Browsing by Author "Pinto, Ana Luísa"
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- Characterization of anti-listeria bacteriocins isolated from shellfish: Potential antimicrobials to control non-fermented seafoodPublication . Pinto, Ana Luísa; Fernandes, Melissa; Pinto, Cristina; Albano, Helena; Castilho, Fernanda; Teixeira, Paula; Gibbs, Paul A.This work had as main objectives to characterize two bacteriocins produced by lactic acid bacteria (LAB) previously isolated from non-fermented seafood, in order to evaluate their potential as new food protective agents. The two bacteriocinogenic isolates were identified by Polymerase Chain Reaction (PCR) using genusand species-specific primers, and confirmed by 16S rDNA sequencing, as Enterococcus faecium and Pediococcus pentosaceus. The antimicrobial spectrum of each strain included several indicator microorganisms, some of them also isolated from seafood. Growth of Listeria innocua, L. monocytogenes, Staphylococcus aureus, Bacillus cereus and other LAB species were inhibited, although no inhibition of Gram-negative microorganisms was observed. Proteolytic, but not lipolytic or glycolytic enzymes, completely inactivated the antimicrobial effect of both cell-free supernatants confirming the proteinaceous nature of the inhibitors. The antimicrobial activity was maintained after treatment with NaCl, SDS, Triton X-100, Tween 20, Tween 80 and EDTA after 2 h or 5 h of exposure and both bacteriocins were stable over a wide range of pH and temperatures. Production of bacteriocin by E. faecium (bacALP7)was detected initially at exponential phase and reached a maximum activity of 25,600 AU/ml in the early stationary phase, whereas bacteriocin production by P. pentosaceus ALP57 (bacALP57) reached the maximum at exponential phase with 12,800 AU/ml. The bacteriocins did not kill L. monocytogenes ESB54 nor L. innocua 2030c however, cellular growth was reduced. The partially purified bacteriocins, bacALP7 and bacALP57, were below 6.5 kDa in size as determined by Tricine-SDS gel electrophoresis. E. faecium and P. pentosaceus contained DNA fragments corresponding in size to those recorded for enterocin B and pediocin PA-1, respectively. Sequencing of the fragments from both bacteriocins confirmed the homology. To our knowledge, for the first time two LAB producing bacteriocins similar to pediocin PA-1 and enterocin B, were isolated from non-fermented shellfish. The adaptation of the cultures to seafood matrices may be advantageous in terms of application as a biopreservation strategy for reduction of L. monocytogenes levels in seafood products.
- Prevalence and serotyping of Listeria monocytogenes in Portuguese live bivalve molluscs sampled in various steps along the sanitary control processPublication . Pinto, Ana Luísa; Teixeira, Paula; Castilho, Fernanda; Felício, Maria Teresa; Pombal, FilomenaThe prevalence and contamination levels of Listeria monocytogenes were investigated in live bivalve molluscs for human consumption, collected in various steps of the commercial and sanitary circuits in the North of Portugal. Samples of di¡erent species were collected per lot before and after depuration treatment in two depuration units and further, when placed in retail markets. Listeria monocytogenes was isolated from 4% of the samples although with very low contamination levels (less than 100MPNg 1). None of the environmental (non-depurated) samples were contaminated with the food-borne pathogen. The positive samples involved depurated and commercialized samples from two lots, one in each circuit. Two serotypes (1/2a and 3b) were detected among the isolated strains. This study demonstrated the possibility of occurrence of L.monocytogenes contamination of live bivalvemolluscs along the sanitary control circuit, including at the retailmarket level. As L. monocytogenes grows well at refrigerated temperatures and in high salt matrixes such as seafood its presence in these products should not be overlooked.