Application of PCR typing for investigating Listeria monocytogenes transmission along the Gorgonzola PDO production chain

Introduction: Literature shows silage as a likely source of Listeria monocytogenes for milking cows, which shed the pathogen in farm environments (Nightingale, 2004). Presence of L. monocytogenes in farms could lead to its introduction and persistence in processing plants (Lomonaco et al. 2008), as happens in the production chain of Gorgonzola Protected Designation of Origin (PDO). In fact, its peculiar production processes allow survival and growth of Listeria monocytogenes, which may contaminate production environments as well as cheese rinds. Purpose: This study was aimed at evaluating the presence of the pathogen in dairy farms (N=20), and in the plant of Gorgonzola PDO cheese they give milk to. In addition, genetic typing was performed in order to detect possible transmission links. Methods: A survey was carried out over 2 years in Piedmont (Italy); samples were collected from primary production (feed, N=240; feces, N=40; milk filters, N=40, and milk, N=80) and from manufacturer processing environments/equipments (N=108). L. monocytogenes detection was performed using ISO 11290-1:1996/Amd 1:2004 (2004). For each positive sample, 1 to 5 colonies were confirmed by specie-specific PCR (D'Agostino et al., 2004), and selected for PCR typing (Jersek et al. 1999). Results were analyzed by software Bionumerics (Applied Math - Belgium), producing a dendrorgam generated by UPGMA algorithm based on Dice similarity coefficient. Strains were considered identical if their similarity was above 95%. Results: L. monocytogenes was detected in: 10 feed, 3 feces, 1 milk and 8 dairy plant environments. Typing results, showed that all isolates were highly similar (74,5%): a set of 17 isolates (5 PCR profiles) retrieved from milk, feed, and dairy environments were genetically homogeneous (84,4%). Moreover, 2 strains were distributed across farms and 1 was persistent in processing environments. Significance: L. monocytogenes isolated from the selected production chain were genetically homogeneous, indicating a putative transmission. The absence of identity between genotypes retrieved from primary production and from processing environments, may be related to environment adaptation of the strains (Verghese et al. 2011). Milk may not be considered the primary source of L. monocytogenes. contamination of the producing plant, considering the low isolation frequency in milk. Thus the introduction of the pathogen may be due to non-compliances with Good Hygienic Practices (GHP) (i.e. unloading truck tanks). These findings strengthen the need for strict application of GHP in order to prevent contamination. Funds granted by Regione Piemonte - Direzione Sviluppo Agricoltura - 2007