Effect of Lactobacillus cultures on microbiological,chemical and odour changes during storage of rainbow trout fillets

The purpose of this study was to investigate the effect of applying two Lactobacillus protective cultures on microbiological, chemical and odour changes during storage of refrigerated vacuum‐packed rainbow trout (Oncorrynchus mykiss) fillets. Lactobacillus sakei CECT 4808 and Lb. curvatus CECT 904^T were inoculated individually or in combination into rainbow trout fillets. The samples were vacuum‐packed and stored at 4 ± 0.5 °C and were assessed during a 20‐day storage period for microbiological (Enterobacteriaceae, Pseudomonas spp., lactic acid bacteria (LAB), H₂S‐producing bacteria, yeasts and moulds) and chemical (pH, total volatile basic nitrogen, lipid oxidation) parameters and off‐odour. Samples inoculated with Lb. sakei and with both strains had significantly (P≤0.05) lower counts of all microbiological spoilage indicator organisms (Enterobacteriaceae, Pseudomonas spp., H₂S‐producing bacteria, yeasts and moulds) than those inoculated with Lb. curvatus or the controls. All chemical parameters examined and off‐odour scores were also significantly (P≤0.05) improved in the samples inoculated with Lb. sakei or with both strains compared with those inoculated with Lb. curvatus or the controls. Lb. sakei generally showed a better preservative effect (P≤0.05) than the combination of strains. Inoculation with Lb. sakei CECT 4808 could provide an additional hurdle to improve shelf‐life of refrigerated vacuum‐packaged trout fillets, as inoculation with this strain resulted in extension of shelf‐life by 5 days. Furthermore, this strain showed some antioxidative ability, which could retard lipid oxidation in fish. Copyright © 2006 Society of Chemical Industry     

Study of the Lactobacillus sakei protective effect towards spoilage bacteria in vacuum packed cooked ham analyzed by PCR-DGGE

The effectiveness of Lactobacillus sakei B-2 inoculated as a protective culture on the inhibition of spoilage bacteria on sliced vacuum packed cooked ham was investigated by using culture-dependent and -independent approaches. Total microbial DNA was directly extracted from both control and treatment samples, and subjected to a nested PCR protocol, PCR–DGGE analysis was used to identify and monitor the dynamic changes in the microbial population, followed by partial 16S rDNA sequencing. The DGGE profile demonstrated that the protective culture effectively suppressed growth of predominant spoilage bacteria L. sakei, Lactobacillus curvatus and Leuconostoc mesenteroides in cooked ham during storage at 4 °C, however, growth of uncultured Leuconostoc was not inhibited. The shelf-life of this product inoculated with L. sakei B-2, at levels of 5.91 ± 0.04 log₁₀ CFU g⁻¹ was 35 days, compared to 15 days of control samples, when the ham was stored at 4 °C.     

Inhibition by Lactobacillus sakei of other species in the flora of vacuum packaged raw meats during prolonged storage

The abilities of five Lactobacillus sakei strains and one Lactococcus lactis strain to retain inhibitory activity against several target organisms in the flora of product during 12 weeks storage of vacuum-packaged lamb and beef was investigated. L. sakei strains were generally found capable of developing dominant populations on both beef and lamb. L. lactis 75 grew poorly on lamb did not inhibit co-inoculated Brochothrix thermosphacta. Lamb inoculated with the Sakacin-A producer L. sakei Lb706 had lower Listeria monocytogenes populations than lamb inoculated with a bacteriocin-negative variant. In beef packs inoculated with Clostridium estertheticum spores and L. sakei strain 27, 44 or 63, the development of blown-pack spoilage was delayed by up to one week. Campylobacter jejuni inoculated onto beef was recovered from fewer packs when it was co-inoculated with 3000 CFU cm⁻² of L. sakei strain 27, 44 or 63. Observed inhibition did not always correlate with inhibition observed in earlier media-based studies, supporting the view that functionality identified using simple media-based screening methods may not be replicated in the complex environment of stored foods, and vice-versa. These findings further define a set of L. sakei strains with potential for the extended bio-preservation of minimally processed fresh beef and lamb.     

Application of the bacteriocinogenicLactobacillus sakeiCTC494 to prevent growth of Listeria in fresh and cooked meat products packed with different atmospheres

The production of sakacin K byLactobacillus sakeiCTC494 at different temperatures and pH was evaluated before its application as a bioprotective culture againstListeriain different meat-based food packaging systems, that is, oxygen-permeable film, under vacuum and under a modified atmosphere (20% CO₂: 80% O₂) and stored at 7°C. TheLactobacillusculture produced the bacteriocin at a range of temperatures from 4°C to 30°C and at initial pH from 5·5 to 6·5.Listeriainhibition in raw minced pork, poultry breasts and modelized cooked pork could not be achieved by the sole application of vacuum or a modified atmosphere. Inoculation ofLb. sakeiCTC494 or sakacin K inhibited the growth ofListeriato different extents in all the products studied in each system; the greatest inhibition being observed in the vacuum packaged samples of poultry breasts and cooked pork, and in the modified atmosphere packaged samples of raw minced pork. Addition of sakacin K resulted in immediate bactericidal action againstListeriain every product and atmosphere studied.     

Molecular and technological characterization of lactic acid bacteria from traditional fermented sausages of Basilicata region (Southern Italy)

Lactic acid bacteria (LAB) from traditional fermented sausages of the Basilicata region were investigated by ARDRA-PCR and RAPD-PCR for taxonomic identification at species and strain level and characterized on the basis of the growth and acidification at different temperatures, incubation times, levels of NaCl and KNO₂, hydrolysis of sarcoplasmatic and myofibrillar proteins and antimicrobial, peptide/amino acid release and nitrate reductase activities.Lactobacillus sakei was the predominant species (67%) followed by Pediococcus pentosaceus (16%), Leuconostoc carnosum (8%), Lactobacillus plantarum (4%), Lactobacillus brevis (2%) and Leuconostoc pseudomesenteroides (2%). The technological characterization revealed that most of the isolates had good acidifying and proteolytic properties. Moreover, Lb. sakei strains showed antimicrobial ability, while Leuconostoc strains the highest reduction of nitrates.This work was a preliminary study in the formulation of autochthonous starter cultures in order to standardize the production process of sausages, to preserve their typical organoleptic and sensory characteristics and to improve the quality of final product.     

Lactic acid bacteria as protective cultures against Listeria spp. on cold-smoked salmon

Three bacteriocin producing (Bac⁺) strains of Lactobacillus sakei were used singly and in combination with each other as protective cultures to control the growth of listeria in cold-smoked salmon. Challenge experiments were conducted under practical conditions in a smokehouse. The surface of salmon sides was inoculated with 10⁴ cfu/g of Listeria innocua and 10⁷ cfu/g of Bac⁺ lactic acid bacteria as well as a L. sakei Bac⁻ control. After smoking the counts of listeria and lactic acid bacteria were determined at days 1 and 14. All Bac⁺ L. sakei strains reduced the counts of L. innocua by >2 log units. Strain LTH5754 was an isolate from cold-smoked salmon and achieved even a 5 log reduction of L. innocua within the storage period. In vitro experiments showed, that the Bac⁺ strains were also effective against L. monocytogenes (three strains tested) and L. ivanovii (1 strain). The pH as well the sensorial properties of the smoked salmon were not affected by the L. sakei inocula.     

Analysis of volatile compounds produced by different species of lactobacilli in rye sourdough using multiple headspace extraction

Profiles of volatile organic compound (VOC) produced by nine individual lactic acid bacteria (LAB) during rye sourdough fermentation were compared by automated SPME and GC/MS‐Tof. The dough samples were inoculated with individual strains, placed inside the headspace vials and incubated during next 24 h. The production or loss of VOC‐s was followed by adsorbing volatiles onto 85‐m Car/PDMS fibre in every 4 h. Volatile profiles differed among LAB species and divided LAB into two main groups – hetero‐ and homofermentative. Hetrofermentative LAB (Lactobacillus brevis; Leuconostoc citreum; Lactobacillus vaginalis, Lactobacillus panis) showed high production of acetic acid, CO₂, ethanol, ethylacetate, producing also hexyl acetate, ethyl hexanoate and isopentyl acetate. Whereas homofermentative LAB species (Lactobacillus helveticus; Lactobacillus casei; Lactobacillus sakei; Lactobacillus curvatus) produced a considerable amount of 2,3‐butanedione. Production of l ‐leucine methyl ester was unique for Lb. sakei, Lb. casei and Lb. curvatus strains. Lb. helveticus was the only LAB that produced benzaldehyde.     

The inhibition of the viability of myeloma cells and the production of cytokine by two strains of Lactobacillus sakei from meat

Ten strains of lactic acid bacteria were isolated from a dry fermented sausage and tested for stimulation or inhibition of the viability of Vero and myeloma cells. They did not significantly affect the viability of Vero cells but two isolates (CBL/H and CBL/K) showed a strong and moderate inhibition of the myeloma cell viability (at 10⁸ CFU mL^?1, 17.6 and 33.2%, respectively, survival of myeloma cells). The isolates were identified as Lactobacillus sakei by DNA sequencing of the 16S rRNA products of a polymerase chain reaction. No protective effect was found, at the concentrations used, against cytotoxicity of N‐nitrosamines. To test the effect of L. sakei CBL/H and CBL/K on cytokine production [tumour necrosis factor alpha (TNF‐α), interleukin‐1β (IL‐1β) and interleukin‐8 (IL‐8)], the human macrophage cell line (THP‐1) was cultured in the presence and absence of lipopolysaccharide (LPS). Lactobacillus sakei CBL/H and CBL/K induced IL‐1β and IL‐8 release when cells were stimulated with and without LPS. However there was TNF‐α release only in the presence of the LPS.     

Assessment of technological attributes of autochthonous starter cultures in Turkish dry fermented sausage (sucuk)

The study aims to evaluate the technological properties of autochthonous strains (Lactobacillus sakei S15, Lactobacillus plantarum S24, L. plantarum S91, Pediococcus pentosaceus S128b and Staphylococcus carnosus G109) in Turkish dry fermented sausage (sucuk). After 24 h of fermentaPtion, all lactic acid bacteria strains reduced the pH to below 5.0, while the pH in the control group was above 5.3. The number of lactic acid bacteria strains reached 10⁸–10⁹ cfu g⁻¹ during fermentation. Staphylococcus carnosus G109 remained at the inoculation level of 10⁶ cfu g⁻¹ during ripening. Lactobacillus sakei S15 as mono-culture showed higher TBARS values compared to other strains. The control group had the lowest L* value and autochthonous strains caused no significant difference for a* value. According to principal component analysis results, most volatile compounds were positively correlated with the group containing only L. sakei S15.     

Microbial and physicochemical succession in fermented sausages produced with bacteriocinogenic culture of Lactobacillus sakei and semi-purified bacteriocin mesenterocin Y

The influence of the bacteriocinogenic culture Lactobacillus sakei (10⁵/g) and semi-purified bacteriocin mesenterocin Y (2560 AU/kg) on the safety and quality of traditional Croatian fermented sausages was investigated. The addition of Lb. sakei and/or mesenterocin Y reduced microbial counts (P < 0.05) in the final products. After 28 days of ripening, coagulase-negative cocci decreased 1.5–2.0 log, yeasts 1.2–1.4 log and enterococci 1.7–2.7 log. In the case of the addition of Lb. sakei, the lactic acid bacteria count was significantly (P < 0.05) higher at day 7 of ripening, and was accompanied by a lower pH and a higher amount of lactic acid (P < 0.05). In the final product the amount of acetic acid was significantly lower. More intensive proteolysis and an increase in ammonia content were found at the beginning of fermentation, and in the second phase of ripening in the control samples, respectively. The free fatty acid concentration was significantly lower during the entire ripening process compared to the control (P < 0.05). Semi-purified mesenterocin Y did not affect the sensory properties of the sausages, whilst the addition of Lb. sakei enhanced them.     

Effect of selected autochthonous starter cultures on processing and quality characteristics of Greek fermented sausages

Selected autochthonous starter (SAS) cultures (i.e. Lactobacillus sakei 8416, Lactobacillus sakei 4413, and L. sakei 8426, L. plantarum 7423 and L. curvatus 8427) were used as starter cultures in addition to a control treatment in the production of fermented sausages. The SAS cultures had a rapid growth and dominated the fortuitous population of LAB during the whole fermentation and ripening process improving the sensory attributes in comparison to control. Apart from the treatment produced with L. sakei 8416, all other SAS cultures prevented the lipid oxidation to values lower than 1 mg malonaldehyde/kg. The Micrococcaceae count and the redness of the sausages was not affected by the smoking and the acidification during the fermentation in the treatments produced with L. sakei 8416 and L. sakei 4413. The treatment of L. sakei 4413 had the lowest (*P < 0.05) content of all biogenic amines. In comparison to the control, the reduction of tyramine was 13%, tryptamine 55%, cadaverine 60% and putrescine 72%. Sausages produced with SAS cultures L. sakei 4413 and L. sakei 8416 had the highest scores for all sensory attributes. The results indicated that the SAS culture of L. sakei 4413 is the best autochthonous starter culture for fermented sausages.     

Sensory evaluation of black pigmented rice (Oryza sativa cv. Poireton) wine fortified with probiotic Lactobacillus acidophilus ATCC 4356 and Lactobacillus sakei ATCC 15521 using fuzzy logic

Rice wine was prepared in the laboratory from a black pigmented rice (cv. Poireton) using standardized methodology. The wine was fortified with the probiotic strains Lactobacillus acidophilus ATCC 4356 and Lactobacillus sakei ATCC 15521 and allowed to undergo a storage period of 28 days at 4°C. These rice wines were compared with a commercial wine used as a standard reference wine. A panel consisting of 30 judges performed the sensory evaluation. The scores obtained were based on the quality attributes of colour, taste, aroma and alcoholic strength. Analysis of the sensory scores and ranking of the different samples were performed using fuzzy analysis. The quality attributes were also ranked according to their importance in the overall acceptability. Colour and alcoholic strength were accorded the highest importance, compared with aroma and taste, as quality attributes. The rice wines were rated as ‘not satisfactory’, ‘fair’, ‘medium’, ‘good’ and ‘excellent’ on the sensory scale. Out of the wines prepared, black pigmented rice wine fortified with L. acidophilus was the most acceptable and was ranked as ‘good’, which was also at par with the commercial wine. On the other hand, black pigmented rice wine fortified with L. sakei and black pigmented rice wine fortified with L. acidophilus and L. sakei were ranked in the ‘satisfactory’ category followed by the unfortified wine ranked in the ‘fair’ category. Copyright © 2015 The Institute of Brewing & Distilling     

Characterization of Proteolytic Effect of Lactic Acid Bacteria Starter Cultures on Thai Fermented Sausages

The objective of this study was to investigate the effect of starter culture addition on proteolysis of Thai fermented sausages. Sausages inoculated with six different external starter cultures—Pediococcus pentosaceous, Pediococcus acidilactici, Weissella cibaria, Lactobacillus plantarum, Lactobacillus pentosus, and Lactobacillus sakei—were compared with naturally fermented sausages. The results of microbiological analysis indicated that the dominance of lactic acid bacteria (LAB) could inhibit the growth of pathogens and spoilage. Proteolysis was observed during fermentation by the reduction of myofibrillar and sarcoplasmic proteins and the increase in nonprotein nitrogen (NPN) and total free amino acids. The highest increase in concentration of NPN and free amino acids was obtained from sausages inoculated with LAB. Sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE) showed a similar pattern of proteolysis of sarcoplasmic proteins in all sausages, while that of the inoculated sausages with L. plantarum, L. pentsus, and L. sakei exhibited increased degradation of myofibrillar protein bands at 200 and 45 kDa.     

Development and PFGE monitoring of dominance among spoilage lactic acid bacteria from cured meats

Strains of lactic acid bacteria (LAB), isolated from intact vacuum-packaged cured meats obtained at retail, were subjected to single inhibitory factor and mixed-culture dominance tests during challenge with different conditions of pH and temperature in the presence of NaCl and NaNO₂to study reasons for strain dominance. Bacteria were checked for bacteriocin production. Both initial pH and temperature had a significant effect on the growth of spoilage LAB isolates in modified MRS broth. Lactobacilli generally grew better than Lc. mesenteroides at pH 5·5. It appeared that L. curvatus and L. sakei grew faster at elevated temperature (12°C) than Lc. mesenteroides. It was also noted that there was a similar response for strains within the same species to the challenges of NaCl, NaNO₂, pH and temperature. When present in mixed cultures, Leuconostoc strains did not grow well at 2°C and at an initial pH of 5·5 compared with lactobacilli, but performed better at this low temperature as pure cultures than when mixed with lactobacilli. At normal cured meat pH (6·0 and 6·5) and higher temperatures (6°C and 12°C), dominant bacteria always grew from the originally larger bacterial population. When leuconostocs and lactobacilli were present in equal initial numbers, leuconostocs generally did not grow as well as lactobacilli. Lc. mesenteroides grew faster than L. curvatus at 6°C but did not out-compete L. sakei at this temperature (pH ≥ 6·0). Pulsed field gel electrophoresis (PFGE) of SmaI digested genomic DNA successfully distinguished all the strains under study. Difficulties associated with enumeration of LAB having similar biochemical properties were addressed by the development of a composite-simultaneous PFGE method for interspecies qualification using single colonies from agar plates to generate digested DNA for analysis.     

Influence of the addition of lupin sourdough with different lactobacilli on dough properties and bread quality

The aim of this research was to study the effects of solid‐state fermentation (SSF) with Lactobacillus sakei, Pediococcus pentosaceus and P. acidilactici on lupine sourdough parameters and lupine sourdough influence on the physical dough properties and wheat bread quality. The results showed that lactic acid bacteria (LAB) significantly reduced the pH and increased total titratable acidity (TTA) of SSF lupine. The highest protease activity in lupine is excreted by L. sakei (187.1 ± 8.6 PU g^?1), and the highest amylase activity, by P. pentosaceus (155.8 ± 7.5 AU g^?1). Lupine sourdough has a significant effect on the rheological properties of doughs, which affect the baking characteristics of the final product. In conclusion, it can be said that L. sakei, P. pentosaceus and P. acidilactici could be used for lupine SSF, and the addition of up to the 10% SSF lupine products increases the wheat–lupine bread quality.     

Effect of the lactic acid bacteria on the control of listerial activity and shelf life of smoked salmon scraps

The potential protective activity of lactic acid bacteria (LAB) from cold smoked salmon scraps was evaluated towards Listeria monocytogenes. Seventy‐three LAB strains were isolated and identified by biomolecular methods; Lactobacillus curvatus and Lactobacillus sakei prevailed. Three of the strains tested, identified as L. sakei, profile O, had a significant inhibitory activity against L. monocytogenes ATCC 19115 and clones DUP‐1042 and DUP‐18596. The evolution of microbial populations and chemical parameters were determined at time intervals to verify the shelf life. Listeria monocytogenes was isolated in half the packages also exceeding the legal limit. The shelf life of scraps was set at 30 days. Clonal characterisation of L. monocytogenes was performed by ribotyping. DUP‐1042, one of the human pathogen clones, was the most represented pattern. The results suggest further studies aimed at the selection of autochthonous nonspoilage LAB strains as bioprotective agents for cold smoked salmon.     

Inhibition of Listeria monocytogenes and Escherichia coli O157:H7 in liquid broth medium and during processing of fermented sausage using autochthonous starter cultures

The antimicrobial effect of two autochthonous starter cultures of Lactobacillus sakei was evaluated in vitro (in liquid broth medium) and in situ assays. The inactivation of foodborne pathogens Listeria monocytogenes (serotype 4ab No 10) and Escherichia coli O157:H7 ATCC 43888 was investigated during the production of fermented sausage according to a typical Greek recipe using L. sakei strains as starter cultures. The inactivation kinetics were modeled using GInaFiT, a freeware tool to assess microbial survival curves. By the end of the ripening period, the inhibition of L. monocytogenes was significant in treatments with L. sakei 8416 and L. sakei 4413 compared to the control treatment. A 2.2-log reduction of the population of E. coli O157:H7 resulted from the autochthonous starter culture L. sakei 4413 during sausage processing. The use of the autochthonous starter cultures constitutes an additional improvement to the microbial safety by reducing foodborne pathogens.     

The influence of exopolysaccharide-producing lactic acid bacteria on reconstructed ham

The influence of in-situ-formed exopolysaccharides (EPS) by lactic acid bacteria (LAB; Lactobacillus sakeiTMW 1.411 and Lactobacillus plantarumTMW 1.1478) on the yield and texture of reconstructed ham was investigated. No differences in yield (P > 0.05) were observed but weight loss during storage was significantly higher (P < 0.05) for hams produced with L. sakei 1.411. Furthermore, the produced EPS decreased the hardness (P < 0.05; 50.22 N for control compared to 44.81 N) but did not influence the cohesiveness (P > 0.05) of hams. Products with L. plantarum 1.1478 showed no significant differences in comparison to the control. This could be attributed to the lower amount of EPS formed during ripening. L. sakei 1.411 produced 194.49 ± 5.34 mg kg⁻¹ EPS, whereas L. plantarum 1.1478 formed 60.26 ± 2.96 mg kg⁻¹ EPS. Thus, the use of the examined LAB is not recommended in reconstructed ham, since they have no or even a negative influence on the product quality.     

Identification of the lactic acid bacteria in Kimchi according to initial and over-ripened fermentation using PCR and 16S rRNA gene sequence analysis

This paper aimed to identify the lactic acid bacteria species involved in kimchi fermentation at different fermentation periods by using culture-independent 16S rRNA gene clone libraries, and develop polymerase chain reaction for the detection of lactic acid bacteria (LAB). We investigated 6 commercially produced kimchi samples, including kimchi at an initial stage of fermentation and kimchi that was fermented to an over-ripened stage. The results of our study show that the analysis with cultureindependent 16S rRNA gene clone libraries could successfully identify 134 clones, 11 species, including Weissella, Lactobacillus, Pediococcus, and Leuconostoc from the 6 commercial kimchi samples. Weisella koreensis and Lactobacillus brevis were the predominant LAB in the initial stage of kimchi fermentation at 4°C (pH 4.96–5.27 and acidity 0.81–0.88), and Leuconostoc gelidum and Lactobacillus sakei subsp. sakei may play an important role in kimchi fermentation at the over-ripened stage (pH 3.61–3.91 and acidity 1.70–1.79).     

The efficacy of nisin can drastically vary when produced in situ in model cheeses

Nisin, a bacteriocin produced by strains of Lactococcus lactis, has a broad inhibitory effect against Gram-positive bacteria. This study investigated the efficacy of nisin Z against Lactobacillus sakei when produced by a nisin-producing strain L. lactis in model cheeses manufactured with ultrafiltrated milk. These cheeses, containing 0, 4 or 10% of gelatin in their dry matter, were inoculated with both strains. Measurement of Lb. sakei loss of viability was an indirect indicator of nisin in situ efficacy. After 24 h, the loss of viability of Lb. sakei was from 0.73 ± 0.14 to 3.30 ± 0.60 log₁₀ cfu g⁻¹ in the cheeses with 0 and 10% of gelatin, respectively, indicating a better in situ efficacy of nisin when gelatin was incorporated. However, the concentration of nisin produced by Lactococcus was similar (3.5 μg g⁻¹) in all model cheeses when measured using an enzyme-linked immune sorbent assay (ELISA). The growth of Lactococcus was slightly improved when gelatin was incorporated, leading to a higher lactate concentration, which is one of the factors explaining the increased nisin efficacy. These results reinforced previous observations that prediction of nisin efficacy in complex food systems remains difficult.