Increased in vitro adherence and on-farm persistence of predominant and persistent Listeria monocytogenes strains in the milking system

Authors: LATORRE, ALEJANDRA - Cornell University; Van Kessel, Jo Ann; Karns, Jeffrey; ZURAKOWSKI, MICHAEL - Cornell University; PRADHAN, ABANI - Cornell University; SUKHNANAND, SHARINNE - Cornell University; SCHUKKEN, YNTE - Cornell University

Submitted to: Applied and Environmental Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 3/21/2011
Publication Date: 5/23/2011
Citation: Latorre, A., Van Kessel, J.S., Karns, J.S., Zurakowski, M., Pradhan, A., Sukhnanand, S., Schukken, Y. 2011. Increased in vitro adherence and on-farm persistence of predominant and persistent Listeria monocytogenes strains in the milking system. Applied and Environmental Microbiology. 77(11):3676-3684.

Interpretive Summary: In the US approximately 2500 people are diagnosed annually with Listeriosis, a foodborne disease caused by Listeria monocytogenes. Immunocompromised individuals are at an elevated risk for infection and pregnant women who become infected are at high risk for fetal loss. Dairy farms have been implicated as major reservoirs for human pathogenic L. monocytogenes strains and human outbreaks have been caused by ingestion of unpasteurized, raw milk, raw milk products, or products that were contaminated after pasteurization. Controlling the presence of L. monocytogenes at the farm level is critical for minimizing human exposure. There are considerable differences between strains of L. monocytogenes and, among other things, these differences can determine the persistence of a strain at the farm level. The objectives of this research were to study the diversity of L. monocytogenes strains on a single dairy farm, assess strain dynamics (movement) within the farm, identify potential sources of L. monocytogenes in bulk tank milk and in-line milk filters, and assess the biofilm-forming ability of representative strain types. We think that strains that are capable of forming biofilms are more likely to contaminate the milk handling equipment. A total of 248 L. monocytogenes isolates from many farm samples including raw milk, milk filters, feces, and water were studied. Using Pulsed Field Gel Electrophoresis, a method to determine the relatedness between strains, predominant, persistent, and transient strains were identified. Strains from feces and environmental samples were much more diverse than strains from the in-line milk filters or the raw, bulk tank milk. The strains were also tested for their ability to form biofilms. The L. monocytogenes strains that were predominant and persistent showed better biofilm-forming ability than strains that were either sporadic, predominant, or persistent. Our results suggest that the milking system was exposed to several L. monocytogenes types from different sources. Only three strain types, however, were successful in persisting within the milk system, suggesting the presence of strains that are more suitable to a particular ecological environment. These results will help the dairy industry develop management practices that are effective in controlling L. monocytogenes in milk.

Technical Abstract: Dairy farms are a reservoir for Listeria monocytogenes and control of this pathogen at farm level is critical for reducing human exposure. The objectives of this research were to study the diversity of L. monocytogenes strains on a single dairy farm, assess strain dynamics within the farm, identify potential sources of L. monocytogenes in bulk tank milk and in-line milk filters, and assess the biofilm forming ability of representative strain types. A total of 248 L. monocytogenes isolates were analyzed by pulsed-field gel electrophoresis (PFGE). Combined AscI and ApaI restriction analysis yielded 40 PFGE types. The most predominant PFGE types were F (14.9 %), T (28.6 %), and D (22.6 %). A large heterogeneity among PFGE types in isolates from fecal (Simpson’s Index of Diversity SID=0.96) and environmental samples (SID=0.96), and more homogeneity of PFGE types in isolates from milk filters (SID=0.71) and bulk tank milk (SID=0.65) was observed. Six of 17 L. monocytogenes strains (35.3%) were classified in an in-vitro assay as “low biofilm formers”, nine (52.9%) as “medium biofilm formers”, and two (11.8%) as “high biofilm formers”. The L. monocytogenes strains that were predominant and persistent showed better biofilm forming ability than strains that were either sporadic, predominant, or persistent ( p=0.0006). Our results suggest that the milking system was exposed to several L. monocytogenes types from different sources. Only 3 PFGE types, however, were successful in persisting within the milk system, suggesting the presence of strains that are more suitable to a particular ecological environment.