Antilisterial activity of raw sheep milk from two native Epirus breeds: Culture-dependent identification, bacteriocin gene detection and primary safety evaluation of the antagonistic LAB biota.

Raw milk from native small ruminant breeds in Epirus, Greece, is a valuable natural source of autochthonous lactic acid bacteria (LAB) strains with superior biotechnological properties. In this study, two bulk milks (RM1, RM2) from two local sheep yards, intended for traditional Kefalotyri cheese production, were preselected for bacteriocin-like antilisterial activity by in vitro tests. Their antagonistic LAB biota was quantified followed by polyphasic (16S rRNA gene sequencing; IGS for ; a multiplex-PCR for ) identification of 42 LAB (RM1/18; RM2/24) isolates further evaluated for bacteriocin encoding genes and primary safety traits.

Assessment of the Spoilage Microbiota during Refrigerated (4 °C) Vacuum-Packed Storage of Fresh Greek Anthotyros Whey Cheese without or with a Crude Enterocin A-B-P-Containing Extract.

Although fresh whey cheeses are prone to rapid deterioration, mainly by psychrotrophic Gram-negative bacteria and lactic acid bacteria (LAB), data on the specific spoilage species in traditional Greek whey cheeses are scarce. Therefore, this study quantified growth and characterized the primary spoilage bacteria in fresh Anthotyros whey cheeses stored at 4 °C in a vacuum for 40 days, without or with an added 5% (/) of an enterocin A-B-P crude extract (CEntE). Psychrotrophic spp.

Growth Inhibitory and Selective Pressure Effects of Sodium Diacetate on the Spoilage Microbiota of Frankfurters Stored at 4 °C and 12 °C in Vacuum.

This study evaluated microbial growth in commercial frankfurters formulated with 1.8% sodium lactate (SL) singly or combined with 0.25% sodium diacetate (SDA), vacuum-packaged (VP) and stored at 4 °C and 12 °C.

Monitoring Growth Compatibility and Bacteriocin Gene Transcription of Adjunct and Starter Lactic Acid Bacterial Strains in Milk.

Abstract: When developing protective starter cultures for application in cheese technologies, monitoring growth interactions between starter and adjunct lactic acid bacterial (LAB) species and in situ expression of bacteriocin genes in the mixtures is crucial. This study first aimed to monitor the growth of mixed LAB strain populations during milk model fermentations by microbial counts and real-time quantitative PCR. The primary starter strains, Streptococcus thermophilus ST1 and costarter Lactococcus lactis subsp.

Cell Growth Density and Nisin A Activity of the Indigenous Lactococcus lactis subsp. cremoris M78 Costarter Depend Strongly on Inoculation Levels of a Commercial Streptococcus thermophilus Starter in Milk: Practical Aspects for Traditional Greek Cheese Processors.

Abstract: Mixed thermophilic and mesophilic commercial starter cultures (CSCs), particularly those including Streptococcus thermophilus as a primary milk acidifier, have been found to reduce growth and counteract in situ nisin A (NisA+) antilisterial effects by the novel, indigenous Lactococcus lactis subsp. cremoris M78 costarter in traditional Graviera thermized milk cheese curds. Therefore, this model challenge study evaluated growth and in situ NisA+ activity of strain M78 in coculture with S.

Structural enterocin gene profiles and mode of antilisterial activity in synthetic liquid media and skim milk of autochthonous Enterococcus spp. isolates from artisan Greek Graviera and Galotyri cheeses.

The presence of eight common structural enterocin genes, singly or in varying combinations, in the genome of 15 antagonistic Enterococcus spp. previously isolated from artisan Greek Graviera and Galotyri retail cheeses was tested and associated with the mode of enterocin (Ent) antilisterial activity of each isolate in three liquid culture media. The isolates were assigned to nine distinct strain genotypes of E.

Major ecological shifts within the dominant nonstarter lactic acid bacteria in mature Greek Graviera cheese as affected by the starter culture type.

Traditional Greek Graviera cheese is often produced from thermized milk to control undesirable bacterial contaminants. Since thermization also reduces the desirable lactic acid bacteria (LAB) microbiota of raw milk, natural undefined or commercially defined starters are used. This study evaluated effects of the type of starter added to bulk thermized milk on the microbiology of mature (day-90) Graviera cheese.

Approaches for enhancing in situ detection of enterocin genes in thermized milk, and selective isolation of enterocin-producing Enterococcus faecium from Baird-Parker agar.

Enterococci are naturally selected for growth in thermized ewes'/goats' milk mixtures used for traditional cooked hard cheese processing in Greece. A culture-independent PCR-based approach was applied to detect the presence of enterocin-encoding genes in naturally culture-enriched thermized milk (TM). Portions of TM (63 °C, 30 s) collected from a commercial cheese plant before addition of starters were fermented at 37 °C for 48 h to facilitate growth of indigenous enterococci.

Hurdle factors minimizing growth of while counteracting antilisterial effects of a novel nisin A-producing subsp. costarter in thermized cheese milks.

The capacity of growth, survival, and adaptive responses of an artificial contamination of a three-strain cocktail in factory-scale thermized (65 °C, 30 s) Graviera cheese milk (TGCM) was evaluated. Bulk TGCM samples for inoculation were sequentially taken from the cheese making vat before process initiation (CN-LM) and after addition of a commercial starter culture (CSC), the CSC plus the nisin A-producing (NisA+) costarter strain subsp. M78 (CSC + M78), and all ingredients with the rennet last (CSC + M78-RT).

Growth, nisA Gene Expression, and In Situ Activity of Novel Lactococcus lactis subsp. cremoris Costarter Culture in Commercial Hard Cheese Production.

This study evaluated in situ expression of the nisA gene by an indigenous, nisin A-producing (NisA+) Lactococcus lactis subsp. cremoris raw milk genotype, represented by strain M78, in traditional Greek Graviera cheeses under real factory-scale manufacturing and ripening conditions. Cheeses were produced with added a mixed thermophilic and mesophilic commercial starter culture (CSC) or with the CSC plus strain M78 (CSC+M78).

Enhanced Control of Listeria monocytogenes by Enterococcus faecium KE82, a Multiple Enterocin-Producing Strain, in Different Milk Environments.

Enterococcus faecium KE82, isolated from traditional Greek Graviera cheese, was identified in pure broth cultures in vitro as a multiple enterocin-producing bacterial strain possessing the structural entA, entB, and entP enterocin genes. E. faecium KE82 was further assessed for in situ antilisterial activity in raw milk (RM) and commercially thermized milk (TM; 63°C for 30 s) in the presence of the indigenous microbiota and in sterile raw milk (SRM; 121°C for 5 min) with or without the addition of two commercial starter culture (CSC) strains Streptococcus thermophilus and Lactococcus lactis .

Growth interactions and antilisterial effects of the bacteriocinogenic Lactococcus lactis subsp. cremoris M104 and Enterococcus faecium KE82 strains in thermized milk in the presence or absence of a commercial starter culture.

Traditional Greek cheeses are often produced from thermized milk (TM) with the use of commercial starter cultures (CSCs), which may not inhibit growth of Listeria monocytogenes completely. Therefore, this study evaluated the behavior of an artificial L. monocytogenes contamination in commercially TM (63 °C; 30 s) inoculated with a CSC plus Lactococcus lactis subsp.

Behavior of Staphylococcus aureus in culture broth, in raw and thermized milk, and during processing and storage of traditional Greek Graviera cheese in the presence or absence of Lactococcus lactis subsp. cremoris M104, a wild, novel nisin A-producing raw milk isolate.

This study was conducted to evaluate the behavior of Staphylococcus aureus during processing, ripening, and storage of traditional Greek Graviera cheese in accordance with European Union Regulation 1441/2007 for coagulase-positive staphylococci in thermized milk cheeses. Lactococcus lactis subsp. cremoris M104, a wild, novel nisin A-producing (NisA+) strain, also was evaluated as an antistaphylococcal adjunct.

Characterization of a wild, novel nisin a-producing Lactococcus strain with an L. lactis subsp. cremoris genotype and an L. lactis subsp. lactis phenotype, isolated from Greek raw milk.

Several molecular taxonomic studies have revealed that many natural (wild) Lactococcus lactis strains of dairy origin which are phenotypically representative of the L. lactis subspecies lactis cluster genotypically within subspecies cremoris and vice versa. Recently, we isolated two wild nisin-producing (Nis(+)) L.

FTIR-based polyphasic identification of lactic acid bacteria isolated from traditional Greek Graviera cheese.

This study used a combination of phenotypic, physical (Fourier Transformed Infra-Red [FTIR] spectroscopy) and molecular (RFLP and SSCP analysis of 16S rRNA genes) methods to identify the lactic acid bacteria (LAB) flora present in traditional Greek Graviera cheese after five weeks of ripening. A total of 300 isolates collected from high dilution plates of TSAYE (incubated at 30 °C), M-17 (22 °C) and M-17 (42 °C) agar media were clustered by FTIR and then representative strains of each cluster were cross-identified blindly by all methods. Based on their FTIR spectra, 282 isolates were LAB grouped in 28 clusters.

Microbial stability and safety of traditional Greek Graviera cheese: characterization of the lactic acid bacterial flora and culture-independent detection of bacteriocin genes in the ripened cheeses and their microbial consortia.

The microflora of four batches of traditional Greek Graviera cheese was studied at 5 weeks of ripening, and 200 lactic acid bacteria (LAB) isolates were phenotypically characterized and screened for antilisterial bacteriocins. The cheeses were also analyzed for organic acids by high-performance liquid chromatography and for the potential presence of 25 known LAB bacteriocin genes directly in cheese and their microbial consortia by PCR. All batches were safe according to the European Union regulatory criteria for Listeria monocytogenes, Salmonella, enterobacteria, and coagulase-positive staphylococci.

Changes in the microbial composition of raw milk induced by thermization treatments applied prior to traditional Greek hard cheese processing.

The microbiological quality, safety, and composition of mixtures of ewe's and goat's milk (90:10) used for cheesemaking were evaluated before and after thermization at 60 and 67 degrees C for 30 s. Such mild thermal treatments are commonly applied to reduce natural contaminants of raw milk before processing for traditional hard Greek cheeses. Raw milk samples had an average total bacterial count of 7.

Fate of Listeria monocytogenes on fully ripened Greek Graviera cheese stored at 4, 12, or 25 degrees C in air or vacuum packages: in situ PCR detection of a cocktail of bacteriocins potentially contributing to pathogen inhibition.

The behavior of Listeria monocytogenes on fully ripened Greek Graviera cheese was evaluated. Three batches (A, B, and C) were tested. Batches A and C were prepared with a commercial starter culture, while in batch B the starter culture was combined with an enterocin-producing Enterococcus faecium Graviera isolate.

Leuconostoc carnosum associated with spoilage of refrigerated whole cooked hams in Greece.

A polyphasic taxonomic approach was used to identify a major atypical group of gas-forming, arginine-negative lactic acid bacteria associated with spoilage of whole (nonsliced) refrigerated (4 degrees C) cooked hams produced in two Greek industrial meat plants. Biochemical characterization revealed that the ham isolates shared their phenotypic properties with Leuconostoc carnosum, Weissella viridescens, and Weissella hellenica. However, gas chromatographic analysis of cellular fatty acids clearly differentiated the ham isolates from the Weissella spp.

Survival of Escherichia coli O157:H7 in Galotyri cheese stored at 4 and 12 degrees C.

Post-process contamination of fresh acid-curd cheeses with Escherichia coli O157:H7 may pose a risk considering the low infectious dose and the ability of the pathogen to survive in acidic foods. To evaluate its survival in Galotyri, a traditional Greek acid-curd cheese, portions (0.5 kg) of two commercial fresh products, one artisan (pH 3.