Listeria Counts Research Articles

Survival of Listeria monocytogenes and Salmonella Enteritidis in sea bream (Sparus aurata) fillets packaged under enriched CO2 modified atmospheres

The effect of packaging of fresh sea bream fillets under different modified atmospheres (high levels of CO2 combined with N2) on the behaviour of Listeria monocytogenes and Salmonella Enteritidis was studied. Two different strains of each pathogen were spread over the sea bream fillets stored at either 0°C or 4°C for 16days under standard supermarket lighting conditions (14h a day). The atmospheres investigated were 60% CO2/40% N2, 70% CO2/30% N2, 80% CO2/20% N2 and air as control. Mesophilic, psychrotrophic and Enterobacteriaceae microbial counts were also performed in control batches. The results indicate that all the CO2 concentrations used favoured the control of pathogen bacteria inoculated, when compared with air conditions especially, when combined with storage at 0°C. Both storage temperature (0°C and 4°C) resulted in a decrease in Salmonella count. The results were more significant for Listeria strains, because during the storage at 4°C Listeria count increased in all batches, while for samples stored at 0°C a significant reduction in microbial load except in control batch was observed. Sensitivity differences of inoculated strains against temperature and CO2 concentrations were also discussed.

In situ production of pediocin PA-1 like bacteriocin by different genera of lactic acid bacteria in soymilk fermentation and evaluation of sensory properties of the fermented soy curd.

The lactic acid bacteria (LAB) are found to produce bacteriocins with enhanced nutritive properties in the fermented foods. In the present study, the ability of LAB cultures (Pediococcus acidilactici NCIM 5424, Enterococcus faecium NCIM 5423 and Lactobacillus plantarum Acr2) to produce pediocin PA-1 like bacteriocin was evaluated during soymilk fermentation. The isolates E. faecium NCIM 5423 and Lb. plantarum Acr2 were able to produce bacteriocin as well as ferment soymilk within 6h of incubation. Upon plating the cultures E. faecium NCIM 5423 and Lb. plantarum Acr2 in soymilk were found to be viable even after 15days of storage at 4°C. No significant variation was observed in the viable counts of E. faecium NCIM 5423 and Lb. plantarum Acr2 (P>0.05). The effect of crude bacteriocin on Listeria cells was evidenced through scanning electron microscope (SEM) photographs wherein cell membrane damage was observed. On co-cultivation of E. faecium NCIM 5423 and Lb. plantarum Acr2 individually with Listeria monocytogenes ScottA a decrease in the Listeria count was observed within 24h of incubation. However, during co-cultivation of ScottA with P. acidilactici NCIM 5424, no significant difference was observed in the viable counts (P>0.05). The pH, titratable acidity, pediocin activity, anti-oxidant property and sensory attributes for E. faecium NCIM 5423 were studied. It was observed that E. faecium NCIM 5423 fermented soymilk had an acceptable sensory score during storage period. Hence, such culture can be an ideal starter for development of functional foods with longer shelf life.

Evaluation of the dry sheet medium (Compact Dry LS) for screening of total Listeria count in food samples

The Compact Dry LS (CD-LS), ready-to-use dry sheet selective medium for Listeria species, was evaluated for inclusivity and exclusivity by using 107 strains including 20 Listeria species strains. All tested Listeria species strains other than Listeria seeligeri that grew as blue colored colony on CD-LS. CD-LS were compared with those on Oxford agar and ALOA agar as conventional methods using Listeria species inoculated from 100 food samples. The correlation coefficients between CD-LS and Oxford agar, and CD-LS and ALOA agar were 0.983 and 0.978, respectively. Our results suggested CDLS was a suitable alternative medium for screening of Listeria species.

Application of ozone sprays as a strategy to improve the microbial safety and quality of salmon fillets

Ozone research investigating the efficacy of spray application mechanisms for balancing microbial safety with chemical quality attributes of high lipid content fish is lacking. The objectives of this study were to investigate the influence of aqueous spray treatments of 1 mg/L and 1.5 mg/L ozone on the microbial and chemical quality indices of Atlantic salmon fillets inoculated with Listeria innocua as a surrogate species replacing the pathogen Listeria monocytogenes. In order to simulate industry processing parameters, number of passes under spray nozzles (1, 2, and 3) was also investigated.Listeria counts resulting from treatment with three passes of 1 mg/L ozone sprays were significantly lower (p ≤ 0.05) than non-ozonated controls on days 0, 3, 6, and 10 of refrigerated storage at 4 °C. At both ozone concentrations, Listeria counts were significantly influenced (p ≤ 0.05) by the number of passes under spray nozzles with increasing passes resulting in increasing reductions. The residual antimicrobial effectiveness of ozone at both concentrations against aerobic bacterial populations on treated samples was limited to <6 days storage at 4 °C. Lipid oxidation analyses indicate that ozone concentration did not significantly affect (p ≤ 0.05) TBARS but did influence headspace propanal with 1 mg/L ozone treatments yielding approximately 30% higher propanal values than spray treatments of 1.5 mg/L ozone. Both TBARS and propanal levels were significantly influenced (p ≤ 0.05) by storage time, yet the number of spray passes did not significantly affect either measure of lipid oxidation. Our results indicate that ozone sprays at concentrations up to 1.5 mg/L are effective in reducing initial counts of aerobic bacterial populations and in significantly reducing (p ≤ 0.05) counts of L. innocua without causing significant increases in lipid oxidation levels in farmed Atlantic salmon fillets during refrigerated storage at 4 °C.

Evaluation of the effects of fermentation of buffalo curd and acidity on survival kinetics of &lt;i&gt;Listeria monocytogenes&lt;/i&gt;

Listeria monocytogenes , a food-borne pathogen often found in milk and milk products, causes listeriosis in pregnant women, newborns, elderly and immuno-suppressed people. There have been reports on survival of Listeria in various milk products in the world, but comparable studies on the survival of Lis-teria in buffalo curd have not been reported. Therefore, the present study investigated the effects of fer-menting buffalo milk and acidity on survival kinetics of L. monocytogenes . L. monocytogenes FSTLC2 and lactic acid starter cultures were aseptically introduced to boiled and cooled buffalo milk (fat 6%, protein 4%) and the mix was allowed to ferment at ambient temperature (26 ± 2 o C) for 18h. The Lis-teria count on Modified Oxford Agar (MOX; Oxoid Ltd.), lactic acid bacteria count on MRS Agar (Oxoid Ltd.), pH and titratable acidity were determined in the beginning and at 2h intervals during fer-mentation. The effect of pH/acidity on Listeria was determined by introducing Listeria to buffalo milk with pH values (pH 4.0, 4.5, 5.0, 5.5) adjusted using 88% lactic acid (BDH Chemicals) and enumerating Listeria on MOX Agar at 12h intervals for 96h. It was observed in the present study that Listeria count decreased over time and after 16h of fermentation of buffalo milk at ambient temperature, Listeria can-not be detected on MOX Agar. The pH value decreased from 6.8 to 4.1 and titratable acidity (lactic acid %) increased from 0% to 1.2 % during fermentation. Fermentation appeared to be an effective preser-vation technique in eliminating Listeria in buffalo curd. The pH value of 5.5 appeared to be the critical pH for inactivation of Listeria as no growth of Listeria was observed below pH 5.5. The total inactiva-tion of Listeria in buffalo milk appeared to be due to lowering of pH coupled with increasing titratable acidity as well as action of bacteriocins, especially Nisin, produced by lactic acid bacteria during fer-mentation. It is clear that fermentation is an effective tool in inactivating Listeria in buffalo milk. Keywords : Listeria monocytogenes , Buffalo curd, Fermentation, Inactivation, Acidity DOI: http://dx.doi.org/10.4038/tare.v13i4.3292 TARE 2010; 13(4): 94-98

Airborne Listeria spp. in the red meat processing industry

The aim of this study was to determine the potential presence of the airborne Listeria spp. and its correlation with the aerobic mesophilic bacteria and Listeria carcass contamination in three red meat slaughtering and three processing plants. Airborne L. seeligeri and L. innocua were determined using 8 (5.06%, n = 158) air samples taken on the locations characteristic for aerosol generating and in a chilly environment. The positive airborne samples of Listeria spp. were in an insignificant (P &amp;gt; 0.05) relation with the highest airborne bacteria counts. On the carcass, only 1 positive case (0.69%, n = 144) of L. innocua was determined, presumably owing to the low airborne Listeria counts and its unpredictable settling rates. In addition, insignificant (P &amp;gt; 0.05) influences of air moisture and airflow on the airborne Listeria were found. Nevertheles, the methods currently used to determine the airborne Listeria and its relationships to aerosol viable mesophilic bacteria and carcass contamination need to be reconsidered in future investigations.

Bacteriophage biocontrol of Listeria monocytogenes on soft ripened white mold and red-smear cheeses.

Soft-ripened cheeses belong to the type of food most often contaminated with Listeria monocytogenes, and they have been implicated in several outbreaks of listeriosis. Bacteriophages represent an attractive way to combat foodborne pathogens without affecting other properties of the food. We used the broad host range, virulent Listeria phage A511 for control of L. monocytogenes during the production and ripening phases of both types of soft-ripened cheeses, white mold (Camembert-type) cheese, as well as washed-rind cheese with a red-smear surface (Limburger-type). The surfaces of young, unripened cheese were inoculated with 10(1)-10(3) cfu/cm(2)L. monocytogenes strains Scott A (serovar 4b) or CNL 10(3)/2005 (serovar 1/2a). Phage was applied at defined time points thereafter, in single or repeated treatments, at 3 × 10(8) or 1 × 10(9) pfu/cm(2). With Scott A (10(3) cfu/cm(2)) and a single dose of A511 (3 × 10(8) pfu/cm(2)) on camembert-type cheese, viable counts dropped 2.5 logs at the end of the 21 day ripening period. Repeated phage application did not further inhibit the bacteria, whereas a single higher dose (1 × 10(9) pfu/cm(2)) was found to be more effective. On red-smear cheese ripened for 22 days, Listeria counts were down by more than 3 logs. Repeated application of A511 further delayed re-growth of Listeria, but did not affect bacterial counts after 22 days. With lower initial Listeria contamination (10(1)-10(2) cfu/cm(2)), viable counts dropped below the limit of detection, corresponding to more than 6 logs reduction compared to the control. Our data clearly demonstrate the potential of bacteriophage for biocontrol of L. monocytogenes in soft cheese.

INHIBITORY EFFECT OF CLOVE OIL (SYZIUM AROMATICUM) AGAINST LISTERIA MONOCYTOGENES CELLS INCUBATED IN FRESH‐CUT SALMON

ABSTRACT The ability of Listeria monocytogenes to survive and grow at refrigeration temperature in a wide spectrum of foods is a public health concern. Essential oil of clove (Syzium aromaticum) was investigated for its ability to inhibit the growth of this pathogen in food. Three strains of L. monocytogenes (106 cfu/g) were inoculated on fresh‐cut salmon and stored (4C and 25C) for 2 weeks. The effect of clove oil (1% and 2%) on the growth of cells was determined. The results proved that all strains survived and grew at 4C and 25C. In addition, we noted that the growth was inhibited under storage conditions in the presence of clove oil (1% or 2%). Listeria counts in treated samples were 1–4 log10 cfu/g less compared with controls at different intervals during storage. The results revealed the potential of clove oil as a natural preservative in fresh salmon.PRACTICAL APPLICATIONS Listeria monocytogenes continues to be a significant threat to the safety of the food supply. It was isolated from a variety of foods and ready‐to‐eat products and generated a serious public health problem. Because of its growth capacity at refrigerating temperatures and its survival in diverse environments, the control of L. monocytogenes is becoming a challenge.Several agents were used to suppressing contamination by L. monocytogenes, such as the addition of biopreservatives and essential oil. Spices and herbs are commonly used in food preservation because they do not have a strong effect on food quality unlike chemical and synthetic compounds. Clove oil, contains eugenol which when tested on various agar media, has antimicrobial properties, and it was able to restrict the proliferation of L. monocytogenes in food products. For this reason, clove oil has good potential as an antilisterial substance in food preservation as it may be more acceptable to consumers.

Population dynamics of two antilisterial cheese surface consortia revealed by temporal temperature gradient gel electrophoresis

BackgroundSurface contamination of smear cheese by Listeria spp. is of major concern for the industry. Complex smear ecosystems have been shown to harbor antilisterial potential but the microorganisms and mechanisms involved in the inhibition mostly remain unclear, and are likely related to complex interactions than to production of single antimicrobial compounds. Bacterial biodiversity and population dynamics of complex smear ecosystems exhibiting antilisterial properties in situ were investigated by Temporal temperature gradient gel electrophoresis (TTGE), a culture independent technique, for two microbial consortia isolated from commercial Raclette type cheeses inoculated with defined commercial ripening cultures (F) or produced with an old-young smearing process (M).ResultsTTGE revealed nine bacterial species common to both F and M consortia, but consortium F exhibited a higher diversity than consortium M, with thirteen and ten species, respectively. Population dynamics were studied after application of the consortia on fresh-produced Raclette cheeses. TTGE analyses revealed a similar sequential development of the nine species common to both consortia. Beside common cheese surface bacteria (Staphylococcus equorum, Corynebacterium spp., Brevibacterium linens, Microbacterium gubbeenense, Agrococcus casei), the two consortia contained marine lactic acid bacteria (Alkalibacterium kapii, Marinilactibacillus psychrotolerans) that developed early in ripening (day 14 to 20), shortly after the growth of staphylococci (day 7). A decrease of Listeria counts was observed on cheese surface inoculated at day 7 with 0.1-1 × 102 CFU cm-2, when cheeses were smeared with consortium F or M. Listeria counts went below the detection limit of the method between day 14 and 28 and no subsequent regrowth was detected over 60 to 80 ripening days. In contrast, Listeria grew to high counts (105 CFU cm-2) on cheeses smeared with a defined surface culture.ConclusionsThis work reports the first population dynamics study of complex smear ecosystems exhibiting in situ antilisterial activity. TTGE revealed the presence of marine lactic acid bacteria that are likely related to the strong Listeria inhibition, as their early development in the smear occurred simultaneously with a decrease in Listeria cell count.

Survival of Listeria monocytogenes after cleaning and sanitation of wooden shelves used for cheese ripening

The effect of cleaning and heat disinfection processes of wooden shelves used for cheese ripening on the survival of Listeria monocytogenes was examined. The cut boards were inoculated with a suspension containing 5.5 × 107 colony forming units (CFU)/ml of L. monocytogenes. Survival of L. monocytogenes was investigated in the wood shavings. During the 24-hour incubation period in a humidity chamber, Listeria counts increased by 0.8 log units on average (n = 9), compared to results 1 h after incubation, indicating the absence of antimicrobial properties of the wood in use. Additionally, after incubation for 24 h at room temperature, the boards were cleaned by soaking them for 15 min in a solution of hot alkaline detergent followed by brushing and rinsing with warm water. Some of the cleaned boards were subsequently heat treated at 80°C for 5 min and at 65°C for 15 min, respectively. The cleaning procedure alone was not sufficient to render L. monocytogenes from the upper 2 mm wood layer inactive. In the case of both temperature-time combinations for heat disinfection, however, L. monocytogenes was not detectable. The present study shows that the use of wooden shelves does not affect the hygienic safety of cheeses if such shelves are in good repair and are thoroughly cleaned and sanitized by heat treatment. Therefore, there is no reason to replace wood employed in cheese ripening processes with other materials.

Combined effect of enterocin AS-48 and high hydrostatic pressure to control food-borne pathogens inoculated in low acid fermented sausages

The single and combined effects of enterocin AS-48 and high hydrostatic pressure (HHP) on Listeria monocytogenes, Salmonellaenterica, and Staphylococcus aureus was investigated in fuet (a low acid fermented sausage) during ripening and storage at 7 degrees C or at room temperature. AS-48 (148 AU g(-1)) caused a drastic 5.5 log cfu g(-1) decrease in L. monocytogenes (P<0.001) and a significant (P<0.01) inhibition (1.79 logs) for Salmonella at the end of ripening (10 d). After pressurization (400 MPa) and storage Listeria counts remained below 5 cfu g(-1) in all fuets containing AS-48 (pressurized or not). HHP alone had no anti-Listeria effect. HHP treatment significantly reduced Salmonella counts, with lowest levels in pressurized fuets with AS-48. S. aureus showed similar growth for all treatments and storage conditions. These results indicate that AS-48 can be applied alone to control L. monocytogenes and combined with HHP treatment to control Salmonella in fuets.

Specific detection and quantitative enumeration ofListeriaspp. using fluorescentin situhybridization in combination with filter cultivation (FISHFC)

To design a rapid specific method for enumeration of viable Listeria spp. using the fluorescence in situ hybridization with filter cultivation (FISHFC) method. The probe, Lis-1400, was designed from the 23S rRNA region of the Listeria genome, and labelled with 5'-carboxy-tetramethyl-rhodamine-N- hydroxy-succinimide-ester. Fluorescence was observed for all Listeria species but not for any organisms from the other genera, suggesting Lis-1400 is highly specific for Listeria spp. For purposes of filter cultivation prior to hybridization, hydrophilic polypropylene membrane filters gave better contrast between fluorescing colonies and background fluorescence. This was because of a high S/N ratio (fluorescence intensity of each microcolony/fluorescence intensity of background noise) after FISH treatment. Results were achievable in 14 h using Lis-1400-aided FISHFC as compared with 4-7 days required for confirmation of Listeria spp. by conventional plate count methods. Moreover, viable Listeria counts in selected food samples showed no significant differences between Lis-1400-aided FISHFC and conventional methods. The Lis-1400-aided FISHFC method is more efficient than conventional methods for enumeration of viable Listeria spp. in food samples. For enumeration of Listeria spp., Lis-1400-aided FISHFC method is equally accurate yet faster than conventional plate count methods, and can be valuable in the control of listeriosis.

Growth and bacteriocin production by lactic acid bacteria in vegetable broth and their effectiveness at reducing Listeria monocytogenes in vitro and in fresh-cut lettuce

The fresh-cut fruit and vegetable industry is searching for alternatives to replace chemical treatments with biopreservative approaches that ensure the safety of the product and fulfil consumer preferences for minimally processed foods. In this study, the use of bacteriocins produced by lactic acid bacteria has been tested as a substitute for chemical disinfection of fresh-cut iceberg lettuce. First, the ability of several non-plant origin bacteriocinogenic strains (nisin Z +, plantaricin C +, lacticin 481 +, coagulin + or pediocin PA-1 +) to grow in a lettuce extract at 4 °C, 10 °C and 32 °C was tested. All strains were able to grow, but bacteriocin production was predominantly detected at 32 °C. Addition of bacteriocinogenic supernatants (nisin +, coagulin + and a nisin–coagulin + cocktail) to tryptic-soy agar plates inoculated with Listeria monocytogenes reduced Listeria counts by approximately 1–1.5 log units compared with the control plates without bacteriocin, after 48 h of storage at 4 °C. The effect of washing with bacteriocin-containing solutions on survival and proliferation of Listeria monocytogenes was also evaluated in fresh-cut lettuce packaged in macro-perforated polypropylene bags and stored for 7 days at 4 °C. Washing fresh-cut lettuce with these solutions decreased the viability of Listeria monocytogenes by 1.2–1.6 log units immediately after treatment, but, during storage at 4 °C, bacteriocin treatments only exerted minimal control over the growth of the pathogen. Natural microbiota were little affected by bacteriocins during storage.

Effectiveness of cell-adsorbed bacteriocin produced by Lactobacillus curvatus CWBI-B28 and selected essential oils to control Listeria monocytogenes in pork meat during cold storage

To study the effectiveness of a combination of cell-adsorbed bacteriocin (CAB; a suspension of producer cells on which maximum bacteriocin has been immobilized by pH adjustments) of a Lactobacillus curvatus strain with oregano or savory essential oil to control Listeria monocytogenes in pork meat at 4 degrees C. The antimicrobial activity of the CAB and six different essential oils was tested by the well diffusion assay against L. monocytogenes M, Escherichia coli 10536 and Salmonella serotype Typhi CWBI-H1. The anti-Listeria activity of the CAB and oregano or savory essential oils was also investigated in pork meat. The results of the well diffusion assay showed that CAB was only inhibitory to L. monocytogenes while savory and oregano essential oils were the most active against the three indicator bacteria. In pork meat, Listeria counts have declined from c. 10(2) CFU g(-1) to below the detectable limit during the first week of storage in samples treated with CAB or oregano essential oil and in those treated with CAB combined with oregano or savory essential oil. However, the counts of L. monocytogenes have increased after the third week of storage in all samples with the exception of those treated with the combination of CAB and oregano essential oil. The combination of CAB with savory essential oil resulted in a 2-week delay of the growth rebound compared with samples treated with CAB alone. Addition of oregano or savory essential oil exhibited a synergistic effect with CAB to control L. monocytogenes in pork meat during storage at 4 degrees C. The combination of CAB with oregano or savory essential oil may be effectively used in meat industry to enhance the safety and stability of meat products.

Comparison of Media and Sampling Locations for Isolation of Listeria monocytogenes in Queso Fresco Cheese

Listeriosis associated with Hispanic-style soft cheese is an ongoing public health concern. Although rapid detection methods based on molecular and immunological technologies have been applied successfully for detecting Listeria monocytogenes in foods, obtaining isolates of the pathogen is a critical procedure for epidemiologic studies and regulatory analysis. Oxford agar, a medium recommended by the U.S. Food and Drug Administration Bacteriological Analytical Manual (BAM) to isolate L. monocytogenes from cheese, is unable to differentiate L. monocytogenes from other Listeria species. Hence, two selective isolation media, L. monocytogenes blood agar (LMBA) and Rapid 'L. mono agar (RLMA), were compared with Oxford agar for isolating L. monocytogenes from cheese. Queso fresco cheese was inoculated at 100 or 101 CFU/g with a five-strain mixture of L. monocytogenes or with the five-strain L. monocytogenes mixture and Listeria innocua. Cheese samples were stored at 21, 12, and 4°C and Listeria counts were determined at 3, 7, and 10 days; 7, 10, 14, 21 days; and 2, 4, 8, and 12 weeks postinoculation, respectively. Surface and interior cheese samples as well as liquid exudate produced during storage were assayed individually to determine differences in Listeria contamination at different sampling locations. L. monocytogenes was more easily differentiated from L. innocua on RLMA than LMBA and Oxford agar. Similar L. monocytogenes counts (ca. 104 CFU/g) were obtained on the last sampling day on the surface and interior of cheese samples (P > 0.05) for all storage temperatures and both initial inoculation levels, but smaller cell numbers were detected in the exudate produced during storage. In addition, simultaneous inoculation of L. innocua with L. monocytogenes did not affect the final L. monocytogenes counts in the cheese. The amount of exudate released from the cheese and decrease of pH correlated with storage temperature. More exudate was produced and a greater decrease of pH occurred at 21°C than at 12 or 4°C. Our results indicate that RLMA is a suitable medium for isolating L. monocytogenes from queso fresco cheese. Higher counts of L. monocytogenes were obtained from surface and interior samples of cheese than from the exudate of the cheese during storage. In addition, pH may be a useful indicator of improperly stored queso fresco cheese.

Bacteriocin Activity by Lactobacillus curvatus CWBI-B28 To Inactivate Listeria monocytogenes in Cold-Smoked Salmon during 4°C Storage

The inhibition effectiveness of a bacteriocin produced by Lactobacillus curvatus CWBI-B28 against Listeria monocytogenes was investigated in cold-smoked salmon during storage at 4°C. Three bacteriocin-based strategies for the control of L. monocytogenes in foods (i.e., producing bacteriocin in situ, spraying with partially purified bacteriocin, and packaging in bacteriocin-coated plastic film), plus a newly developed method that uses cell-adsorbed bacteriocin (i.e., a suspension of producer cells on which maximum bacteriocin has been immobilized by pH adjustments), were assessed. Although all the approaches inactivated L. monocytogenes in cold-smoked salmon, various efficacy levels were observed. The behavior of L. monocytogenes was similar in samples treated with either partially purified bacteriocin or in situ bacteriocin production. In both of these cases, the counts of the pathogen declined to below the detectable limit of 0.7 log CFU/cm2 within the first week, but a ∼0.95- and 1.3-log increase, respectively, occurred after day 14. The bioactive packaging film resulted in a slower inactivation of the pathogen but prevented any subsequent increase in the CFU throughout 22 days of storage at 4°C. Application of the cell-adsorbed bacteriocin was shown to be the most effective means, as it resulted in a complete inactivation of the pathogen within 3 days, and no increase in Listeria counts occurred up to 22 days.

Effect of immersion solutions containing enterocin AS-48 on Listeria monocytogenes in vegetable foods.

The effect of immersion solutions containing enterocin AS-48 alone or in combination with chemical preservatives on survival and proliferation of Listeria monocytogenes CECT 4032 inoculated on fresh alfalfa sprouts, soybean sprouts, and green asparagus was tested. Immersion treatments (5 min at room temperature) with AS-48 solutions (25 microg/ml) reduced listeria counts of artificially contaminated alfalfa and soybean sprouts by approximately 2.0 to 2.4 log CFU/g compared to a control immersion treatment in distilled water. The same bacteriocin immersion treatment applied on green asparagus had a very limited effect. During storage of vegetable samples treated with immersion solutions of 12.5 and 25 microg of AS-48/ml, viable listeria counts were reduced below detection limits at days 1 to 7 for alfalfa and soybean sprouts at 6 and 15 degrees C, as well as green asparagus at 15 degrees C. Only a limited inhibition of listeria proliferation was detected during storage of bacteriocin-treated alfalfa sprouts and green asparagus at 22 degrees C. Treatment with solutions containing AS-48 plus lactic acid, sodium lactate, sodium nitrite, sodium nitrate, trisodium phosphate, trisodium trimetaphosphate, sodium thiosulphate, n-propyl p-hydroxybenzoate, p-hydoxybenzoic acid methyl ester, hexadecylpyridinium chloride, peracetic acid, or sodium hypochlorite reduced viable counts of listeria below detection limits (by approximately 2.6 to 2.7 log CFU/g) upon application of the immersion treatment and/or further storage for 24 h, depending of the chemical preservative concentration. Significant increases of antimicrobial activity were also detected for AS-48 plus potassium permanganate and in some combinations with acetic acid, citric acid, sodium propionate, and potassium sorbate.

Antilisterial Activity of a Carnobacterium piscicola Isolated from Brazilian Smoked Fish (Surubim [Pseudoplatystoma sp.]) and Its Activity against a Persistent Strain of Listeria monocytogenes Isolated from Surubim

Data on the prevalence and growth of Listeria monocytogenes in lightly preserved fish products from subtropical and tropical regions are very scarce. Our research describes L. monocytogenes that was detected in 5% of the packages of cold-smoked surubim, a native Brazilian freshwater fish that we analyzed, and shows that the strains isolated were of the same random amplified polymorphic DNA subtype as the strains that were isolated from the same factory 4 years earlier. A bacteriocinogenic strain of Carnobacterium piscicola (strain C2), isolated from vacuum-packed cold-smoked surubim, and two C. piscicola strains, isolated from vacuum-packed, cold-smoked salmon, were capable of limiting or completely inhibiting the growth of an L. monocytogenes (strain V2) isolated from surubim in fish peptone model systems incubated at 10°C. Mono-cultures of L. monocytogenes reached 108 CFU/ml (g), whereas the growth of L. monocytogenes was completely inhibited by C. piscicola C2. The bacteriocinogenic C. piscicola A9b+ and its nonbacteriocinogenic mutant A9b− reduced maximum Listeria levels by 2 to 3 log units. Both bacteriocinogenic C. piscicola strains prevented listerial growth in cold-smoked fish juices (surubim and salmon). Although the carnobacteria grew poorly on cold-smoked surubim at 10°C, the strains were able to reduce maximum Listeria counts by 1 to 3 log units in an artificially inoculated product (surubim). We conclude that Brazilian smoked fish products harbor L. monocytogenes and should be stabilized against the growth of the organism. C. piscicola C2 has the potential for use as a bioprotective culture in surubim and other lightly preserved fish, but further studies are required to optimize its effect.

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.

Control of Listeria monocytogenes in model sausages by enterocin AS-48

In this work we describe the control of Listeria monocytogenes CECT 4032 in sausage by adding the enterocin AS-48 producer strains Enterococcus faecalis A-48-32 and Enterococcus faecium S-32-81, and also by adding a semi-purified preparation of the bacteriocin. Addition of preformed AS-48 caused a significant decrease ( P<0.01) in the number of viable listeria even at the lowest bacteriocin concentration tested (112 AU/g). At a higher concentration (225 AU/g) listeria were below the detection level (1.99 log units/g) in meat at 3 days of incubation but growth of listeria was observed again after 9 days. For an AS-48 concentration of 450 AU/g, no viable listeria were detected after 6 and 9 days of incubation. When E. faecalis A-48-32 was used as inoculum at approximately 10 7 cfu/g, listeria counts decreased progressively from start of experiment, being below detection level at day 9. The best results were obtained with E. faecium S-32-81, since listeria were undetectable at 6 days of incubation. Bacteriocin concentrations in samples reached concentrations of 60 and 80 AU/g for strains A-48-32 and S-32-81, respectively. These results clearly indicate that AS-48 can be used in the control of L. monocytogenes in sausages.