Molecular ecology of Listeria monocytogenes: Evidence for a reservoir in milking equipment on a dairy farm

Authors: LATORRE, ALEJANDRA - Cornell University; Van Kessel, Jo Ann; Karns, Jeffrey; ZURAKOWSKI, MIKE - Cornell University; PRADHAN, ABANI - Cornell University; ZADOKS, RUTH - Cornell University; BOOR, KATHRYN - Cornell University; SCHUKKEN, YNTE - Cornell University

Submitted to: Applied and Environmental Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 12/18/2008
Publication Date: 3/1/2009
Citation: Latorre, A.A., Van Kessel, J.S., Karns, J.S., Zurakowski, M.J., Pradhan, A.K., Zadoks, R.N., Boor, K.J., Schukken, Y.H. 2009. Molecular ecology of Listeria monocytogenes: Evidence for a reservoir in milking equipment on a dairy farm. Applied and Environmental Microbiology. 75(5):1315-1323.

Interpretive Summary: Listeria monocytogenes can cause listeriosis in humans. This illness, despite being underreported, is an important public health concern in the Unites States and worldwide. Listeria are ubiquitious in the environment and are frequently isolated from various locations in and around dairy farms. As a result, raw milk is occasionally contaminated with Listeria monocytogenes. This objective of this study was to determine the presence and distribution of Listeria monocytogenes on a dairy farm and to identify potential sources for bulk milk contamination. Listeria monocytogenes was isolated from many farm samples including raw milk, milk filters, feces, and water. Using Pulsed Field Gel Electrophoresis, a method to determine the relatedness between strains, it was shown that the same strains persisted in the milk and milk filters despite regular, rigorous cleaning of the milking equipment. These results suggest that a biofilm of Listeria monocytogenes was persisting in the milking equipment on this farm. This information will help the dairy industry understand potential weak points in milking equipment hygiene and to target these areas with new approaches to decrease pathogen contamination of raw milk.

Technical Abstract: A longitudinal study aimed to detect Listeria monocytogenes on a New York State dairy farm, was conducted between February 2004 and July 2007. Fecal samples were collected every six months from all lactating cows. Approximately 20 environmental samples were obtained every three months. Bulk milk samples and in-line milk filters were collected weekly and swabs from milking equipment and the milking parlor environment were collected in May 2007. Fifty of 716 fecal samples (7%) and 24 of 304 environmental samples (7.9%) were positive for L. monocytogenes. A total of 71 of 115 in-line milk filter samples (61.7%) and 34 of 172 bulk milk samples (19.7%) were positive for L. monocytogenes. In-line milk filters had a greater proportion of L. monocytogenes compared to bulk milk samples (P <0.05). L. monocytogenes was isolated from 6 of 40 (15%) sampling-sites in the milking parlor and milking equipment. In-line milk filters showed a significantly greater proportion of L. monocytogenes positive samples compared to the other sources (P <0.05). Bulk milk also had greater proportion of L. monocytogenes positive samples than the other sources (P <0.05), except for samples from milking parlor and milking equipment. A total of 36 isolates were selected for Pulsed-Field Gel Electrophoresis (PFGE) typing. Seven PFGE types and two subtypes were distinguished with the restriction endonuclease AscI, and seven PFGE types and one subtype were found when using ApaI. The same L. monocytogenes PFGE types were repeatedly found in samples collected from the bulk milk and milk filters over the course of the study. This persistence of strains despite regular cleaning of the milking equipment is highly suggestive of the presence of L. monocytogenes biofilms in the milking equipment.