used to identify unique genes associated with capsular polysaccharide synthesis as amplification targets in a multiplex PCR assay to discriminate between serotype 1, 2, and 6 strains. The specificity of serotype specific gene targets was evaluated against 47 reference strains representing 12 known serovars of M. haemolytica and 101 field isolates identified through antisera agglutination as serotypes 1, 2, or 6. The results suggest this simple and cost-effective serotype specific PCR assay can be used as an alternative to agglutination based techniques to serotype the majority of M. haemolytica collected from bovines, thus averting the need to use animals and invest in expensive sera development for agglutination assays. In addition, the gene targets identified in this study can be used in silico to identify serotype 1, 2, and 6 strains in sequenced M. haemolytica isolates without the need for culture based analysis.