Reduction of Lactococcus lactis phage contamination in whey by means of membrane filtration: Impact of phage morphology and of bacterial host cells functioning as “phage fishing tool”

For dairy bacteriophages, the impact of phage morphology and whey components on phage retention efficiencies in filtration processes is not known. Therefore, the influence of (i) five differently shaped Lactococcus lactis phages as well as (ii) differently treated bacterial host cells added to native whey were analysed via cross-flow membrane filtration experiments with polyethersulfone membranes. Filtration efficiency with respect to phage retention and whey protein permeation was investigated. Compared with untreated native whey, phage retention was significantly higher (up to 4.8 log units) in the experiments using whey that was spiked with mechanically disrupted lactococcal host cells. However, the retention of phages commonly found in the dairy environment (i.e., members of the 936, c2, and P335 phage groups) did not correlate with the phage morphology. Results of this model study clearly demonstrate that phage titres can be reduced in whey (containing a sufficient protein concentration) by flat sheet membrane filtration processes.     

Lactococcus lactis phages from the perspective of their diversity,thermal and biocidal resistance

Lactic acid bacteria (LAB), particularly Lactococcus lactis, are of great significance in dairy fermentations. Many LAB strains are susceptible to attack by phages that affect their technological, biochemical and physiological functions. Phages of L. lactis are a serious concern because of the economic importance of this bacterium in the dairy industry. Members of L. lactis phages belonging to the P335, 936 and c2 groups are more problematic for the dairy industry. Many phages of the 936 group are resistant to various thermal and biocidal treatments commonly used in the dairy industry. This article reviews the diversity of L. lactis bacteriophages of the P335, 936 and c2 groups and discusses their interaction with their bacterial hosts. In addition, this review provides an overview of the resistance of L. lactis phages to thermal treatments and chemical biocides, and highlights some novel strategies to destroy these phages.     

Characterisation of technologically proficient wild Lactococcus lactis strains resistant to phage infection

The aim of this work was to establish whether Lactococcus lactis strains isolated from spontaneous dairy fermentations exhibited useful milk-processing capabilities and resistance to bacteriophage infection in order to be used as components in starter formulations. The 33 out of 100 isolates of L. lactis, originated from farmhouse cheeses, were found to be resistant to a collection of 34 phages belonging to the c2 and 936 groups. Six of the isolates were discarded as potential starters because they were lysogenic and other five because they produced tyramine. Plasmid and chromosomal profiles of the 22 remaining isolates allowed their classification into 16 different strains. All of these were good lactic acid producers from lactose, moderately proteolytic and, in eight cases, diacetyl production from citrate was observed. The mechanism(s) leading to the phenotype of phage resistance was identified for all the strains used in this study. Inhibition of adsorption was the most frequent one, although genetic determinants for some abortive infection systems were also detected (abiB, abiG and abiI). Frequently, more than one mechanism was present in the same strain. One of the strains, L. lactis IPLA542, was selected as a model starter for pilot fermentations. It clotted milk normally both in the absence and in the presence of phage at concentrations that completely abolished the process when promoted by a phage-susceptible strain.     

Salmonella bacteriophage diversity reflects host diversity on dairy farms

Salmonella is an animal and human pathogen of worldwide concern. Surveillance programs indicate that the incidence of Salmonella serovars fluctuates over time. While bacteriophages are likely to play a role in driving microbial diversity, our understanding of the ecology and diversity of Salmonella phages is limited. Here we report the isolation of Salmonella phages from manure samples from 13 dairy farms with a history of Salmonella presence. Salmonella phages were isolated from 10 of the 13 farms; overall 108 phage isolates were obtained on serovar Newport, Typhimurium, Dublin, Kentucky, Anatum, Mbandaka, and Cerro hosts. Host range characterization found that 51% of phage isolates had a narrow host range, while 49% showed a broad host range. The phage isolates represented 65 lysis profiles; genome size profiling of 94 phage isolates allowed for classification of phage isolates into 11 groups with subsequent restriction fragment length polymorphism analysis showing considerable variation within a given group. Our data not only show an abundance of diverse Salmonella phage isolates in dairy farms, but also show that phage isolates that lyse the most common serovars causing salmonellosis in cattle are frequently obtained, suggesting that phages may play an important role in the ecology of Salmonella on dairy farms.     

Differentiation of Streptococcus thermophilus strains in commercial Direct Vat Set yoghurt starter

Based on inhibition of acid production, 100 individual colonies of Streptococcus thermophilus isolated from commercial Direct Vat Set yoghurt starter DVS-M were typed into 2 groups of sensitive and insensitive to phage φ001, which was isolated from the lysis culture of the turbidity test. The ratio between the sensitive and insensitive isolates in DVS-M was calculated at 3:2. Whole-cell protein profile (WCPP) and randomly amplified polymorphic DNA (RAPD) analysis showed that both WCPP and RAPD patterns between sensitive and insensitive isolates were distinct, whereas isolates with the same sensitivity or insensitivity shared common WCPP and RAPD patterns. It suggested that all the sensitive isolates belong to one S. thermophilus strain and all the insensitive isolates belong to another one. This study revealed that DVS-M yoghurt starter consists of 2 S. thermophilus strains with different phage sensitivity.     

Molecular subtyping and virulence gene analysis of Listeria monocytogenes isolates from food

A total of 67 Listeria monocytogenes isolates from 698 raw meat samples were characterized for molecular serogroup identification and antimicrobial susceptibility. Approximately one third (32.8%) of the isolates belonged to molecular serogroup 1/2a, 3a, followed by 1/2c, 3c (26.9%), 1/2b, 3b, 7 (22.4%), 4b, 4d, 4e (16.4%) and 4a, 4c (1.5%). Most of the L. monocytogenes isolates were susceptible to 14 antimicrobials tested but several were resistant to tetracycline, ciprofloxacin and nitrofurantoin. An additional 30 L. monocytogenes isolates from chicken and produce in our collection were also included to determine the presence of significant virulence markers. All 97 isolates carried inlC and inlJ except for a lineage III isolate 110-1. Most Listeriolysin S (LLS)-carrying isolates (11/12) belonged to lineage I, whereas the remaining one isolate belonged to lineage III. Five 4b, 4d, 4e isolates including two from turkey and three from produce belonged to Epidemic Clone I (ECI). Four molecular serogroup associated mutation types that lead to premature stop codons (PMSCs) in inlA were identified. PFGE and inlA sequence analysis results were concordant, and different virulence potential within 1/2a, 3a and 4b, 4d, 4e isolates were observed. The study revealed that a subset of isolates from meat and produce belonged to ECI, harbored inlC, inlJ and LLS, and produced full length InlA, suggesting that they be capable of causing human illness.     

Biodiversity of lactococcal bacteriophages isolated from 3 Gouda-type cheese-producing plants

This study reports on the identification and characterization of bacteriophages isolated from cheese-production facilities that use undefined, mixed starter cultures. Phage screening was carried out on whey samples isolated from 3 factories, 2 utilizing one particular undefined starter mixture and 1 utilizing another undefined starter mixture. Phage screening was carried out using 40 strains isolated from the 2 mixed starter cultures, and phages were profiled using host range, electron microscopy, multiplex PCR, and DNA restriction analysis. Twenty distinct lactococcal phages were identified based on host range and DNA restriction profiles, all belonging to the 936-type phage species. Nineteen of these phages were found to be able to infect both recognized subspecies of Lactococcus lactis. Restriction of phage DNA isolated using a newly developed guanidinium thiocyanate disruption method showed that the genomes of the 20 isolated phages were between 26 and 31 kb in size. It is evident from this study that the use of mixed starters creates an ideal environment for the proliferation of different phages with slightly varying host ranges. Furthermore, in this environment, members of the 936-type phage species clearly dominated the phage population.     

Analysis of the complete genome sequence of the lactococcal bacteriophage bIBB29

Bacteriophage bIBB29 was isolated from a whey sample originating from an industrial biotechnological process, disturbed by a bacteriophage attack. Phage bIBB29 was determined to be active against three phage-resistant strains of Lactococcus lactis. It belongs to the 936 species containing virulent phages with isometric head and short non-contractile tail. One-step growth kinetics of bIBB29 phage showed that its latent time was 23 min, and the burst size was about 130 bacteriophages. The complete nucleotide sequence of the virulent L. lactis bacteriophage bIBB29 comprises 29305 nucleotides and is the sixth phage genome of the 936 species published until now. The G+C content of the bIBB29 genome (34.7%) is similar to that of its host and also to that of other phages from the 936 species. The bIBB29 genome counts 54 open reading frames organized in three typical clusters, corresponding to the early, middle and late expressed genes. Only 20 protein products of the predicted genes were found to have their homologs among proteins with known function. The early expressed region in the genomes of 936 group members displays the highest divergence, whereas the late and middle regions share high similarities, with the exception of five genes. The genome of bIBB29 shares the highest overall nucleotide similarity with bIL170 (87%), and the lowest with phage 712 (77%). The host range analysis showed that despite the high level of similarity between the receptor binding protein (RBP) of phage bIBB29 and P475, they have a different host range. This implies that RBP is not a sufficient factor for host range.     

Persistent Listeria monocytogenes subtypes isolated from a smoked fish processing facility included both phage susceptible and resistant isolates

Contamination of Ready-To-Eat foods with Listeria monocytogenes can typically be traced back to post-processing contamination from environmental sources; contamination is often linked to subtypes that persist in food associated environments. Although phage-based biocontrol strategies have been proposed for controlling this pathogen, information on the efficacy of phage treatment against diverse L. monocytogenes subtypes from food associated environments is still limited. We identified subtypes that were repeatedly found (“persistent”) in a smoked fish processing facility by using EcoRI ribotyping data for isolates obtained in 1998–2009. PFGE analysis of 141 isolates (9 ribotypes) supported persistence for up to 11 years. Characterization of selected isolates, representing persistent subtypes, against a panel of 28 listeriaphages showed a wide range of likelihood of phage susceptibility, ranging from 4.6% (for 7 ribotype DUP-1043A isolates) to 95.4% (for 7 ribotype DUP-1044A isolates). In challenge studies with 10⁵ and 10⁶ CFU/ml L. monocytogenes, using phage cocktails and a commercial phage product at different phage-host ratios, one isolate (ribotype DUP-1043A) was not affected by any treatment. A reduction in L. monocytogenes counts of up to 4 log units was observed, after 8 h of treatment, in isolates of two ribotypes, but subsequent re-growth occurred. Survivor isolates obtained after 24 h of treatment showed decreased susceptibility to individual phages included in the phage cocktail, suggesting rapid emergence of resistant subtypes.     

Comparison of antimicrobial resistance patterns and phage types of Salmonella Typhimurium isolated from pigs,pork and humans in Belgium between 2001 and 2006

Infections with non-typhoid Salmonella represent a major problem in industrialized countries.The emergence and spread of antimicrobial-resistant pathogens, among them Salmonella, has become a serious health hazard worldwide. One of the most commonly isolated non-typhoid Salmonella serovars in pigs, pork and humans is Salmonella Typhimurium. In this study the comparison of the incidences of resistance to nine antimicrobials, resistance patterns and phage types between S. Typhimurium isolated from pigs (n = 581), pork (n = 255) and humans (n = 1870) in Belgium in the period 2001 to 2006 was performed.Resistance to the antimicrobials ampicillin, chloramphenicol, streptomycin, sulfonamides and tetracycline was frequently observed and varied between 23.5% and 83.1%. Resistance ranged from 15.6% to 20.7% for the combination trimethoprim–sulfonamides and from 3.4% to 5.8% for nalidixic acid. Resistance to the critical important antimicrobials cephalosporins and fluoroquinolones was found sporadically (≤ 1.2%). Resistance to the different antimicrobials was observed to be similar in S. Typhimurium isolates from the various origins. Twenty-seven antimicrobial resistance patterns representing in total 75.2%, 89.0% and 89.6% of the isolates from pigs, pork and humans respectively were found to be common among the three groups and 73 combinations antimicrobial resistance pattern/phage type were found to be common among pork and human isolates, representing 70.1% of the pork isolates and 51.0% of the human isolates. The high percentage of isolates that have a common resistance pattern, and in a less pronounced way a common combination phage type/resistance pattern, are in agreement with the hypothesis of transfer of antimicrobial resistant Salmonella from pigs via the consumption of pork to humans as one of the possible pathways. The most prevalent combination in Belgium within both the pork isolates (7.4%) and the human isolates (13.2%) was S. Typhimurium DT104 resistant to ampicillin, chloramphenicol, streptomycine, sulfonamides and tetracycline.     

Lactococcal 936-species phage attachment to surface of Lactococcus lactis

The interactions of the 936-species phages sk1, jj50, and 64 with the cell surface of Lactococcus lactis LM0230 were analyzed. Cell envelopes (walls + plasma membrane), cell wall, or plasma membrane from L. lactis ssp. lactis LM0230 each inactivated the phages in vitro. However, other 936-species phages kh and P008, which do not infect strain LM0230, were not inactivated by any of the subcellular fractions. Treating cell walls or plasma membrane with the cell wall hydrolase mutanolysin eliminated inactivation of phage sk1. This suggested that intact cell wall fragments were required for inactivation. A role for plasma membrane in phage sk1 inactivation was further investigated. Boiling, washing in 2 M KCl, 8 M urea, or 0.1 M Na(2)CO(3)/pH 11, or treating the plasma membrane with proteases did not reduce adsorption or inactivation of phage. Adding lipoteichoic acid or antibodies to lipoteichoic acid did not reduce inactivation of phage in a mixture with membrane, suggesting that lipoteichoic acid was not involved. Inactivation by envelopes or cell wall correlated with ejection of DNA from the phage sk1 capsid. Although calcium is required for plaque formation, it was not required for adsorption, inactivation, or ejection of phage DNA by envelopes or cell wall. The results suggest that at least for phages sk1, jj50, and 64, adsorption and phage DNA injection into the host does not require a host membrane protein or lipoteichoic acid, and that cell wall components are sufficient for these initial steps of phage infection.     

Novel conjugative plasmids from the natural isolate Lactococcus lactis subspecies cremoris DPC3758: a repository of genes for the potential improvement of dairy starters

A collection of 17 natural lactococcal isolates from raw milk cheeses were studied in terms of their plasmid distribution, content, and diversity. All strains in the collection harbored an abundance of plasmids, including Lactococcus lactis ssp. cremoris DPC3758, whose 8-plasmid complement was selected for sequencing. The complete sequences of pAF22 (22,388 kb), pAF14 (14,419 kb), pAF12 (12,067 kb), pAF07 (7,435 kb), and pAF04 (3,801 kb) were obtained, whereas gene functions of technological interest were mapped to pAF65 (65 kb) and pAF45 (45 kb) by PCR. The plasmids of L. lactis DPC3758 were found to encode many genes with the potential to improve the technological properties of dairy starters. These included 3 anti-phage restriction/modification (R/M) systems (1 of type I and 2 of type II) and genes for immunity/resistance to nisin, lacticin 481, cadmium, and copper. Regions encoding conjugative/mobilization functions were present in 6 of the 8 plasmids, including those containing the R/M systems, thus enabling the food-grade transfer of these mechanisms to industrial strains. Using cadmium selection, the sequential stacking of the R/M plasmids into a plasmid-free host provided the recipient with increased protection against 936- and c2-type phages. The association of food-grade selectable markers and mobilization functions on L. lactis DPC3758 plasmids will facilitate their exploitation to obtain industrial strains with enhanced phage protection and robustness. These natural plasmids also provide another example of the major role of plasmids in contributing to host fitness and preservation within its ecological niche.     

Screening for and characterization of Lactococcus lactis bacteriophages with high thermal resistance

Fifty-six Lactococcus lactis phage isolates collected from different German dairies and obtained from a starter culture manufacturer were tested for their heat resistance. About 40% of these isolates resisted treatment at 80 °C for 5 min when they were heated in milk. The most resistant phage isolate, P1532, was collected from sour cream. Plaque-formation was still detectable even after heating at 97 °C for 5 min. The second heat-resistant one, P680, showed some plaque-forming ability after heating at 95 °C for 5 min. Kinetic parameters for the thermal inactivation of these two resistant phages were determined for temperatures ranging from 70 to 97 °C. The inactivation of phage P1532 in skim milk and in buffer medium were found to follow first-order kinetics and did not exhibit tailing, whereas in the inactivation curves of phage P680 tailing was observed. The D-value of P1532 at pasteurization temperature of 72 °C was calculated as 112 min.     

Rhyzopertha dominica adult mortalities after exposure to indigenous Beauveria bassiana isolates from stored-grain pests: Effects of certain factors in sampling process

In search for potential entomopathogenic fungi for microbial control of Rhyzopertha dominica, 75 Beauveria bassiana isolates previously recovered from stored-grain insects sampled in Adana and Mersin provinces of Turkey were tested and the mortality levels were used to evaluate possible effects of three sampling factors on encountering potential isolates. In the bioassays, twenty adults were exposed to 1000 ppm spores mixed into wheat kernels and adult mortalities after 14 days were used for statistical analysis. The relations between mortalities and 1) species of host insects from which isolates were obtained, 2) collection season –summer or autumn– of host insects from storage facilities, and 3) time of host insect's death –before or after collection– were evaluated. Considerable variations in mortalities were detected (13.67%–100%) depending on isolates. Amongst twelve Adana isolates, isolate 155657 caused the highest mortality. Eleven Mersin isolates were found to be effective against R. dominica. There was no correlation between R. dominica mortality and insect species from which isolates were obtained. Statistically, the frequency of potential isolates was higher in Adana samples collected in summer months compared to that in autumn months; however, sampling season did not have a significant effect in Mersin. The frequency of potential B. bassiana isolates was significantly higher when isolation was made from hosts that died in laboratory after sample collection compared to those isolated from hosts already dead at the time of collection. This study demonstrated that stored-grain insects are a good source of finding various B. bassiana isolates with different efficacy levels. At least for stored-grain insects, specifically sampling targeted species does not significantly increase the chance of finding highly potential isolates. However, retaining alive sampled insects can increase the likelihood of encountering isolates with high efficacy.     

Molecular characteristics and virulence potential of Listeria monocytogenes isolates from Chinese food systems

In this study, we examined Listeria monocytogenes isolates from Chinese food sources in an attempt to gain further insights on the molecular characteristics and virulence potential of this important foodborne pathogen. Of the 88 L. monocytogenes food isolates recovered, 42 (47.7%) were of serovars 1/2a or 3a; 23 (26.1%) of serovars 1/2b or 3b; 15 (17.0%) of 1/2c or 3c; 6 (6.8%) of serovars 4b, 4d or 4e; and 2 (2.2%) of serovars 4a or 4c. In contrast to inlAB locus conserved in all serovars, internalin cluster between ascB and dapE varies with different serovars, with inlC2DE, inlGC2DE and inlGHE predominantly in serovars 1/2b or 4b, serovar 1/2a and serovar 1/2c. While inlF existed in all the inlGHE- and inlGC2DE-containing isolates but 17.4% of those having inlC2DE, lmo2026 existed in all the inlGHE-containing isolates but 20.0% of those bearing inlGC2DE, suggesting that inlF might have co-evolved with inlGC2DE and inlGHE while lmo2026 with inlGHE only. With the exception of serovar 4a isolate, most serovar isolates demonstrated remarkable ability to form plaques on L929 cells and produced significant mouse mortality irrespective of the internalin gene organization and whether an intact actA gene is present or not. These results indicate that majority of these food isolates may have the potential to cause human diseases if ingested via contaminated foods. Given that serovar 4b accounts for nearly half of human clinical listeriosis cases documented, the relative low proportion of serovar 4b food isolates suggests that this serovar is probably more tolerant of the adverse conditions in the host's stomach and/or more efficient in entering host cells than serovars 1/2a, 1/2b and 1/2c.     

Identification and antimicrobial resistance of Streptococcus uberis and Streptococcus parauberis isolated from bovine milk samples

The conventional identification of Streptococcus uberis/parauberis group (n = 137) in clinical and subclinical bovine mastitis samples originating from 111 different farms was compared with identification based on 16 and 23S rRNA gene HindIII RFLP patterns used as operational taxonomic units in numerical analyses. On the basis of ribopattern analysis only 2 isolates belonged to S. parauberis, which is thus not a frequent cause of bovine intramammary infections in Finland. According to in vitro antimicrobial susceptibility testing, Streptococcus uberis is susceptible to β-lactam antibiotics. The prevalence of erythromycin (15.6%) and oxytetracycline (40.6%) resistance of clinical S. uberis isolates was higher than reported previously among subclinical isolates. The 2 subclinical S. parauberis isolates were susceptible to all the antimicrobials tested.     

Expanding the diversity of oenococcal bacteriophages: Insights into a novel group based on the integrase sequence

Temperate bacteriophages are a contributor of the genetic diversity in the lactic acid bacterium Oenococcus oeni. We used a classification scheme for oenococcal prophages based on integrase gene polymorphism, to analyze a collection of Oenococcus strains mostly isolated in the area of Bordeaux, which represented the major lineages identified through MLST schemes in the species. Genome sequences of oenococcal prophages were clustered into four integrase groups (A to D) which were related to the chromosomal integration site. The prevalence of each group was determined and we could show that members of the int_B- and int_C-prophage groups were rare in our panel of strains. Our study focused on the so far uncharacterized members of the int_D-group. Various int_D viruses could be easily isolated from wine samples, while int_D lysogens could be induced to produce phages active against two permissive O. oeni isolates. These data support the role of this prophage group in the biology of O. oeni. Global alignment of three relevant int_D-prophages revealed significant conservation and highlighted a number of unique ORFs that may contribute to phage and lysogen fitness.     

Analytical specificity and sensitivity of a real-time polymerase chain reaction assay for identification of bovine mastitis pathogens

Intramammary infection (IMI), also known as mastitis, is the most frequently occurring and economically the most important infectious disease in dairy cattle. This study provides a validation of the analytical specificity and sensitivity of a real-time PCR-based assay that identifies 11 major pathogen species or species groups responsible for IMI, and a gene coding for staphylococcal β-lactamase production (penicillin resistance). Altogether, 643 culture isolates originating from clinical bovine mastitis, human, and companion animal samples were analyzed using the assay. The isolates represented 83 different species, groups, or families, and originated from 6 countries in Europe and North America. The analytical specificity and sensitivity of the assay was 100% in bacterial and β-lactamase identification across all isolates originating from bovine mastitis (n = 454). When considering the entire culture collection (including also the isolates originating from human and companion animal samples), 4 Streptococcus pyogenes, 1 Streptococcus salivarius, and 1 Streptococcus sanguis strain of human origin were identified as Streptococcus uberis, and 3 Shigella spp. strains were identified as Escherichia coli, decreasing specificity to 99% in Strep. uberis and to 99.5% in E. coli. These false-positive results were confirmed by sequencing of the 16S rRNA gene. Specificity and sensitivity remained at 100% for all other bacterial targets across the entire culture collection. In conclusion, the real-time PCR assay shows excellent analytical accuracy and holds much promise for use in routine bovine IMI testing programs. This study provides the basis for evaluating the assay's diagnostic performance against the conventional bacterial culture method in clinical field trials using mastitis milk samples.     

Characterization of lactococcal bacteriophages isolated from Slovenian dairies

Summary This is the first study on lactic acid bacteria bacteriophages isolated from dairy plants in Slovenia. Over a period of 2 years, thirteen lactococcal phages were isolated from the whey samples taken in three dairy plants. Phages were characterized by morphological properties, host range, restriction patterns, genome size and similarity of phage protein genes. All phages belonged to the Siphoviridae family. Five phages had prolate heads (52–61 × 40–52 nm) and 85–120 nm long non‐contractile tails (morphotype B2). Eight phages had small isometric heads (46–63 nm in diameter) and long non‐contractile tails (morphotype B1). The phages with the prolate heads showed a broader host range than the small isometric‐headed phages. The genome sizes of prolate phages were estimated between 15.9 and 16.3 kb while the genome sizes of small isometric‐headed phages were between 20.0 and 39.9 kb. Based on DNA restriction patterns, genome sizes and multiplex PCR analysis prolate phages were classified into the c2 group and were similar to the phage type c6A while all of the phages with small isometric heads were found to be related to the phage species 936.     

Isolation and identification of Enterococcus faecium from seafoods: Antimicrobial resistance and production of bacteriocin-like substances

A collection of isolates from uncooked seafoods (molluscs, fish, and fish fillets) were identified as Enterococcus faecium species and studied in further detail. Isolates were clustered in well-defined genomic groups according to food origin after ERIC-PCR analysis. Four isolates (FR 1-2, FB 1-3-B, FB 3-1, FTA 1-2) decarboxylated lysine, ornithine, and tyrosine. Isolate FR 1-2 also decarboxylated histidine. Most isolates were sensitive to antibiotics of clinical use, but resistance was detected more frequently towards nitrofurantoin (50%), erythromycin (33.33%) or rifampicin (33.33%) to quinupristin/dalfopristin (12.5%). Resistance to β-lactams or vancomycin was not detected. The enterococcal antigen A was the presumed virulence trait detected most frequently. None of isolates carried haemolysin/cytolysin genes. Twelve isolates produced anti-listerial activity. Among them, seven isolates also produced bacteriocin-like inhibitory substances against other enterococci, and one isolate was also able to inhibit Staphylococcus aureus. Three isolates only were active against Listeria monocytogenes, and two only were active against enterococci. One bacteriocinogenic isolate carried the enterocin A structural gene, but genes corresponding to other enterocins (EntB, EntP, EntQ, Ent1071, EntL50A/EntL50B, and Ent31) were not detected. Bacteriocin-producing enterococci lacking undesirable traits (such as antibiotic resistance or biogenic amine production) or their produced bacteriocins could be potential candidates to aid in preservation of seafoods and other food products as well.