Lactic acid bacteria biodiversity in Italian marinated seafood salad and their interactions on the growth of Listeria monocytogenes

The aim of this research was to study the biodiversity of lactic acid bacteria (LAB) in marinated seafood salad (pH 5.0) and their interaction on the growth of Listeria monocytogenes. LAB were highly present in the samples considered in this study, reaching values of 8.0 log cfu/g at the end of product’s shelf-life. A high biodiversity in terms of LAB species and strains was detected by means of RAPD–PCR within the 171 bacterial isolates collected. Among them Lactobacillus curvatus, Lactobacillus sanfranciscensis and Enterococcus were present in all the salad batches considered. Three challenge tests against L. monocytogenes were carried out in order to assess the growth of this pathogen in the presence of dominant populations of LAB. L. monocytogenes tended to decrease in time, suggesting that a stable concentration of LAB inhibits the development of this pathogenic micro-organism.     

Lactic acid bacteria in an alginate film inhibit Listeria monocytogenes growth on smoked salmon

Antimicrobial packaging with lactic acid bacteria incorporated into the film matrix is a novel approach for controlling the growth of food-borne pathogens in ready-to-eat food. The overall objective of this study was to assess the effect of two strains of lactic acid bacteria (LAB) and nisin trapped in an alginate matrix, on Listeria monocytogenes growth on vacuum packed cold-smoked salmon. A film was formulated containing two LAB strains and nisin (100 IU/mL). LAB viability and bacteriocin like substance production (BLS) were assessed using the plate antagonism technique. To check the film antagonistic activity, pieces of salmon (4.0 × 4.0 cm²), inoculated with L. monocytogenes at a final concentration of 10⁴ CFU/cm², were covered with film containing both LAB strains plus nisin and stored at 4 °C. L. monocytogenes colonies on OXA agar were counted after 0, 7, 14, 21 and 28 days to evaluate pathogen inhibition. All treatments led to effective diffusion of the BLS that inhibited L. monocytogenes for 20 days after film preparation, with inhibition zones of 5.7 cm² for film coupons of 8 mm in diameter. After 28 days, salmon pieces covered with the film without inhibitors showed an increase of 2.4 log cycles in L. monocytogenes growth. In contrast, films with either LAB strain or a combination of both strains and nisin had a bacteriostatic effect on the pathogen over a period of 28 days, which exceeds the industrial standard shelf life for smoked salmon. The results demonstrate that these films inhibit L. monocytogenes growth on salmon during refrigerated storage.     

Non starter lactic acid bacteria biofilms: A means to control the growth of Listeria monocytogenes in soft cheese

The possibility to consider non starter lactic acid bacteria (NSLAB) biofilms as a means to control the growth of Listeria monocytogenes in soft cheeses was evaluated. In particular, the aptitude to form biofilm of four NSLAB strains (Lactobacillus plantarum DSM1055, Lactobacillus casei DSM20011, Lactobacillus curvatus DSM20019 and Lactobacillus paracasei DSM20207) was investigated. All tested strains were able to form biofilm on stainless steel and the highest quantities were produced when NSLAB were present simultaneously (about 6 Log CFU cm^?2). Then, experimental cheeses were made in presence of chips containing 7-days NSLAB biofilms and inoculated with L. monocytogenes (about 2 Log CFU g^?1). Results demonstrated that NSLAB biofilms can be considered a useful means to delay the growth of L. monocytogenes in soft cheeses: the maximum cell load attained at the stationary phase was about 6 Log CFU g^?1 in experimental cheeses against about 7 Log CFU g^?1 observed in control samples.     

Antimicrobial activity of lacticin 3147 against oenological lactic acid bacteria. Combined effect with other antimicrobial agents

This study investigated the potential use of lacticin 3147 to control growth of wine lactic acid bacteria (LAB) aiming to develop new approaches for reducing the content of sulphites in wine. Lacticin 3147, potassium metabisulphite and eucalyptus extract interactions were also evaluated. Inhibitory activity of the bacteriocin demonstrated to be strain-dependent and the highest antimicrobial effect was established at the exponential growth phase. Low lacticin 3147 IC₅₀ values (10.46–57.97 UA/mL) were measured for most of the strains, except for Lactobacillus casei CIAL-51 which were remarkably resistant (IC₅₀ = 141.70 UA/mL); Oenococcus oeni CIAL-117 were the most sensitive strain (IC₅₀ = 10.46 UA/mL). A synergistic effect of either lacticin 3147 or eucalyptus extract in combination with metabisulphite in inhibiting LAB growth was measured. Addition of lacticin 3147 to growing cultures of Lactobacillus plantarum CIAL-92 caused effective cell lysis, decrease of cell viability and extensive membrane damage, differing from the effects detected after addition of eucalyptus extract. To our knowledge, this is the first study that reports the inhibitory activity of this bacteriocin against oenological LAB and its potential role in the wine industry. The potential of eucalyptus extract as antimicrobial agent in winemaking is also highlighted. A combination of these two antimicrobials with metabisulphite could represent a promising approach to preserve LAB spoilage in wine and, consequently, eliminate or reduce the addition of SO₂.     

Inhibitor activities of two Lactobacillus strains,isolated from sourdough,against rope-forming Bacillus strains

The effect of two different sourdoughs, produced with Lactobacillus plantarum LMO25 and Lactobacillus alimentarius LMO7, with antimicrobial activities on inhibition of rope-forming Bacillus strains in wheat bread was studied. Addition of 15% or 20% low pH (pH 3.5–4.0) sourdough to bread dough, which were produced by using two strains (Lb. plantarum LMO25 and Lb. alimentarius LMO7) separately, prevented the generation of visual rope caused by Bacillus subtilis and Bacillus licheniformis. However, adding 10% sourdough was not enough to prevent the generation of visual rope. When repeated with sourdoughs with a higher pH (pH > 4), additives at 10% or 15% did not prevent the generation of rope, whereas additives at 20% prevented the generation of visual rope caused by both B. subtilis and B. licheniformis.     

Microbial profile of buffalo sausage during processing and storage

A study was made on the microbial levels of buffalo sausage during preparation and storage at 4 ± 1 °C. Microbial counts in raw minced meat were, total plate count (TPC) (log cfu/g) 5.41 ± 0.25; coliforms (MPN/g) 23.2; Staphylococcus aureus (log cfu/g) 1.57 ± 0.11; yeasts and molds (log cfu/g) 2.29 ± 0.07 and lactic acid bacteria (LAB) (log cfu/g) 0.60 ± 0.20. Sausage emulsion showed similar trend in microbial counts with minimal microbial contamination during the preparation of emulsion. Cooked buffalo sausage gave the following microbial counts: TPC (log cfu/g) 3.75 ± 0.31; coliforms (MPN/g) 0.2; LAB (log cfu/g) 0.07 ± 0.01; yeast and molds (log cfu/g) 0.72 ± 0.07. S. aureus, Clostridium perfringens and Bacillus cereus were not detected in cooked sausages. These results indicate that steam cooking for 45 min followed in the study was effective in reducing the microbial counts substantially. The investigation revealed that shelf life of cooked buffalo sausage was 31 days in either vacuum or CO₂ at 4 ± 1 °C. The results indicated that spoilage of vacuum packed cooked buffalo sausage was likely due to LAB while microflora other than LAB may be responsible for spoilage of CO₂ packed cooked buffalo sausage. The study suggests that measures such as low initial microbial counts, hygienic precautions during preparation of sausage, steam cooking for 45 min, vacuum or CO₂ packing and storage at 4 ± 1 °C would control the microbial growth and provide wholesomeness and safety to the buffalo sausage.     

Antimicrobial properties of selected essential oils in vapour phase against foodborne bacteria

The aim of this study was to identify antimicrobial properties of essential oils in vapour phase. In vitro antibacterial activity against five foodborne bacteria (Escherichia coli, Listeria monocytogenes, Pseudomonas aeruginosa, Salmonella enteritidis, Staphylococcus aureus) was evaluated by disc volatilization method. The results were expressed as minimum inhibitory concentrations (MIC) in μl/cm³ of air. Thirteen of the 27 essential oils were active at least against one bacterial strain in the range of tested concentrations (0.0083–0.53 μl/cm³). The best results were shown by Armoracia rusticana (MIC 0.0083 μl/cm³) against all of the strains, followed by Allium sativum > Origanum vulgare > Thymus vulgaris > Satureja montana, Thymus pulegioides > Thymus serpyllum > Origanum majorana > Caryopteris x clandonensis, Hyssopus officinalis, Mentha villosa, Nepeta x faassenii, Ocimum basilicum var. grant verte. In conclusion, certain essential oils are highly effective in vapour phase and could be used in control of foodborne bacterial pathogens.     

In vitro control of growth and ochratoxin A production by butylated hydroxyanisole in Aspergillus section Nigri species

This study was carried out to determine the efficacy of phenolic antioxidant butylated hydroxyanisole (BHA) under different interacting water activity (a_W) and temperature regimes on the lag phase and growth rate by Aspergillus section Nigri strains. In this experiment four A. section Nigri strains were used. Peanut meal extract agar (PMEA) was prepared at 2%. The a_W of the medium was adjusted to 0.995, 0.980 and 0.930, BHA at 1, 5, 10 and 20 mmol l^?1 was added to the basic medium. The plates were inoculated and incubated for 30 days at 18 and 25 °C. Radial growth rates (mm d^?1) and lag phase (h) were calculated. In control treatments, the growth rate decreased as water activity reduced in all strains assayed. At all a_W levels tested, BHA at 20 mmol l^?1 completely inhibited growth. In general, at 10 mmol l^?1 and 0.995 and 0.980a_W level, a significant reduction respect to control was observed. This antioxidant completely inhibited OTA production, at concentrations of 20 mmol l^?1, regardless of a_W used by all the strains evaluated.     

Effect of starter culture on growth of Staphylococcus aureus in sucuk

The effects of starter cultures, starter A (Pediococcus acidilactici + Lactobacillus curvatus + Staphylococcus xylosus), starter B (Lactobacillus sakei + Staphylococcus carnosus) and ripening period on growth of S. aureus were investigated in sucuk (Turkish dry-fermented sausage) production. Sucuk batters were inoculated with S. aureus (10⁶/g). S. aureus, Micrococcus/Staphylococcus, lactic acid bacteria and Enterobacteriaceae counts were determined during ripening, and pH and a_w were analyzed. Both starter cultures and ripening period had significant effects on the S. aureus, lactic acid bacteria, Micrococcus/Staphylococcus, Enterobacteriaceae counts and pH value (P < 0.01). The numbers of S. aureus increased by 1 log on the 3rd day in the control group, whereas a reduction was observed during ripening period in sucuk with starter cultures.     

In vitro antifungal activity of lactic acid bacteria against mycotoxigenic fungi and their application in loaf bread shelf life improvement

Food spoilage caused by mycotoxigenic fungi represents an important food safety problem. Lactic acid bacteria (LAB) are used as starter cultures in a larger number of food products. In this study, 16 strains of LAB were cultivated in MRS broth under anaerobiosis. Then, cell free supernatants were obtained by centrifugation and their antifungal activity against Aspergillus parasiticus and Penicillium expansum was tested using the disc-diffusion method. Furthermore, the LABs that showed in vitro antifungal activity were used in bread fermentation with yeast in order to study fungal growth inhibition and aflatoxin (AF) reduction in processed bread previously inoculated with A. parasiticus. The compounds present in the fermented medium of six LAB strains induced inhibition of P. expansum growth, whereas five probiotic strains produced antifungal compounds against A. parasiticus. The analysis by liquid chromatography coupled to mass spectrometry in tandem showed a reduction of the AF content in bread samples fermented with yeast and LABs. The reduction of AFs ranged from 84.1 to 99.9%. Moreover, bread sample studies showed a shelf life increase of about 3–4 days.     

Microbiology of retail mung bean sprouts vended in public markets of National Capital Region,Philippines

This study is the first attempt in the Philippines to conduct a region-wide assessment of the microbiological quality of retailed mung bean sprouts. Production and vending practices of selected stakeholders were also determined. Ninety-four percent of the samples tested positive for the presence of Salmonella spp. and some samples had Coliform and Escherichia coli counts as high as 5.90 and 5.50 log₁₀ CFU · g⁻¹, respectively. The TPC, YMC and LAB were established to be as high as 11.38, 5.90 and 10.47 log₁₀CFU · g⁻¹, respectively. The poor microbiological quality of most of the tested sprouts was attributed to unhygienic sprout production and retailing practices. To improve the microbiological quality of the retailed sprouts, adherence to sprouting industry best practices is strongly recommended.     

Characteristics of Batzos cheese made from raw,pasteurized and/or pasteurized standardized goat milk and a native culture

Experimental cheeses were made from raw (R), raw with starter (RS), pasteurized with starter (PS) and standardized, pasteurized with starter (PSS) goat milk to study the influence of the starter and pasteurization on the quality of Batzos cheese. Lactococcus lactis subsp lactis strains from raw milk cheese used as starter proliferated significantly (P < 0.05) only in PSS cheese. Heat treatment lowered (P < 0.05) the levels of most microbial groups. However, TC and LAB counts were higher (P < 0.05) in PSS cheese and this was accompanied by lower (P < 0.05) pH; thus, a higher rate of decrease of undesirable microorganisms in PSS cheese was recorded. Degradation of αs-casein was in R > RS and in PS > PSS, while a small reduction of β-casein during ripening and storage was recorded. The proportions of aminoacids and lipolysis products increased throughout ripening and storage.     

Prevalence of virulence genes in Escherichia coli isolated from food in Casablanca (Morocco)

This study was undertaken to determine the frequency of potentially pathogenic Escherichia coli (E. coli) strains among E. coli isolated from Casablanca, Morocco. The E. coli strains were isolated from ground beef (n = 140), turkey (n = 200), sausage (n = 120), seafood (n = 60), domestic water (n = 35) and well water (n = 50). The prevalence of E. coli was 48%, 45%, 35.5%, 30%, 8.3%, 0%, for well water, ground beef, turkey, sausage, sea food and domestic water, respectively. Two hundreds E. coli isolates were tested for the presence of 17 virulence genes associated with strains causing intestinal and extra-intestinal infections. The virulence genes included stx1, stx2, lt, st, hlyA, aggA, saa, astA, iucD, cnf1, eaeA, bfpA, ial, ipaH, afa, pap and sfa. PCR showed that 37% (74) of E. coli isolates carried one or more of these virulence genes. No virulence genes were found in E. coli strains isolated from sea food samples. In contrast, 10% of the ground beef samples, 18% of the turkey samples, 17.5% of sausage samples and 6% of well water contained specific factors for intestinal E. coli pathogens.     

In vitro and in situ screening of lactic acid bacteria and propionibacteria antifungal activities against bakery product spoilage molds

In the agri-food context, bioprotective cultures represent an interesting alternative to chemical preservatives. The aim of this study was to evaluate the in vitro and in situ antifungal activity of lactic acid bacteria (LAB) and propionibacteria against bakery product spoilage fungi. Firstly, the biodiversity of fungal contaminant in pound cakes and milk bread rolls, as well as the resistance to chemical preservatives of representative isolates (n = 21), were studied. Aspergillus and Eurotium species were the most dominant spoilage fungi and the most resistant towards the tested chemical preservatives. They were followed by Penicillium, Cladosporium and Wallemia spp. Secondly, an in vitro screening showed that the most active isolates against selected fungal targets belonged to the Leuconostoc spp., Lactobacillus reuteri and Lactobacillus buchneri groups among the 270 tested LAB, as well as to the Propionibacterium freudenreichi and Propionibacterium acidipropionici species for the 50 tested propionibacteria. Finally, in situ tests showed that cultures of isolates belonging to the Leuconostoc citreum, Lactobacillus sakei, Lactobacillus plantarum, Lactobacillus spicheri, L. reuteri and Lactobacillus brevis species could delay one or several target fungal growths after bakery product surface spraying. Moreover, different strain cultures led to delayed fungal growths after incorporation in milk bread rolls preparation. The combination of in vitro and in situ approaches allowed for the identification of bacteria exhibiting antifungal activity, providing future prospects for use as bioprotective cultures in bakery products.     

Lactic acid bacteria bioprotection applied to the malting process. Part I: Strain characterization and identification of antifungal compounds

Lactic acid bacteria (LAB) are of interest to cereal and beverage industry due to their contribution to the microbial safety and quality of fermented food/beverages. The main objectives of this study were to optimize the production of an antifungal cell-free-supernatant (cfs), based on lactic acid bacteria, using a wort-base substrate; and the subsequent identification of its active acid-base antifungal compounds. Two antifungal strains (Lactobacillus amylovorus DSM19280 and Lactobacillus reuteri R29) and a negative control strain (L. amylovorus DSM20552) were used. Barley based malt extract (wort) was produced and used as a fermentation substrate. Worts fermented with LAB were characterized in detail (total cell counts, pH, and total titratable acids (TTA) over time (0–120 h). A microtiter plate and an overlay agar based assay were used to evaluate the antifungal activity of the LAB cfs. Sugars, organic acids of the fermented and non-fermented wort (control) were analysed by HPLC. LAB antifungal metabolites were quantified using a QuEChERS method and an HPLC-UV/PDA system. Results show that wort produced from barley malt is a suitable substrate for LAB and dependent on the species and the fermentation time. The type of antifungal compounds can vary significantly. L. amylovorus DSM19280 cfs inhibited Fusarium culmorum spores at levels of 10⁴ spores.mL⁻¹, for 7 days, whereas L. reuteri R29 cfs inhibited up to 10⁵ spores.mL⁻¹ for the same time. Thirteen acid-base antifungal compounds were identified in L. reuteri R29 cfs and twelve in L. amylovorus DSM19280. Among them, phenyllactic, OH-phenylactic and benzoic acids were present at a significant concentration. This study demonstrates that the LAB wort-base cfs exhibits potent antifungal activity and is the ideal substrate for applications in a wide range of cereal products.     

Antimicrobial activity of clove (Syzgium aromaticum) oil in inhibiting Listeria monocytogenes on chicken frankfurters

The ability of Listeria monocytogenes to survive and grow at refrigeration temperature in some ready to eat (RTE) poultry products is a public health concern. The inhibitory effect of clove oil (1% and 2%, v/w) applied to the surface of RTE chicken frankfurters was determined on seven strains of L. monocytogenes inoculated at low (10²–10³ cfu/g) or high cell numbers (10⁴–10⁶ cfu/g), and stored at 5 °C for 2 weeks or at 15 °C for 1 week. All strains of L. monocytogenes survived and grew on control frankfurters at 5 °C and 15 °C but growth was inhibited under both storage conditions in the presence of either 1% or 2% clove oil. Depending on the sensory considerations, the addition of clove oil to frankfurters may be an effective strategy to control L. monocytogenes in chicken frankfurters.     

Use of lactic acid bacteria (LAB) biofilms for the control of Listeria monocytogenes in a small-scale model

The antilisterial activity in biofilms developed in a small-scale model by two LAB (lactic acid bacteria) bacteriocin producers (Lactobacillus plantarum 35d, Enterococcus casseliflavus IM 416K1) and by two non-producers (L. plantarum 396/1, Enterococcus faecalis JH2-2) was evaluated against Listeria monocytogenes NCTC 10888. The LAB biofilms showed the capability to influence the survival and the multiplication of the pathogen with differences among the strains. L. plantarum 35d displayed the highest efficacy reducing L. monocytogenes by 5.4 log in the planktonic population and by 3.9 log in the adherent population at the end of the experiment (10 days). L. plantarum 396/1 reduced L. monocytogenes by 3.8 log in the adherent cells and by 4.9 log in the planktonic cells and this outcome could be attributed to the pH reduction.The E. casseliflavus IM 416K1 biofilm caused a L. monocytogenes reduction of 3.7 log in the adherent cells and of 4.8 log in the planktonic cells and the role of the bacteriocin production seemed to be predominant as the pH values did not significantly decrease. This hypothesis is confirmed by a slight capability to influence the L. monocytogenes survival by the non-bacteriocinogenic E. faecalis JH2-2. Studies performed with L. monocytogenes in co-culture with a Pseudomonas putida strain, revealed a reduction of the antilisterial activity only for the biofilms produced by lactobacilli.     

Pathogenic microorganism survival in dulce de leche

In order to evaluate the survival of Salmonella typhimurium, Listeria monocytogenes, Escherichia coli O157:H7 and Staphylococcus aureus in dulce de leche, aliquots of this sweet were experimentally contaminated with these pathogenic microorganisms at 10³ (C1) and 10¹ (C2) bacterial cells per gram, and later analysed to evaluate microorganism viability after storage for 0, 1, 2, 3, 5, 10, 20 and 30 days. Salmonella and L. monocytogenes were recovered from all aliquots. E. coli O157:H7 and S. aureus were recovered from all aliquots at C1 concentration, and up to the 5th and 10th day, respectively, at C2 concentration. Relevant public-health danger pathogenic microorganisms were found to survive up to 30 days in dulce de leche.     

Biological control of postharvest diseases of peach with Cryptococcus laurentii

Cryptococcus laurentii was evaluated for its activity in reducing postharvest gray mold decay, blue mold decay and Rhizopus decay of peach caused by Botrytis cinerea, Penicillium expansum and Rhizopus stolonifer respectively, and in reducing natural decay development of peach fruits. The concentrations of antagonist had significant effects on biocontrol effectiveness: the higher the concentrations of the antagonist, the lower the disease incidence. At concentrations of C. laurentii at 1 × 10⁹ CFU ml⁻¹, the gray mold decay was completely inhibited after 4 days incubation at 25 °C, while the control fruit had 50% decay, when inoculated with B. cinerea spores suspension of 1 × 10⁵ spores ml⁻¹; no complete control of the blue mold or Rhizopus mold was observed, when peach fruits were stored at 25 °C for 4 days (challenged with P. expansum) or 5 days (challenged with R. stolonifer) respectively, but the decay was distinctly prevented, the incidence of blue mold or Rhizopus mold was reduced by 78.6% or 80% respectively, compared with control, at challenged with P. expansum or R. stolonifer spores suspension of 5 × 10⁴ spores ml⁻¹, respectively. C. laurentii significantly reduced the natural development of decay and did not impair quality parameters of fruit following storage at 2 °C for 30 days followed by 20 °C for 7 days.     

Effects of Zataria multiflora Boiss. essential oil and nisin on Bacillus cereus ATCC 11778

The effects of different concentrations of Zataria multiflora Boiss. essential oil (EO, 0.0%, 0.005%, 0.015%, 0.03% and 0.045%) and nisin (N, 0.0, 0.25, 0.5, 1.5 and 2.5 μg ml⁻¹), pH values (7.4, 6.5 and 6.0), temperatures (Ts, 10, 20 and 30 °C) and storage times (Ds, up to 43 days) on log₁₀ probability percentage of growth initiation (log P%) of one vegetative cell of Bacillus cereus in brain heart infusion broth were evaluated in a factorial design study. The log P% of the organism was significantly affected (P < 0.01) by the values of EO, N, pH, T and D.The combinations of T ⩾ 20 °C, EO ⩽ 0.03% and pH values (7.4, 6.5 and 6.0) could not obviously affect the growth of the organism in this study. Whereas, the strong inhibitory action was observed by increasing EO concentration to 0.045 at T ⩾ 20 °C and selected pH values (7.4, 6.5 and 6.0) and by decreasing temperature to 10 °C at EO ⩾ 0.015% and pH values used in this study. The inhibitory effect of N also was enhanced by decreasing storage temperature to 10 °C at the selected pH values (7.4, 6.5 and 6.0) in this study.The growth of the organisms was strongly affected by increasing EO concentration to 0.03% in combination with N concentrations used at the selected temperatures in this study. The growth of the organism was completely inhibited at combinations EO ⩾ 0.015%, N ⩾ 1.5 μg ml⁻¹, T ⩽ 30 °C and pH ⩽ 7.4 during 43 days of storage in this study. This synergistic effect of EO and N was enhanced in lower pH values (6.5 and 6.0) in the present study. The growth of organism was completely inhibited at combinations of EO ⩾ 0.005 and N ⩾ 1.5 μg ml⁻¹ at pH = 6.0, and EO ⩾ 0.03 and N ⩾ 0.5 μg ml⁻¹ at pH ⩽ 6.5 during the study at the selected Ts (30, 20 and 10 °C).