Safety characterization of Enterococcus isolated from kefir.

CARASI P; URDACI MC; DE ANTONI GL; SERRADELL MA

Rosario

Congreso; IX Congreso de SAMIGE; 2013

Kefir is a viscous beverage produced through fermentation of milk with mixed microbial population confined to ?kefir grains? as a starter culture. Many health-promoting properties have been associated with kefir-consumption. The genus Enterococcus belongs to the lactic acid bacteria (LAB). Enterococci can be found in a variety of foods and dairy products and are natural inhabitants of the gastrointestinal tract. Enterococcal strains are relevant for their technological properties, however, not all of them are recognized as safe. The assessment of Enterococcus for QPS (qualified presumption of safety) has been performed by European Food Safety Authority in 2010, reaching the conclusion that a strain-specific evaluation is necessary to assess the risk associated to the intentional use of enterococci in the food chain. The aim of this study was to isolate and characterize Enterococcus strains present in kefir grains and fermented milk. Six different kefir grains were used as starter cultures: CIDCA AGK1, AGK2, AGK3, AGK4, AGK5 and AGK11. Bacterial isolation was performed using disrupted kefir grains or milk fermented with kefir grains (10% w/w) during 24hs at 32°C. For direct isolation Bile Esculin Azide Agar was used meanwhile the enrichment step was performed in KF Streptococcus broth with BCP. For identification, the criterion of Bergey?s manual of systematic bacteriology was used combined with 16S rDNA sequencing and a species-specific PCR. The differential enumeration was performed on MRS supplemented with NaCl 6.5% w/v. Susceptibility to antimicrobials was evaluated by the disk diffusion method and the presence of virulence genes (ccf, espfm, gelE, cylA, agg, acm, van A, van B, van C2) was screened by PCR. Finally, resistance to simulated gastric and intestinal fluids and bacterial binding to mucus were assessed.The 36 Gram-positive coccoid bacteria isolated from kefir were identified as Enterococcus durans and they lacked all tested virulence genes. None of the strains was resistant to vancomycin, ciprofloxacin, erythromycin or linezolid. However, 5 of them were resistant to ampicillin. The quantity of these bacteria in kefir grains was lower than 30 CFU/g and 103-104 CFU/mL were quantified in fermented milk. Survival rates after exposure to simulated gastric and intestinal fluids ranged from 5.0 to 13.5 %, meanwhile, the degree of adhesion to mucus ranged from 6.9 to 22.7 %. We conclude that the presence of Enterococcus durans in kefir´s microbiota does not represent a health risk because of their susceptibility to clinically relevant antibiotics and the lack of genes associated with virulence. Moreover, their potential functionality was strengthened by their resistance to gastrointestinal conditions and their ability to adhere to mucus. Enterococcus strains isolated from kefir should be further studied to understand their role in the probiotic properties of this dairy product.