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Applied and Environmental Microbiology, October 2008, p. 6348-6357, Vol. 74, No. 20
0099-2240/08/$08.00+0     doi:10.1128/AEM.00913-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.

Sources of Clostridia in Raw Milk on Farms{triangledown} ,{dagger}

Marie-Claude Julien,1,{ddagger} Patrice Dion,1 Carole Lafrenière,2 Hani Antoun,3 and Pascal Drouin4*

Département de phytologie, Pavillon Charles-Eugène Marchand, 1030 avenue de la médecine, Université Laval, Québec, Québec, Canada G1V 0A6,1 Agriculture et Agroalimentaire Canada, Ferme de recherche sur les bovins de boucherie de Kapuskasing, 445 boulevard de l'Université, Rouyn-Noranda, Québec, Canada J9X 5E4,2 Département des sols et de génie agroalimentaire, Pavillon Paul-Comtois, 2425 rue de l'agriculture, Université Laval, Québec, Québec, Canada G1V 0A6,3 Unité de recherche en agroalimentaire, Université du Québec en Abitibi-Témiscamingue, 445 boulevard de l'Université, Rouyn-Noranda, Québec, Canada J9X 5E44

Received 22 April 2008/ Accepted 19 August 2008

A PCR-denaturing gradient gel electrophoresis (DGGE) method was used to examine on-farm sources of Clostridium cluster I strains in four dairy farms over 2 years. Conventional microbiological analysis was used in parallel to monitor size of clostridial populations present in various components of the milk production chain (soil, forage, grass silage, maize silage, dry hay, and raw milk). PCR amplification with Clostridium cluster I-specific 16S rRNA gene primers followed by DGGE separation yielded a total of 47 operational taxonomic units (OTUs), which varied greatly with respect to frequency of occurrence. Some OTUs were found only in forage, and forage profiles differed according to farm location (southern or northern Québec). More clostridial contamination was found in maize silage than in grass silage. Milk represented a potential environment for certain OTUs. No OTU was milk specific, indicating that OTUs originated from other environments. Most (83%) of the OTUs detected in raw milk were also found in grass or maize silage. Milk DGGE profiles differed according to farm and sampling year and fit into two distinct categories. One milk profile category was characterized by the presence of a few dominant OTUs, the presence of which appeared to be more related to farm management than to feed contamination. OTUs were more varied in the second profile category. The identities of certain OTUs frequently found in milk were resolved by cloning and sequencing. Clostridium disporicum was identified as an important member of clostridial populations transmitted to milk. Clostridium tyrobutyricum was consistently found in milk and was widespread in the other farm environments examined.

* Corresponding author. Mailing address: Unité de recherche et de développement en agroalimentaire, Université du Québec en Abitibi-Témiscamingue (UQAT), 445 Boulevard de l'Université, Rouyn-Noranda, Québec, Canada J9X 5E4. Phone: (819) 762-0971, ext. 2260. Fax: (819) 797-4727. E-mail: pascal.drouin{at}uqat.ca

{triangledown} Published ahead of print on 29 August 2008.

{dagger} Supplemental material for this article may be found at http://aem.asm.org/.

{ddagger} Present address: Département de phytologie, Pavillon Paul-Comtois, 2425 rue de l'agriculture, Université Laval, Québec, Québec, Canada G1V 0A6.

Applied and Environmental Microbiology, October 2008, p. 6348-6357, Vol. 74, No. 20
0099-2240/08/$08.00+0     doi:10.1128/AEM.00913-08
Copyright © 2008, American Society for Microbiology. All Rights Reserved.





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