Evaluation of stored lamb bio-preserved using a three-strain cocktail of Lactobacillus sakei

Commercially prepared lamb was stored at −1.5 °C after inoculation with a combination of three Lactobacillus sakei strains previously shown to inhibit spoilage and pathogenic bacteria of importance to the meat industry. Between 6 and 14 weeks storage samples were evaluated for growth of inoculated strains, production of fermentation end-products and sensory acceptance of the cooked product. All three L. sakei strains flourished during storage, formed consistently dominant populations and were associated with lower surface pH and increased levels of lactic and acetic acids. Inoculated samples were determined to be as equally acceptable for smell, acidity, rancidity and overall liking as un-inoculated controls.     

The survival of Campylobacter jejuni in red meats stored at different temperatures

The survival of 4 strains of Campylobacter jejuni was studied in raw minced beef and raw pork sausage mixture stored in plastic stomacher bags at freezer temperatures (−19°C) for up to 10 weeks, refrigirator temperatures (< 10°C) for 6 days and 22°C for 24 h. At each of the 3 storage temperatures survival was better in minced beef. Similarly, there was less variation in percentage survival between the 4 strains in minced beef than in sausage mixture after storage at each temperature. Detailed studies were carried out with one strain of C. jejuni. Viable counts were relatively unchanged in minced beef at refrigerator temperatures and 22°C, but showed a decrease in corresponding samples of sausage mixture. At freezer temperatures decreases in count of approximately 1 log unit were observed during the first week for both meats followed by a more gradual decrease. The effect of desiccation by exposure was studied in minced beef and lamb outer carcass meat (breast) at refrigerator temperatures (≤ 10°C). Decreases in viable count were observed in lamb carcass meat after 32 h although large variations were sometimes observed between duplicate samples for the same strain of C. jejuni. Counts were unchanged in exposed minced beef after storage for 48 h.     

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.     

Comparative analysis of acid resistance in Listeria monocytogenes and Salmonella enterica strains before and after exposure to poultry decontaminants. Role of the glutamate decarboxylase (GAD) system

Data on the ability of chemical poultry decontaminants to induce an acid stress response in pathogenic bacteria are lacking. This study was undertaken in order to compare the survival rates in acid broths of Listeria monocytogenes and Salmonella enterica strains, both exposed to and not exposed to decontaminants. The contribution of the glutamate decarboxylase (GAD) acid resistance system to the survival of bacteria in acid media was also examined. Four strains (L. monocytogenes serovar 1/2, L. monocytogenes serovar 4b, S. enterica serotype Typhymurium and S. enterica serotype Enteritidis) were tested before (control) and after exposure to trisodium phosphate, acidified sodium chlorite, citric acid, chlorine dioxide and peroxyacids (strains were repeatedly passed through media containing increasing concentrations of a compound). Stationary-phase cells (10⁸ cfu/ml) were inoculated into tryptic soy broth (TSB) acidified with citric acid (pH 2.7 and 5.0) with or without glutamate (10 mM) added, and incubated at 37 °C for 15 min. Survival percentages (calculated from viable colonies) varied from 2.47 ± 0.67% to 91.93 ± 5.83%. L. monocytogenes cells previously exposed to acid decontaminants (citric acid and peroxyacids) showed, when placed in acid TSB, a higher (P < 0.05) percentage of survival (average 38.80 ± 30.52%) than control and pre-exposed to non-acidic decontaminants strains (22.82 ± 23.80%). Similar (P > 0.05) survival percentages were observed in previously exposed to different decontaminants and control Salmonella strains. The GAD acid resistance system did not apparently play any role in the survival of L. monocytogenes or S. enterica at a low pH. This study demonstrates for the first time that prior exposure to acidic poultry decontaminants increases the percentage of survival of L. monocytogenes exposed to severe acid stress. These results have important implications for the meat industry when considering which decontaminant treatment to adopt.     

Effect of X-ray treatments on inoculated Escherichia coli O157: H7, Salmonella enterica, Shigella flexneri and Vibrio parahaemolyticus in ready-to-eat shrimp

This study was conducted to evaluate the inactivation effect of X-ray treatments on Escherichia coli O157: H7, Salmonella enteric (S. enterica), Shigella flexneri (S. flexneri) and Vibrio parahaemolyticus (V. parahaemolyticus) artificially inoculated in ready-to-eat (RTE) shrimp. A mixed culture of three strains of each tested pathogen was used to inoculate RTE shrimp. The shrimp samples were inoculated individually with selected pathogenic bacteria then aseptically placed in sterile plastic cups and air-dried at 22 °C for 30 min (to allow bacterial attachment) in the biosafety cabinet prior to X-ray treatments. The inoculated shrimp samples were then placed in sterilized bags and treated with 0.1, 0.2, 0.3, 0.5, 0.75, 1.0, 2.0, 3.0 and 4.0 kGy X-ray at ambient temperature (22 °C and 60% relative humidity). Surviving bacterial populations were evaluated using a non-selective medium (TSA) with the appropriate selective medium overlay for each bacterium; CT-SMAC agar for E. coli O157: H7, XLD for S. enterica and S. flexneri and TCBS for V. parahaemolyticus. More than a 6 log CFU reduction of E. coli O157: H7, S. enterica, S. flexneri and V. parahaemolyticus was achieved with 2.0, 4.0, 3.0 and 3.0 kGy X-ray, respectively. Furthermore, treatment with 0.75 kGy X-ray significantly reduced the initial microflora on RTE shrimp samples from 3.8 ± 0.2 log CFU g⁻¹ to less than detectable limit (<1.0 log CFU g⁻¹).     

Bioconversion of anthocyanin glycosides by Bifidobacteria and Lactobacillus

Eight strains of Lactobacillus plantarum, 6 strains of Lactobacillus casei, Lactobacillus acidophilus LA-5, and Bifidobacterium lactis BB-12 were screened for β-glucosidase activity. We then proceeded to investigate the enzymatic potential of selected strains for bioconversion of delphinidin and malvidin glycosides to their metabolites. L. plantarum and L. casei strains showed the highest cell-envelope associated β-glucosidase activity. Intracellular β-glucosidase activity from B. lactis BB-12 was up to 287-fold higher than that of the other strains. The L. acidophilus strain showed low β-glucosidase activity, both, intra and extracellularly. No aglycons were detected in bacterial extract reactions with anthocyanin glycosides. Delphinidin-3-glucoside underwent chemical degradation to form mainly gallic acid, although delphinidin-3-glucoside degradation due to B. lactis BB-12 and enzymatic activity towards chemically-formed metabolites due to L. casei LC-01 were observed. Incubation of malvidin-3-glucoside with B. lactis BB-12, L. plantarum IFPL722, and L. casei LC-01 cell-free extracts led to different patterns of gallic, homogentisic and syringic acid formation.     

Modelling the growth of Listeria monocytogenes in fresh green coconut (Cocos nucifera L.) water

The behaviour of Listeria monocytogenes in the fresh coconut water stored at 4 °C, 10 °C and 35 °C was studied. The coconut water was aseptically extracted from green coconuts (Cocos nucifera L.) and samples were inoculated in triplicate with a mixture of 5 strains of L. monocytogenes with a mean population of approximately 3 log₁₀ CFU/mL. The kinetic parameters of the bacteria were estimated from the Baranyi model, and compared with predictions of the Pathogen Modelling Program so as to predict its behaviour in the beverage. The results demonstrated that fresh green coconut water was a beverage propitious for the survival and growth of L. monocytogenes and that refrigeration at 10 °C or 4 °C retarded, but did not inhibit, growth of this bacterium. Temperature abuse at 35 °C considerably reduced the lagtimes. The study shows that L. monocytogenes growth in fresh green coconut water is controlled for several days by storage at low temperature, mainly at 4 °C. Thus, for risk population this product should only be drunk directly from the coconut or despite the sensorial alterations should be consumed pasteurized.     

Control of food spoiling bacteria in cooked meat products with nisin,lacticin 3147, and a lacticin 3147-producing starter culture

Nisin, in the form of the commercial product Nisaplin, and lacticin 3147 in whey powdered form were added to minced pork-meat in amounts of 0.15% (w/w) and 1.5% (w/w), respectively. The meat was cooked and inoculated with a Staphylococcus aureus strain of meat origin and a Listeria innocua strain at a level of 10⁷ or 10⁵ CFU g^–1. The batches were stored vacuum-packaged for 21 days at 8 °C. Nisin and lacticin 3147 immediately reduced the L. innocua population at the time of inoculation. Nisin showed higher inhibitory activity than lacticin 3147. During the storage period, a slight L. innocua growth was observed in the batches inoculated with the larger inoculum, and a bacteriostatic effect was observed against Listeria in the batches inoculated with 10⁵ CFU g^–1. Nisin maintained a constant S. aureus population in the cooked batch inoculated with 10⁷ CFU g^–1, although the bacteriocin was capable of reducing the amount of S. aureus by 90% in the batch inoculated with 10⁵ CFU g^–1. On the other hand, lacticin 3147 did not show an inhibitory effect against S. aureus in the cooked meat. The starter culture Lactococcus lactis DPC 303-T4 (containing the conjugative plasmid encoding production of lacticin 3147) was inoculated in a portion of a Longissimus dorsi pork muscle with brine. L. lactis DPC 303-T4 performed a good fermentation, but lacticin 3147 production was not found after 7 days at 12 °C of storage.     

Indigenous microorganisms from iceberg lettuce with adherence and antagonistic potential for use as protective culture

The antagonistic effect of the indigenous microflora on pathogens was investigated by using Salmonella Typhimurium, Staphylococcus aureus and Listeria innocua as challenge microorganisms. 18 strains strongly adherent to the leaves of iceberg lettuce were selected and their taxonomic status was determined with the aid of API NE20, BBL CRYSTAL™ E/NF systems and 16 S rDNA sequencing. In the majority of cases (15/19), the identification by biochemical testing did not agree with that of 16 S rDNA sequencing. Six strains exerted antagonistic effects on S. Typhimurium and L. innocua, when overnight cultures or cell free culture supernatants were investigated in the agar diffusion test. In vivo experiments showed that the inoculation of lettuce with P. putida LTH 5878 by dipping and pre-incubation decreased the numbers of S. Typhimurium, L. innocua and S. aureus below the level of detection (< 100 cfu/g) after storage for 7–8 d at 4 °C. The dependency of the antagonistic efficacy on the numbers of P. putida LTH 5878 was determined using point inoculation. The study of the effect of the pseudomonades/pathogen ratio on the reduction of pathogens showed that the antagonistic activity of Pseudomonas is stronger against S. Typhimurium than against L. innocua. A major reduction was achieved for the counts of L. innocua at ratios of greater than 100:1 whereas for S. Typhimurium a ratio of 0.1:1 was already effective. The sensory properties of the inoculated and stored lettuce were in general better than those of the untreated control.

Industrial relevance

The increasing importance of minimally processed ready-to-eat (RTE) vegetables has initiated many studies using surface treatments for microbial decontamination. However, most treatments such as the use of chlorine or ozone have not proven highly effective or desirable. This study is of high relevance because it evaluates the potential of endogenous microbial populations with high surface adherence properties as a competitive tool against pathogenic microorganisms. This is an intriguing concept which deserves further development because of its potential as an new biological food preservation process.     

The fate of Listeria monocytogenes during the manufacture of Camembert-type cheese: A comparison between raw milk and milk treated with high hydrostatic pressure

Camembert-type cheese was produced from: raw bovine milk; raw milk inoculated with 2 or 4 log CFU/ml Listeria monocytogenes; raw milk inoculated with L. monocytogenes and subsequently pressure-treated at 500 MPa for 10 min at 20 °C; or uninoculated raw milk pressure-treated under these conditions. Cheeses produced from both pressure-treated milk and untreated milk had the typical composition, appearance and aroma of Camembert. Curd and cheese made from inoculated, untreated milk contained large numbers of L. monocytogenes throughout production. An initial inoculum of 1.95 log CFU/ml in milk increased to 4.52 log CFU/g in the curd and remained at a high level during ripening, with 3.85 log CFU/g in the final cheese. Pressure treatment inactivated L. monocytogenes in the raw milk at both inoculum levels and the pathogen was not detected in any of the final cheeses produced from pressure-treated milk. Therefore high pressure may be useful to inactivate L. monocytogenes in raw milk that is to be used for the production of soft, mould-ripened cheese.

Industrial relevance

This paper demonstrates the potential of high pressure (HP) for treatment of raw milk to be used in the manufacture of soft cheeses. HP treatment significantly reduced the level of Listeria monocytogenes in the raw milk and so allowed the production of safer non-thermally processed camembert-like soft cheese.     

Angiotensin-converting enzyme inhibitory activity of milk fermented by wild and industrial Lactococcus lactis strains

Angiotensin I-converting enzyme inhibitory (ACEI) activity was evaluated and compared in <3 KDa water-soluble extracts (WSE) isolated from milk fermented by wild and commercial starter culture Lactococcus lactis strains after 48 h of incubation. The highest ACEI activities were found in WSE from milk inoculated with wild L. lactis strains isolated from artisanal dairy products and commercial starter cultures. On the other hand, the lowest ACEI activities were found in WSE from milk inoculated with wild strains isolated from vegetables. Moreover, the IC₅₀ values (concentration that inhibits 50% activity) of WSE from artisanal dairy products were the lowest, indicating that these fractions were the most effective in inhibiting 50% of ACE activity. In fact, a strain isolated from artisanal cheese presented the lowest IC₅₀ (13 μg/mL). Thus, it appears that wild L. lactis strains isolated from artisanal dairy products and commercial starter cultures showed good potential for the production of fermented dairy products with ACEI properties.     

Tunisian Salvia officinalis L. and Schinus molle L. essential oils: their chemical compositions and their preservative effects against Salmonella inoculated in minced beef meat

The essential oils (EOs) extracted from the aerial parts of cultivated Salvia officinalis L. and the berries of Schinus molle L. were analysed by gas chromatography–mass spectrometry (GC–MS) and 68 and 67 constituents were identified, respectively. The major constituents were 1,8-cineole (33.27%), β-thujone (18.40%), α-thujone (13.45%), borneol (7.39%) in S. officinalis oil and α-phellandrene (35.86%), β-phellandrene (29.3%), β-pinene (15.68%), p-cymene (5.43%) and α-pinene (5.22%) in S. molle oil.In its second part, the present study was conducted to evaluate the in vitro antimicrobial activity of both studied EOs. For this purpose, paper disc-diffusion method and broth microdilution test were used. The disc-diffusion method showed significant zone of lysis against all the pathogens studied (gram-negative and gram-positive bacteria, yeast). These activities remained stable after six months, and decreased approximately by 20% after one year of storage of the EOs at 4 to 7 °C. On comparing the efficiency of both EOs, S. officinalis EO exhibited higher antibacterial activity against the majority of strains and especially against Candida albicans (two fold more active according to the inhibition zones values). The minimal inhibitory concentrations (MICs) were reported between 4.5 mg/ml and 72 mg/ml on nutrient broth. The particular chemotype of each EO may be involved in its specific antimicrobial behaviour.Furthermore, the inhibitory effect of these EOs were evaluated against two foodborne pathogens belonging to Salmonella genus, experimentally inoculated (10³ CFU/g) in minced beef meat, which was mixed with different concentrations of the EO and stored at 4 to 7 °C for 15 days. Although the antibacterial activities of both EOs in minced beef meat were clearly evident, their addition had notable effects on the flavour and taste of the meat at concentrations more than 2% for S. molle and 1.5% for S. officinalis. One solution to the above-mentioned problem may be the use of combinations of different food preservation systems. In this context, each of the EOs has been used along with low water activity (addition of NaCl) in addition to low refrigeration temperatures. Results on the Salmonella growth showed that some combinations could be recommended to eliminate germs from minced raw beef. By using this method, a stable and, from a microbiological point of view, safe meat can be produced without substantial loss in sensory quality.Results obtained herein, may suggest that the EOs of S. officinalis and S. molle possess antimicrobial activity, and therefore, they can be used in biotechnological fields as natural preservative ingredients in food and/or pharmaceutical industry.     

Behavior of Escherichia coli O157:H7, Listeria monocytogenes, and Salmonella Typhimurium in teewurst, a raw spreadable sausage

The fate of Listeria monocytogenes, Salmonella Typhimurium, or Escherichia coli O157:H7 were separately monitored both in and on teewurst, a traditional raw and spreadable sausage of Germanic origin. Multi-strain cocktails of each pathogen (ca. 5.0 log CFU/g) were used to separately inoculate teewurst that was subsequently stored at 1.5, 4, 10, and 21 °C. When inoculated into commercially-prepared batter just prior to stuffing, in general, the higher the storage temperature, the greater the lethality. Depending on the storage temperature, pathogen levels in the batter decreased by 2.3 to 3.4, ca. 3.8, and 2.2 to 3.6 log CFU/g for E. coli O157:H7, S. Typhimurium, and L. monocytogenes, respectively, during storage for 30 days. When inoculated onto both the top and bottom faces of sliced commercially-prepared finished product, the results for all four temperatures showed a decrease of 0.9 to 1.4, 1.4 to 1.8, and 2.2 to 3.0 log CFU/g for E. coli O157:H7, S. Typhimurium, and L. monocytogenes, respectively, over the course of 21 days. With the possible exceptions for salt and carbohydrate levels, chemical analyses of teewurst purchased from five commercial manufacturers revealed only subtle differences in proximate composition for this product type. Our data establish that teewurst does not provide a favourable environment for the survival of E. coli O157:H7, S. Typhimurium, or L. monocytogenes inoculated either into or onto the product.     

Reduction of spoilage of chilled vacuum-packed lamb by psychrotolerant clostridia

Methods for the reduction of spoilage, of lamb, by psychrotolerant clostridia were investigated including exposure to air, hot and cold water spray washing and tyndallisation. Initially vegetative cells of psychrotolerant clostridia associated with spoilage of chilled vacuum-packed meat were exposed to aerobic cooked meat medium at room temperature (21 °C) to determine how long they remained viable. Survival of strains varied from 2 h to 3 days. Vegetative cells of Clostridium estertheticum subsp. estertheticum survived 7 days at 10 °C with little reduction in viable numbers. This ruled out exposure to air as a practical method for reducing spoilage. Trials were also carried out on chilled vacuum-packed lamb inoculated with spores of Cl. estertheticum subsp. estertheticum. The time until inoculated packs reached the loss of vacuum stage varied from 38 to 53 days. Hot and cold water washing extended the shelf life by 12 to 13 days in comparison to untreated packs.     

Evaluation of the effect of defrosting practices of ground beef on the heat tolerance of Listeria monocytogenes and Salmonella Enteritidis

The effect of common defrosting practices of ground beef, including (i) defrosting in the refrigerator (5 °C for 15 h), (ii) defrosting at room temperature (25 °C for 12 h) and (iii) defrosting in the microwave, on the heat tolerance of artificially inoculated Listeria monocytogenes and Salmonella Enteritidis, was studied. The thermal inactivation of S. Enteritidis was not, overall, affected by defrosting practices. In contrast, defrosting at room temperature resulted, overall, in an increased heat tolerance of L. monocytogenes compared to the rest tested defrosting practices. Inactivation kinetics of the two pathogens for the different defrosting practices were determined by fitting the data to the Weibull model. The δ parameter of the Weibull model (heat challenge time (min) required for the first 1-log reduction) for S. Enteritidis and for defrosting at 25 °C, microwave defrosting, defrosting at 5 °C and for the control (fresh ground beef inoculated with the pathogens just before the heat challenge trials) was 1.13, 1.62, 1.60 and 0.96, respectively, while the corresponding values for L. monocytogenes were 20.13, 10.82, 9.95 and 9.47, respectively. The findings of this study should be useful in risk assessments and in developing food handling guidelines for the consumers.     

Environmental factors modify carbon nutritional patterns and niche overlap between Aspergillus flavus and Fusarium verticillioides strains from maize

This study examined the utilization patterns of key carbon sources (CS, 24: including key sugars, amino acids and fatty acids) in maize by strains of Aspergillus flavus and Fusarium verticillioides under different water activity (a_w, 0.87–0.98 a_w) and temperature (20–35 °C) values and compared the niche overlap indices (NOI) that estimate the in vitro CS utilization profiles [Wilson, M., Lindow, S.E., 1994. Coexistence among epiphytic bacterial populations mediated through nutritional resource partitioning. Applied and Environmental Microbiology 60, 4468–4477.]. The ability to grow in these key CS in minimal media was studied for 120 h in 12 h steps. The NOI was calculated for inter-species (F. verticillioides–A. flavus) and for intra-species (A. flavus–A. flavus) using CS utilization patterns over the range of interacting environmental conditions. 30 °C, over the whole a_w range examined, was found to be optimal for utilization of the maximum number of CS by A. flavus. In contrast, for F. verticillioides this was more so at 20 °C; 25 °C allowed a suboptimal usage of CS for both species. NOIs confirmed the nutritional dominance of A. flavus at 30 °C, especially at lower a_w levels and that of F. verticillioides at 20 °C, mainly at 0.95 a_w. In other conditions of a_w, based on CS utilization patterns, the data indicated that A. flavus and F. verticillioides occupied different ecological niches. The variability in nutritional sources utilization between A. flavus strains was not related to their ability to produce aflatoxins (AFs). This type of data helps to explain the nutritional dominance of fungal species and strains under different environmental conditions. This could be useful in trying to find appropriate natural biocontrol microorganisms to compete with these mycotoxigenic species.     

Role of quantity and quality of fat in meat models inoculated with Listeria innocua or Salmonella Typhimurium treated by high pressure and refrigerated stored

Several variables can influence the effects of high hydrostatic pressure processing (HPP), but the role of fat in the treated sample is still uncertain. We designed a model by which controlling the known variables we could elucidate that role. We applied 400 MPa for 2 min to minced chicken samples inoculated with Listeria innocua and Salmonella Typhimurium mixed with 10% and 20% of three fat types with different fatty acid composition. Microbial counts were performed during 60 days of refrigerated storage either at 2 °C or 8 °C.Immediately after HPP bacterial growth was independent of the type and percentage of fat content, but a possible effect of type of fat could be observed after 60 days of cold storage.     

Evaluation of stored lamb bio-preserved using a three-strain cocktail of Lactobacillus sakei

Commercially prepared lamb was stored at −1.5 °C after inoculation with a combination of three Lactobacillus sakei strains previously shown to inhibit spoilage and pathogenic bacteria of importance to the meat industry. Between 6 and 14 weeks storage samples were evaluated for growth of inoculated strains, production of fermentation end-products and sensory acceptance of the cooked product. All three L. sakei strains flourished during storage, formed consistently dominant populations and were associated with lower surface pH and increased levels of lactic and acetic acids. Inoculated samples were determined to be as equally acceptable for smell, acidity, rancidity and overall liking as un-inoculated controls.     

Effect of modified atmosphere and temperature abuse on the growth from spores and cereulide production of Bacillus weihenstephanensis in a cooked chilled meat sausage

The effect of modified atmosphere packaging (MAP) on the germination and growth of toxin producing psychrotolerant Bacillus spp is not well described. A model agar system mimicking a cooked meat product was used in initial experiments. Incubation at refrigeration temperature of 8 °C for 5 weeks of 26 Bacillus weihenstephanensis including two emetic toxin (cereulide) producing strains showed that B. weihenstephanensis is sensitive to MAP containing CO₂. The sensitivity to 20% CO₂ was dependent on strain and oxygen level, being increased when oxygen was excluded from the MAP. Growth from spores was observed at the earliest within 2 weeks when 20% CO₂ was combined with 2% O₂ and in 3 weeks when combined with “0”% O₂ (the remaining atmosphere was made up from N₂). Results were validated in a cooked meat sausage model for two non-emetic and one emetic B. weihenstephanensis strain. The packaging film oxygen transfer rates (OTR) were 1.3 and 40 ml/m²/24 h and the atmospheres were 2% O₂/20% CO₂ and “0”% O₂/20% CO₂. Oxygen availability had a large impact on the growth from spores in the MAP meat sausage, only the most oxygen restricted condition (OTR of 1.3 ml/m²/24 h and “0”% O₂/20 % CO₂) inhibited growth of the three strains during 4 weeks storage at 8 °C. Cereulide production was undetectable during storage at 8 °C irrespective of choice of the MAP (quantified by liquid chromatography mass spectrometry/mass spectrometry). MAP storage at 8 °C for 1 and 3 weeks followed by opening of packages and temperature abuse for 1.5 h daily at 20 °C during 1 week resulted in increased cell counts and variable cereulide production in the meat sausage. A pre-history at 8 °C for 1 week in MAP with OTR of 1.3 or 40 ml/m²/24 h and 2% O₂ resulted in cereulide concentrations of 0.816 – 1.353 µg/g meat sausage, while a pre-history under the most oxygen restricted condition (OTR of 1.3 ml/m²/24 h, “0”% O₂/20 % CO₂) resulted in minimal cereulide production (0.004 µg/g meat sausage) at abuse condition. Extension of MAP storage at 8 °C for 3 weeks followed by abuse resulted in a substantially reduced cereulide production.Data demonstrates that MAP can be used to inhibit growth of a psychrotolerant toxin producing Bacillus spp. during chill storage at 8 °C, and substantially reduce the risk of emetic food poisoning at abuse condition. Results are of relevance for improving safety of ready to eat processed chilled foods of extended durability.     

Selection and evaluation of seafood-borne psychrotrophic lactic acid bacteria as inhibitors of pathogenic and spoilage bacteria

In this study, inhibitory psychrotrophic lactic acid bacteria were isolated and investigated for future use in biopreservation of seafood products. Screening of 5575 colonies isolated from various seafood products resulted in the selection of 132 colonies presenting inhibitory properties. Among them, 52 isolates had characteristics of LAB and showed growth at 15 °C but not at 30 °C. The inhibition spectrum of these 52 isolates against 14 target strains (Gram-positive and -negative) showed inhibition of typical seafood spoiling and pathogenic bacteria and enabled the formation of seven interesting clusters. Sequencing of the 16S rRNA gene of a representative isolate from each cluster identified three Leuconostoc gelidum, two Lactococcus piscium, one Lactobacillus fuchuensis and one Carnobacterium alterfunditum. Theses strains did not produce histamine nor tyramine, and showed no particular antibiotic resistance profile. Growth rate as a function of temperature was tested for one L. piscium and one L. gelidum isolate and confirmed their psychrotrophic behavior. One out of seven isolates showed bacteriocin-like activity. The inhibition mechanisms of the other isolates are still unknown but may be due to competition for substrate. Absence of a bacteriocin-like component could be a positive point to gain rapid authorization for food application in France. This collection of LAB is now ready for testing on products.