title: Genetic characterization and antimicrobial resistance of Staphylococcus aureus isolated from bovine milk in Tunisia.
abstract: Staphylococcus aureus is a major agent of bovine mastitis in dairy herds, causing economic losses in dairy industry worldwide. In addition, milk and milk-products contaminated by Staph. aureus can cause harmful human diseases. The aim of this study was to characterize Staph. aureus strains isolated from dairy farms in Tunisia. Bulk tank milk (n = 32) and individual cow milk (n = 130) samples were collected during the period of 2013-2014. Forty-three Staph. aureus isolates were recovered and typed by spa typing, 16S-23S rRNA intergenic spacer (RS-PCR) and multiplex PCRs for 22 virulence genes. Antimicrobial resistance was also investigated with a disc diffusion test. A selected subsample of 22 strains was additionally genotyped by multilocus sequence typing. Seventeen spa types were recovered, and t2421 (n = 10), t521 (n = 6) and t2112 (n = 5) were the most common. Fourteen different RS-PCR genotypes grouped into 11 clusters were detected in our study, with predominance of the RVIgenotype (n = 24). Eight sequence types were identified and Clonal Complex 97, corresponding to RS-PCR cluster R, was the most common (n = 10), followed by CC1 (n = 4), CC15 (n = 3) and other four accounting for one or two strains. Different combinations of virulence genes were reported, and enterotoxin genes were present in few strains (seh, n = 4; sea, n = 2; sea and seh, n = 2; sec and sel, n = 2). The majority of strains were resistant only to penicillin; only one strain was found to be multiresistant and no methicillin-resistant Staph. aureus was demonstrated. Our study reported the isolation of CC97 from bovine milk in Tunisia for the first time and confirmed the relevance of this lineage in intramammary infection in cows.
  
  
  CONCLUSIONS
  This paper describes the characteristics of Staphylococcus aureus isolated from bulk tank and individual cow milk in Tunisia. All strains were genotyped by spa typing and RS-PCR, a method based on the amplification of the 16S-23S rRNA intergenic spacer region, and multiplex PCRs for 22 virulence genes. A selected subsample of strains was also genotyped by multilocus sequence typing. All strains were tested for antimicrobial resistance. Our study evidences a predominance of strains belonging to Clonal Complex 97. Methicillin-resistant strains were not detected, and overall low level of antimicrobial resistance was reported.
authors.affiliation: Université de Tunis El Manar, Institut de la Recherche Vétérinaire de Tunisie, Tunis, Tunisie.|Laboratory Wastewater Treatment, and Research Center of Water Technologies, Techno park Borj Cedria, Soliman, Tunisia.
authors.forename: M
authors.initials: M
authors.lastname: Ben Said
authors.affiliation: Université de Tunis El Manar, Institut de la Recherche Vétérinaire de Tunisie, Tunis, Tunisie.
authors.forename: M S
authors.identifier: http://orcid.org/0000-0001-9673-193X
authors.initials: MS
authors.lastname: Abbassi
authors.affiliation: Istituto Zooprofilattico Sperimentale della Lombardia e dell'Emilia Romagna, Lodi, Italy.
authors.forename: V
authors.initials: V
authors.lastname: Bianchini
authors.affiliation: Université de Tunis El Manar, Institut de la Recherche Vétérinaire de Tunisie, Tunis, Tunisie.|Laboratory Wastewater Treatment, and Research Center of Water Technologies, Techno park Borj Cedria, Soliman, Tunisia.
authors.forename: S
authors.initials: S
authors.lastname: Sghaier
authors.affiliation: Institute of Agricultural Biology and Biotechnology, National Research Council, Lodi, Italy.
authors.forename: P
authors.initials: P
authors.lastname: Cremonesi
authors.affiliation: Istituto Zooprofilattico Sperimentale della Lombardia e dell'Emilia Romagna, Lodi, Italy.
authors.forename: A
authors.initials: A
authors.lastname: Romanò
authors.affiliation: Piattaforma Genomica, Parco Tecnologico Padano, Lodi, Italy.
authors.forename: V
authors.initials: V
authors.lastname: Gualdi
authors.affiliation: Laboratory Wastewater Treatment, and Research Center of Water Technologies, Techno park Borj Cedria, Soliman, Tunisia.
authors.forename: A
authors.initials: A
authors.lastname: Hassen
authors.affiliation: Istituto Zooprofilattico Sperimentale della Lombardia e dell'Emilia Romagna, Lodi, Italy.
authors.forename: M V
authors.initials: MV
authors.lastname: Luini
mesh_terms: D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology*; D002417:Cattle; D024881:Drug Resistance, Bacterial; D004768:Enterotoxins / Q000235:genetics / Q000378:metabolism; D005260:Female; D005506:Food Contamination / Q000032:analysis; D005838:Genotype; D008414:Mastitis, Bovine / Q000382:microbiology*; D008892:Milk / Q000382:microbiology*; D058885:Multilocus Sequence Typing; D016133:Polymerase Chain Reaction; D013203:Staphylococcal Infections / Q000382:microbiology*; D013211:Staphylococcus aureus / Q000187:drug effects / Q000235:genetics* / Q000302:isolation & purification; D014416:Tunisia
chemical_list: D000900:Anti-Bacterial Agents; D004768:Enterotoxins
keywords: 
  Staphylococcus aureus
  ; CC97; antimicrobial resistance; bovine milk; genotyping; virulence genes