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Aims: Lactobacilli have drawn a great deal of attention due to the heterogeneity of their cells, as well as their remarkable ability to adapt to a range of harsh conditions. One such adaptation response by Lactiplantibacillus plantarum was forming a V-shaped cell structure in low acidic pH conditions. This study aims to identify low pH induced-biofilm formation in L. plantarum and Limosilactobacillus reuteri and their antagonistic interactions with an opportunistic pathogen, Pseudomonas aeruginosa PAO1., Methods and Results: Dual species bacterial interactions revealed a Lactobacilli biofilm-mediated suppression of PAO1 biofilms and pyocyanin pigment. Furthermore, filtrates from Lactobacilli grown at low acidic pH (pH 3.5) effectively arrest the pathogenesis of P. aeruginosa PAO1, including biofilm, swarming motility, and pyocyanin formation. Liquid chromatography-mass spectroscopy analysis revealed the presence of several Lactobacilli-secreted metabolites and dipeptides that exhibited strong LasR receptor antagonistic interactions in molecular docking analysis. Prolyl-tyrosine showed the best lasR antagonistic and PAO1 biofilm inhibitory activities of the identified dipeptides., Conclusion: Overall, the results will contribute to the development of anti-infective drugs against P. aeruginosa using Lactobacilli secreted dipeptides. Furthermore, the investigation identifies several dipeptides that can be extracted from V-shaped Lactobacilli for antimicrobial protein therapy against P. aeruginosa and other pathogens., (© The Author(s) 2024. Published by Oxford University Press on behalf of Applied Microbiology International.)

As the second most abundant transition element and a crucial cofactor for many proteins, zinc is essential for the survival of all living organisms. To maintain required zinc levels and prevent toxic overload, cells and organisms have a collection of metal transport proteins for uptake and efflux of zinc. In bacteria, metal transport proteins are well defined for model organisms and many pathogens, but fewer studies have explored metal transport proteins, including those for zinc, in commensal bacteria from the gut microbiota. The healthy human gut microbiota comprises hundreds of species and among these, bacteria from the Lactobacillaceae family are well documented to have various beneficial effects on health. Furthermore, changes in dietary metal intake, such as for zinc and iron, are frequently correlated with changes in abundance of Lactobacillaceae. Few studies have explored zinc requirements and zinc homeostasis mechanisms in Lactobacillaceae, however. Here we applied a bioinformatics approach to identify and compare predicted zinc uptake and efflux proteins in several Lactobacillaceae genera of intestinal relevance. Few Lactobacillaceae had zinc transporters currently annotated in proteomes retrieved from the UniProt database, but protein sequence-based homology searches revealed that high-affinity ABC transporter genes are likely common, albeit with genus-specific domain features. P-type ATPase transporters are probably also common and some Lactobacillaceae genera code for predicted zinc efflux cation diffusion facilitators. This analysis confirms that Lactobacillaceae harbor genes for various zinc transporter homologs, and provides a foundation for systematic experimental studies to elucidate zinc homeostasis mechanisms in these bacteria., (© The Author(s) 2023. Published by Oxford University Press.)

The present work aimed to produce a cupuassu juice (Theobroma grandiflorum) fermented by the probiotic bacterium Lactiplantibacillus plantarum Lp62 and to analyze its antioxidant potential, antimicrobial activity, and resistance to biological barriers. The fermented beverage showed an increase in the content of phenolics, flavonoids, and antioxidant potential. The culture showed antagonistic activity against pathogens, but this result was not observed when the juice was tested. The probiotic strain remained viable under refrigeration, even in an acidified environment, and survived simulated gastrointestinal transit in vitro. L. plantarum Lp62 showed 30% adherence to HT-29 intestinal cells and proved to be safe in terms of antibiotic resistance and production of virulence factors. Fermentation increased the functional characteristics of cupuassu juice. This drink proved to be a good vehicle for the delivery of the probiotic bacteria L. plantarum Lp62., (© The Author(s) 2023. Published by Oxford University Press on behalf of Applied Microbiology International.)

Aims: To analyse lactic acid bacteria (LAB) diversity and technological-functional and safety properties of strains present during spontaneous fermented quinoa sourdoughs., Methods and Results: Fermentation was performed by daily backslopping at 30°C for 10 days. Autochthonous LAB microbiota was monitored by a biphasic approach combining random amplified polymorphic DNA (RAPD)-PCR and rRNA gene sequencing with PCR-denaturing gradient gel electrophoresis (DGGE) analysis. Identification and intraspecies differentiation allowed to group isolates within nine LAB species belonging to four genera. A succession of LAB species occurred during 10-days backslopping; Lactobacillus plantarum and Lactobacillus brevis were detected as dominant species in the consortium. The characterization of 15 representative LAB strains was performed based on the acidifying capacity, starch and protein hydrolysis, γ-aminobutyric acid and exopolysaccharides production, antimicrobial activity and antibiotic resistance., Conclusion: Strains characterization led to the selection of Lact. plantarum CRL1905 and Leuconostoc mesenteroides CRL1907 as candidates to be assayed as functional starter culture for the gluten-free (GF) quinoa fermented products., Significance and Impact of the Study: Results on native LAB microbiota present during quinoa sourdough fermentation will allow the selection of strains with appropriate technological properties to be used as a novel functional starter culture for GF-fermented products., (© 2016 The Society for Applied Microbiology.)

Aims: This study aimed to evaluate the effects of inoculation of strains of lactic acid bacteria (LAB) isolated from sugarcane grown in a Brazil on the quality of corn silage., Methods and Results: Three strains of Lactobacillus buchneri (UFLA SLM11, UFLA SLM103 and UFLA SLM108), five strains of Lactobacillus plantarum (UFLA SLM08, UFLA SLM41, UFLA SLM45, UFLA SLM46 and UFLA SLM105), and one strain of Leuconostoc mesenteroides (UFLA SLM06) were evaluated at 0, 10, 30, 60 and 90 day after inoculating corn forage. The inoculation of the LAB strains did not influence the chemical composition of the silage, but pH, acetic acid and 1,2-propanediol were affected by treatment. The silages inoculated with UFLA SLM11 and SLM108 contained the lowest yeast and filamentous fungi counts during fermentation. Bacteria belonging to the Enterobacteriaceae family, Clostridium genus were detected in the silages inoculated with Lact. buchneri UFLA SLM 11, 103 and 108, as shown by DGGE analysis. Silages inoculated with Lact. buchneri UFLA SLM 11 showed higher aerobic stability., Conclusions: The Lact. buchneri UFLA SLM11 strain was considered promising as a starter culture or inoculant for corn silages., Significance and Impact of the Study: The selection of microbial inoculants for each crop promotes improvement of silage quality. Studies on the chemical and microbiological characteristics of silage provide useful information for improving ensiling techniques., (© 2015 The Society for Applied Microbiology.)

Sake is made from steamed rice, malted rice, and water. Sake production begins with the preparation of a small-scale starter (moto); the quality of moto significantly influences the flavor and richness of sake. In the traditional starter, yamahai-moto, the growth of naturally occurring lactic acid bacteria represses the putrefactive micro-organisms, whereas in the modern starter, sokujo-moto, this is achieved by adding lactic acid. In this study, the successive change in bacterial flora of yamahai-moto was analyzed by pyrosequencing 16S ribosomal RNA genes. Lactobacillus was dominant throughout the process (93-98%). Nitrate-reducing bacteria that have been generally assumed to be the first colonizers of yamahai-moto were scarcely found in the early stage, but Lactobacillus acidipiscis dominated. Lactobacillus sakei drastically increased in the middle stage. This is the first report, though one case study, to show how the early stage microbiota in Japanese yamahai-moto is varyingly controlled without nitrate-reducing bacteria using next-generation sequencing.

Aims: To screen and identify safe micro-organisms used during Douchi fermentation, and verify the feasibility of producing high-quality Douchi using these identified micro-organisms., Methods and Results: PCR-denaturing gradient gel electrophoresis (DGGE) and automatic amino-acid analyser were used to investigate the microbial diversity and free amino acids (FAAs) content of 10 commercial Douchi samples. The correlations between microbial communities and FAAs were analysed by statistical analysis. Ten strains with significant positive correlation were identified. Then an experiment on Douchi fermentation by identified strains was carried out, and the nutritional composition in Douchi was analysed. Results showed that FAAs and relative content of isoflavone aglycones in verification Douchi samples were generally higher than those in commercial Douchi samples., Conclusions: Our study indicated that fungi, yeasts, Bacillus and lactic acid bacteria were the key players in Douchi fermentation, and with identified probiotic micro-organisms participating in fermentation, a higher quality Douchi product was produced., Significance and Impact of the Study: This is the first report to analyse and confirm the key micro-organisms during Douchi fermentation by statistical analysis. This work proves fermentation micro-organisms to be the key influencing factor of Douchi quality, and demonstrates the feasibility of fermenting Douchi using identified starter micro-organisms., (© 2015 The Society for Applied Microbiology.)

Aim: To evaluate the suitability of marine lactic acid bacteria (LAB) as starter cultures for Sargassum sp. fermentation to enhance its antioxidant and anticoagulation activity., Methods and Results: LAB isolated from marine source were characterized for their ability to utilize seaweed as a sole carbon source and applied to Sargassum fermentation. Fermentation period was optimized by monitoring the fermented sample at regular interval for a period of 18 days. Results revealed that a fermentation period of 12 days was effective with maximum culture viability and other desirable characteristics such as pH, total titratable acidity, total and reducing sugars. Under optimum fermentation period, the sample fermented with P1-2CB-w1 (Enterococcus faecium) exhibited maximum anticoagulation activity and antioxidant activity., Conclusions: The study reveals a novel well-defined starter culture from marine origin intended for seaweed fermentation for recovery of bioactive molecules., Significance and Impact of the Study: The study provides information for the enhancement of bioactive molecules in an eco-friendly manner and also paves a way towards the development of wide range of seaweed functional foods., (© 2012 The Society for Applied Microbiology.)

Aims: Apples and apple products are the most notably commodities contaminated with patulin (PAT), which cause detrimental effects on human health and economic problems. The primary objective of this study was to investigate the removal of PAT contamination from apple juice using 10 different inactivated lactic acid bacteria (LAB) strains., Methods and Results: Significant quantities of PAT ranging from 47 to 80% were bound to all tested bacterial strains, whereas Lactobacillus rhamnosus 6224 and Enterococcus faecium 21605 caused a decrease of PAT by 80·4 and 64·5%, respectively. The results showed that the binding of PAT depends on the initial concentration of toxin and the adsorption temperature, also the differences in biomass existed among the 10 bacterial strains. IR analysis was performed to identify potential functional groups and the possible binding sites related to PAT adsorption., Conclusions: The removal of PAT was observed to be strain specific. The results indicated that the biosorption process did not affect the quality of juice. FTIR analysis showed that the cell wall plays a key role in PAT adsorption., Significance and Impact of the Study: Our results proof that inactivated LAB have the potential as a novel and promising adsorbent to bind PAT effectively., (© 2012 The Authors. Journal of Applied Microbiology © 2012 The Society for Applied Microbiology.)

Aim: To screen five strains of lactic acid bacteria (LAB) isolated from rye sourdoughs for the potential production of antimicrobial substances., Methods and Results: Lactobacillus sakei KTU05-06, Pediococcus acidilactici KTU05-7, Pediococcus pentosaceus KTU05-8, KTU05-9 and KTU05-10 isolated from rye sourdoughs were investigated for the production of bacteriocin-like inhibitory substances (BLIS). The supernatants of analysed LAB inhibited growth of up to 15 out of 25 indicator bacteria strains as well as up to 25 out of 56 LAB strains isolated from rye sourdoughs. Moreover, these five LAB were active against ropes-producing Bacillus subtilis and the main bread mould spoilage causing fungi -Aspergillus, Fusarium, Mucor and Penicillium. Lactobacillus sakei KTU05-6 demonstrated the best antibacterial properties and is resistant towards heat treatment even at 100°C for 60 min., Conclusions: The use of LAB-producing antibacterial substances may be a good choice as a co-starter culture to ensure the stability of sourdoughs and to avoid the bacterial and fungi spoilage of the end product., Significance and Impact of the Study: The antimicrobial compounds designated as sakacin KTU05-6, pediocin KTU05-8 KTU05-9, KTU05-10 and AcKTU05-67 were not identical to any other known BLIS, and this finding leads up to the assumption that they might be the novel., (© 2012 The Authors. Journal of Applied Microbiology © 2012 The Society for Applied Microbiology.)

Celiac disease (CD) is an inflammatory disease of the upper small intestine in genetically predisposed individuals caused by glutamine- and proline-rich peptides from cereal storage proteins (gluten) with a minimal length of nine amino acids. Such peptides are insufficiently degraded by gastrointestinal enzymes; they permeate the lymphatic tissue, are bound to celiac-specific, antigen-presenting cells, and stimulate intestinal T-cells. The typical clinical pattern is a flat small intestinal mucosa and malabsorption. Currently, the only therapy is a strict, lifelong gluten-free diet. Recent research has shown that gluten and gluten peptides can be degraded by prolyl endopeptidases from different sources. These peptidases can either be used to produce gluten-free foods from gluten-containing raw materials, or they have been suggested as an oral therapy for CD, in which dietary gluten is hydrolyzed by coingested peptidases already in the stomach, thus preventing CD-specific immune reactions in the small intestine. This would be an alternative for CD patients to the gluten-free diet. Furthermore, microbial transglutaminase could be used to detoxify gluten either by selectively modifying glutamine residues of intact gluten by transamidation with lysine methyl ester or by crosslinking gluten peptides in beverages via isopeptide bonds so that they can be removed by filtration.

Practical removal of radioactivity from polluted soil in Fukushima, Japan was done using a photosynthetic bacterium, Rhodobacter sphaeroides SSI, immobilized in alginate beads. The beads were put in a mesh bag and soaked in which soil was suspended (5 kg of soil/10 L of tap water). The radioactivity of the broth decreased by 31% after 15 d of aerobic treatment. When lactic acid bacterial culture broth was added to the suspend broth, about 50% of the radioactivity was transferred to a suspend broth fraction consisting of small particles from the soil after 3 d of fermentation and 20 s of sedimentation. The results suggest that organic matter in the soil was decomposed by anaerobic digestion and lactic acid fermentation simultaneously, and was then transferred into the liquid as small particles. With combined treatment by anaerobic digestion and lactic acid fermentation for 5 d and immobilized bead aerobic treatment for an additional 19 d, the radioactivity of suspend broth decreased by 66%. The radioactivity of the original soil (10.56 µSv/h) ultimately decreased by 67% (3.52 µSv/h) after the combined treatment.

Although most vitamins are present in a variety of foods, human vitamin deficiencies still occur in many countries, mainly because of malnutrition not only as a result of insufficient food intake but also because of unbalanced diets. Even though most lactic acid bacteria (LAB) are auxotrophic for several vitamins, it is now known that certain strains have the capability to synthesize water-soluble vitamins such as those included in the B-group (folates, riboflavin and vitamin B(12) amongst others). This review article will show the current knowledge of vitamin biosynthesis by LAB and show how the proper selection of starter cultures and probiotic strains could be useful in preventing clinical and subclinical vitamin deficiencies. Here, several examples will be presented where vitamin-producing LAB led to the elaboration of novel fermented foods with increased and bioavailable vitamins. In addition, the use of genetic engineering strategies to increase vitamin production or to create novel vitamin-producing strains will also be discussed. This review will show that the use of vitamin-producing LAB could be a cost-effective alternative to current vitamin fortification programmes and be useful in the elaboration of novel vitamin-enriched products., (© 2011 The Authors. Journal of Applied Microbiology © 2011 The Society for Applied Microbiology.)

Aims: We have developed a PCR-based assay using custom designed panel of primers which allows rapid detection of class IIa bacteriocin-coding genes. To demonstrate the applicability of the developed assay, the method was applied on 40 metagenomic DNA preparations isolated from native microbiota of Polish artisanal cheeses produced in the Tatra Mountains., Methods and Results: The developed assay was designed on the basis of a large scale alignment of class IIa bacteriocin-coding genes. A panel of seven primer pairs with confirmed ability to detect class IIa bacteriocin-coding sequences was obtained. The following study has revealed a superb bacteriocinogenic potential of all forty analysed cheese samples., Conclusions: The majority of obtained sequences were lactic acid bacteria (LAB) related, although some sequences showed significant similarity to bacteriocin-coding sequences present in non-LAB bacteriocin producers. The results suggest that several potentially new bacteriocin-coding sequences were found., Significance and Impact of the Study: The developed assay can be extremely helpful in establishing whether isolates from the environment of interest have a potential of synthesizing antilisterial class IIa bacteriocins. Application of the approach may represent a useful tool contributing to ecological studies looking for valuable probiotic, bacteriocinogenic microbiota developing in foods., (© 2011 The Authors. Letters in Applied Microbiology © 2011 The Society for Applied Microbiology.)

We measured the adhesion of candidate probiotic lactic acid bacteria (LAB) to carp intestinal mucus. The percentage of adherent bacteria varied among strains. Four strains, two with high adhesion and two with low adhesion in vitro, were tested for in vivo colonization ability. Carp were fed LAB-containing feed for 12 d, and then unsupplemented feed until day 33, and the numbers and compositions of intestinal LAB were analyzed during the entire period. LAB with lower in vitro adhesion disappeared quickly from the intestine after LAB feeding stopped. LAB with higher in vitro adhesion remained in the intestine 3 weeks after LAB feeding stopped, indicating a strong correlation between mucus adhesion in vitro and colonization ability in vivo. Next we isolated nine candidate probiotic LAB with high in vitro mucus-binding ability. Three of them were fed to carp, and all three were stably maintained in the intestine.

In this study, we attempted to establish a novel method of screening anti-allergic lactic acid bacteria (LAB). We cloned the human histidine decarboxylase (HDC) promoter into the promoter-less pPhi-Yellow-RPL-dest1 vector and established KU812F cells transduced with this vector (pHDCp-Phi-Yellow/KU812F). After adding LAB to these cells, the change in fluorescence intensity was monitored by flow cytometry. After screening, we identified several LAB strains that downregulated HDC promoter activity. Functional analysis of these LAB strains indicated that two LAB strains inhibited histamine release from KU812F cells, indicating that this assay system can be used to screen for anti-allergic LAB in a high-throughput manner.

One hundred and seventeen strains of plant origin lactic acid bacteria were observed to have interleukin (IL)-12 production-inducing activities using mouse peritoneal macrophages. Pediococcus pentosaceus (KKM122) was chosen for its stable and strong IL-12 production-inducing activity. There was no significant difference in IL-12 activity induced by the KKM122 strain grown in culture conditions of 0% and 6% NaCl. The cell wall components of cells grown in 6% salt condition, however, significantly induced lower IL-12 production as compared with those of cells grown in 0% salt condition. Cell wall components enhanced IL-12 activity by removing cytoplasmic components when KKM122 strain was cultured in 0% salt condition. The immunoenhancing factor was mainly present in the cell wall components. IL-12 production-inducing activities were dependent on both the amount of bacterial cytoplasmic components and the structure of the cell wall components under the NaCl concentration in the culture medium.

Aim: The ability of Lactobacillus casei and Lactobacillus paracasei to modify the azo dye, tartrazine, was recently documented as the result of the investigation on red coloured spoilage in acidified cucumbers. Fourteen other lactic acid bacteria (LAB) were screened for their capability to modify the food colouring tartrazine and other azo dyes of relevance for the textile industry., Methods and Results: Most LAB modified tartrazine under anaerobic conditions, but not under aerobic conditions in modified chemically defined media. Microbial growth was not affected by the presence of the azo dyes in the culture medium. The product of the tartrazine modification by LAB was identified as a molecule 111 daltons larger than its precursor by liquid chromatography-mass spectrometry. This product had a purple colour under aerobic conditions and was colourless under anaerobic conditions. It absorbed light at 361 and 553 nm., Conclusion: LAB are capable of anabolizing azo dyes only under anaerobic conditions. IMPACT AND SIGNIFICANCE OF THE STUDY: Although micro-organisms capable of reducing the azo bond on multiple dyes have been known for decades, this is the first report of anabolism of azo dyes by food related micro-organisms, such as LAB.

Eighteen lactic acid bacteria (LAB) strains, isolated from coffee pulp silages were characterized according to both growth and gallic acid (GA) consumption. Prussian blue method was adapted to 96-well microplates to quantify GA in LAB microcultures. Normalized data of growth and GA consumption were used to characterize strains into four phenotypes. A number of 5 LAB strains showed more than 60% of tolerance to GA at 2 g/l; whereas at 10 g/l GA growth inhibition was detected to a different extent depending on each strain, although GA consumption was observed in seven studied strains (>60%). Lactobacillus plantarum L-08 was selected for further studies based on its capacity to degrade GA at 10 g/l (97%). MRS broth and GA concentrations were varied to study the effect on growth of LAB. Cell density and growth rate were optimized by response surface methodology and kinetic analysis. Maximum growth was attained after 7.5 h of cultivation, with a dilution factor of 1-1/2 and a GA concentration between 0.625 and 2.5 g/l. Results indicated that the main factor affecting LAB growth was GA concentration. The main contribution of this study was to propose a novel adaptation of a methodology to characterize and select LAB strains with detoxifying potential of simple phenolics based on GA consumption and tolerance. In addition, the methodology presented in this study integrated the well-known RSM with an experimental design based on successive dilutions.

Aims: To identify novel actors responsible for the marked adaptation of the Oenococcus oeni species to its environment., Methods and Results: Genomic surveillance of the available genome sequences from O. oeni indicated the presence of a small ORF, encoding a protein named Dps(A). The cloned gene complemented the dps(-) mutant of Escherichia coli and conferred resistance to hydrogen peroxide, wine, and metals. The dps(A) gene was flanked by IS-related elements. The entire region was characterized by an anomalously high GC content compared to those reported for oenococcal genomes. The dps(A) gene was present in 15 of the 38 tested isolates. Positive strains originated from different geographical areas and sources. No change in tolerance to wine or to oxidative stress was observed between O. oeni strains harbouring dps(A) and those not harbouring this gene., Conclusions: Some O. oeni have acquired a functional homologue to the dps gene from E. coli as part of a mobile element., Significance and Impact of the Study: Dps(A) probably increases the bacterial fitness in response to environmental challenges. However, the physiological condition under which it adds a selective advantage to O. oeni during winemaking remains to be found.

Aims: To evaluate the role of the peptidase activities from sourdough lactic acid bacteria (LAB) in the degradation of alpha-gliadin fragments., Methods and Results: Different proline-containing substrates were hydrolysed by LAB indicating pro-specific peptidase activities. Lactobacillus plantarum CRL 775 and Pediococcus pentosaceus CRL 792 displayed the highest tri- and di-peptidase activities, respectively. Lactobacillus plantarum strains hydrolysed more than 60%alpha-gliadin fragments corresponding to the 31-43 and 62-75 amino acids in the protein after 2 h. None of the LAB strains alone could hydrolyse 57-89 alpha-gliadin peptide; however, the combination of L. plantarum CRL 775 and P. pentosaceus CRL 792 led to hydrolysis (57%) of this peptide in 8 h., Conclusions: The capacity of LAB strains to degrade alpha-gliadin fragments was not correlated to individual peptidase activities. Several strains separately degraded the 31-43 and 62-75 alpha-gliadin fragments, while the 57-89 peptide degradation was associated with the combination of peptidase profiles from pooled LAB strains. This is the first report on the peptide hydrolase system of sourdough pediococci and its ability to reduce alpha-gliadin fragments., Significance and Impact of the Study: This study contributes to a better knowledge of sourdough LAB proteolytic system and its role in the degradation of proline-rich alpha-gliadin peptides involved in celiac disease.

Aims: To investigate microbial diversity and population dynamics of spoilage-sensitive modified-atmosphere-packaged (MAP) artisan-type cooked ham in relation to storage temperature., Methods and Results: Modified-atmosphere-packaged cooked ham samples were stored at different temperatures (4, 7, 12 and 26 degrees C). Traditional methods were combined with polymerase chain reaction (PCR)-based techniques, i.e. a culture-dependent, repetitive DNA sequence-based method (rep-PCR) and a culture-independent approach (PCR-denaturing gradient gel electrophoresis of 16S rRNA gene fragments; PCR-DGGE). rep-PCR on DNA extracted from MRS isolates indicated that Leuconostoc carnosum and Enterococcus faecalis prevailed at all temperatures, with the latter becoming more important above 7 degrees C. PCR-DGGE indicated the additional presence of Carnobacterium divergens and Brochothrix thermosphacta at all temperatures. Discriminant analysis related variation within the Leuc. carnosum cluster to the storage temperature. High performance liquid chromatography revealed that lactic acid was the main metabolite because of glucose consumption., Conclusions: Leuconostoc carnosum, C. divergens, E. faecalis and Br. thermosphacta are the main spoilage bacteria of artisan-type MAP cooked ham. Their population dynamics are affected by storage temperature., Significance and Impact of the Study: Temperature can condition the development of spoilage in artisan-type MAP cooked ham, acting at both species and biotype level.

Aims: To investigate antifungal activity produced by lactic acid bacteria (LAB) isolated from malted cereals and to determine if such LAB have the capacity to prevent fungal growth in a particular food model system., Methods and Results: The effect of pH, temperature and carbon source on production of antifungal activity by four LAB was determined. Pediococcus pentosaceus was used to conduct a trial to determine if it is feasible to eliminate Penicillium expansum, the mould responsible for apple rot, using an apple model. Penicillium expansum was incapable of growth during the trial on apple-based agar plates inoculated with the antifungal-producing culture, whereas the mould did grow on apple plates inoculated with an LAB possessing no antifungal activity., Conclusion: Partial characterization of the antifungal compounds indicates that their activity is likely to be because of production of antifungal peptides. The trial conducted showed that the antifungal culture has the ability to prevent growth of the mould involved in apple spoilage, using apples as a model., Significance and Impact of the Study: The ability of an LAB to prevent growth of Pen. expansum using the apple model suggests that these antifungal LAB have potential applications in the food industry to prevent fungal spoilage of food.

Aims: To verify to what degree reducing capacity is a characterizing parameter of a species, and of the strains themselves within a given species, of lactic acid bacteria., Methods and Results: Eighty-eight strains belonging to 10 species of lactic acid bacteria (LAB) isolated from traditional Italian cheeses were studied for their reduction activity: Enterococcus faecalis, Enterococcus faecium, Enterococcus durans, Streptococcus thermophilus, Lactococcus lactis ssp. lactis, Lactobacillus paracasei ssp. paracasei, Lactobacillus plantarum, Lactobacillus delbrueckii ssp. bulgaricus, Lactobacillus helveticus and Pediococcus pentosaceus. It was observed that the lactococci reached minimum redox potential before the lactobacilli. The reduction rate of Enterococcus spp. and L. lactis ssp. lactis was higher than that of the streptococci and Lactobacillus spp. All the P. pentosaceus strains had poor reduction activity compared with the other species., Conclusions: The evolution of the redox potential in milk over a time span of 24 h has been found to be a parameter that characterizes a species: the different courses corresponding to the species in question are clearly evident, and interesting differences can also be noted within the same species., Significance and Impact of the Study: The reduction aptitude of strains might be used to select and adapt appropriate strains for use as starters for dairy products.

Aims: To study the occurrence of histidine, tyrosine and ornithine decarboxylase activity in lactic acid bacteria (LAB) isolated from natural ciders and to examine their potential to produce detrimental levels of biogenic amines., Methods and Results: The presence of biogenic amines in a decarboxylase synthetic broth and in cider was determined by reversed-phase high-performance liquid chromatography (RP-HPLC). Among the 54 LAB strains tested, six (five lactobacilli and one oenococci) were biogenic amine producers in both media. Histamine and tyramine were the amines formed by the LAB strains investigated. Lactobacillus diolivorans were the most intensive histamine producers. This species together with Lactobacillus collinoides and Oenococcus oeni also seemed to produce tyramine. No ability to form histamine, tyramine or putrescine by Pediococus parvulus was observed, although it is a known biogenic amine producer in wines and beers., Conclusions: This study demonstrated that LAB microbiota growing in ciders had the ability to produce biogenic amines, particularly histamine and tyramine, and suggests that this capability might be strain-dependent rather than being related to a particular bacterial species., Significance and Impact of the Study: Production of biogenic amines by food micro-organisms has continued to be the focus of intensive study because of their potential toxicity. The main goal was to identify the microbial species capable of producing these compounds in order to control their presence and metabolic activity in foods.

Aims: Enterococcus faecalis produces a cationic and circular enterocin, AS-48, of 7149 Da, the genetic determinants of which are located within the pMB2 plasmid. We have compared enterocin AS-48 production by different enterococci species with that of other 'safe' lactic acid bacteris (LAB) (GRAS status) and looked into the subsequent application of this enterocin in food production., Methods and Results: In an effort to exploit this system for the heterologous expression of enterocin AS-48, a number of vectors containing the as-48 cluster were constructed and used to transform several LAB strains (genera Enterococcus, Lactococcus and Lactobacillus), Conclusion: Heterologous production of enterocin AS-48 failed when bacteria other than those belonging to the genus Enterococcus were used as hosts, although expression of a partial level of resistance against AS-48 were always detected, ruling out the possibility of a lack of recognition of the enterococcal promoters., Significance and Impact of the Study: Our results reveal the special capacity of species from the genus Enterococcus to produce AS-48, an enterocin that requires a post-transcriptional modification to generate a circular peptide with a wide range of inhibitory activity against pathogenic and spoilage bacteria. Preliminary experiments in foodstuffs using nonvirulent enterococci with interesting functional properties reveal the possibility of a biotechnological application of these transformants.

Aims: The ability of lactic acid bacteria (LAB) to metabolize certain phenolic precursors to vanillin was investigated., Methods and Results: Gas chromatography-mass spectrometry (GC-MS) or HPLC was used to evaluate the biosynthesis of vanillin from simple phenolic precursors. LAB were not able to form vanillin from eugenol, isoeugenol or vanillic acid. However Oenococcus oeni or Lactobacillus sp. could convert ferulic acid to vanillin, but in low yield. Only Lactobacillus sp. or Pediococcus sp. strains were able to produce significant quantities of 4-vinylguaiacol from ferulic acid. Moreover, LAB reduced vanillin to the corresponding vanillyl alcohol., Conclusions: The transformation of phenolic compounds tested by LAB could not explain the concentrations of vanillin observed during LAB growth in contact with wood., Significance and Impact of the Study: Important details of the role of LAB in the conversion of phenolic compounds to vanillin have been elucidated. These findings contribute to the understanding of malolactic fermentation in the production of aroma compounds.

Aims: The aim of this study was to isolate and identify bacteriocin-producing lactic acid bacteria (LAB) issued from Mongolian airag (traditional fermented mare's milk), and to purify and characterize bacteriocins produced by these LAB., Methods and Results: Identification of the bacteria (Enterococcus durans) was carried out on the basis of its morphological, biochemical characteristics and carbohydrate fermentation profile and by API50CH kit and 16S rDNA analyses. The pH-neutral cell-free supernatant of this bacterium inhibited the growth of several Lactobacillus spp. and food-borne pathogens including Escherichia coli, Staphylococcus aureus and Listeria innocua. The antimicrobial agent (enterocin A5-11) was heat stable and was not sensitive to acid and alkaline conditions (pH 2-10), but was sensitive to several proteolytic enzymes. Its inhibitory activity was completely eliminated after treatment with proteinase K and alpha-chymotrypsin. The activity was however not completely inactivated by other proteases including trypsin and pepsin. Three-step purification procedure with high recovery yields was developed to separate two bacteriocins. The applied procedure allowed the recovery of 16% and 64% of enterocins A5-11A and A5-11B, respectively, present in the culture supernatant with purity higher than 99%. SDS-PAGE analyses revealed that enterocin A5-11 has a molecular mass of 5000 Da and mass spectrometry analyses demonstrates molecular masses of 5206 and 5218 Da for fractions A and B, respectively. Amino acid analyses of both enterocins indicated significant quantitative difference in their contents in threonine, alanine, isoleucine and leucine. Their N-termini were blocked hampering straightforward Edman degradation., Conclusions: Bacteriocins A5-11A and B from Ent. durans belong to the class II of bacteriocins., Significance and Impact of the Study: Judging from molecular masses, amino acid composition and spectrum of activities, bacteriocins A5-11A and B from Ent. durans show high degree of similarity with enterocins L50A and L50B isolated from Enterococcus faecium (Cintas et al. 1998, 2000) and with enterocin I produced by Ent. faecium 6T1a, a strain originally isolated from a Spanish-style green olive fermentation (Floriano et al. 1998).

Aims: Acetaldehyde is the major carbonyl compound formed during winemaking and has implications for sensory and colour qualities of wines as well as for the use of the wine preservative SO(2). The current work investigated the degradation of acetaldehyde and SO(2)-bound acetaldehyde by two commercial Oenococcus oeni starters in white wine., Methods and Results: Wines were produced by alcoholic fermentation with commercial yeast and adjusted to pH 3.3 and 3.6. While acetaldehyde was degraded rapidly and concurrently with malic acid at both pH values, SO(2)-bound acetaldehyde caused sluggish bacterial growth. Strain differences were small., Conclusions: Efficient degradation of acetaldehyde can be achieved by commercial starters of O. oeni. According to the results, the degradation of acetaldehyde could not be separated from malolactic conversion by oenococci. While this may be desirable in white winemaking, it may be necessary to delay malolactic fermentation (MLF) in order to allow for colour development in red wines. SO(2)-bound acetaldehyde itself maybe responsible for the sluggish or stuck MLF, and thus bound SO(2) should be considered next to free SO(2) in order to evaluate malolactic fermentability., Significance and Impact of the Study: The current study provides new results regarding the metabolism of acetaldehyde and SO(2)-bound acetaldehyde during the MLF in white wine. The information is of significance to the wine industry and may contribute to reducing the concentration of wine preservative SO(2).

Aims: To study arginine degradation and carcinogenic ethyl carbamate precursor citrulline formation during and after malolactic fermentation (MLF)., Methods and Results: MLF was induced in white wine with two commercial Oenococcus oeni strains under different winemaking conditions regarding the type of alcoholic fermentation (spontaneous, induced) and the lees management (racked, on lees). Arginine degradation and citrulline formation did not occur during malic acid degradation in any treatment. In five of the six treatments in which arginine degradation took place, it occurred 3 weeks after malic acid depletion and significant amounts of citrulline were formed. Presence of yeast lees in wines led to increased citrulline formation., Conclusions: This study suggests that arginine metabolism is inhibited in oenococci at low pH values (< 3.5) and that in the postalcoholic fermentation phase, citrulline formation from arginine degradation can be avoided if MLF is induced by pure cultures of O. oeni with inhibition of the bacterial biomass after malic acid depletion. Residual yeast lees in the wine have been identified as a significant risk factor for increased citrulline formation., Significance and Impact of the Study: Conclusions drawn from this study allow reducing the risk of carcinogenic ethyl carbamate formation from citrulline excretion by wine lactic acid bacteria.

Carnocin KZ213 is an antilisterial bacteriocin produced by Carnobacterium piscicola 213. The effects of pH and temperature were studied during batch fermentation in MRS* medium (modified MRS without ammonium citrate or sodium acetate). The optimal pH for growth is between 6 and 7. The maximum bacteriocin productivity in the supernatant occurs at pH 7. Operating at controlled pH increases the volumetric activity of the free bacteriocin by 8- to 16-fold, compared with uncontrolled pH. No bacteriocin production is observed below pH 6.5. Temperature has a dramatic effect on carnocin KZ213 production. Growth is optimal at 25 degrees C and 30 degrees C, although no bacteriocin production is detected at 30 degrees C. Also, bacteriocin production is observed at 25 degrees C in MRS*, but not in complex APT broth, where growth is optimal. The presence of glucose as a carbon and/or energy source is important for carnocin KZ213 synthesis. Hence, bacteriocin synthesis is regulated by temperature, carbon source and medium composition. Quantification studies of bacteriocin adsorbed onto producer cells show that the majority of the carnocin KZ213 secreted is adsorbed onto the producer cells during growth. Only 15% of the total bacteriocin produced is detected in the cell-free supernatant at the end of growth.

Aims: To evaluate the effect of high-pressure processing (HPP) on Listeria monocytogenes, microbial and chemical changes and shelf-life in chilled cold-smoked salmon (CSS)., Methods and Results: First, challenge tests with L. monocytogenes were carried out using HPP of the product at 0.1 (control), 150, 200 and 250 MPa. Secondly, storage trials with the naturally contaminated product and HPP at 0.1 (control) and 200 MPa were realized. Shelf-life, microbial changes and chemical changes were determined and existing predictive models and multiple compound quality indices evaluated. HPP with 250 MPa did not inactivate L. monocytogenes but significant lag phases of 17 and 10 days were observed at ca 5 and 10 degrees C, respectively. HPP with 200 MPa had a marked effect on both colour and texture of CSS., Conclusions: High-pressure processing was unable to prevent growth of L. monocytogenes or spoilage of chilled CSS. Existing mathematical models allowed growth rates of L. monocytogenes and shelf-life of samples without high-pressure treatments to be predicted., Significance and Impact of the Study: High-pressure processing seems more appropriate for new types of salmon products than for a classical product like CSS where consumers expect specific quality attributes.

Aims: To study the importance of bacteriocin production for the antilisterial effect of a bacteriocinogenic Carnobacterium piscicola strain A9b on growth of Listeria monocytogenes in broth and cold-smoked salmon systems., Methods and Results: Acriflavin treatment of strain A9b resulted in loss of bacteriocin production and of immunity to carnobacteriocin B2. Two plasmids present in the wild-type were lost in the variant that was also more sensitive to bavaricin and leucocin A than the wild-type indicating cross-resistance to class IIa bacteriocins. The growth rate of the bac- mutant was higher than that of the wild-type at 5 and 37 degrees C but not at 25 or 30 degrees C. In salmon juice the maximum cell density of L. monocytogenes was suppressed 3 and 6 log by co-culture with C. piscicola A9b bac- and bac+, respectively, as compared with the control. Sterile filtered cultures of C. piscicola A9b bac- caused a limited suppression of the maximum cell density of L. monocytogenes similar to that observed when sterile buffer was added in equal amounts. Semi-purified carnobacteriocin B2 caused a 3.5 log decline in viable cell count after 6 day of incubation in cold-smoked salmon juice at 5 degrees C. High resistance level to carnobacteriocin B2 was observed for L. monocytogenes cells exposed to semi-purified and in situ produced carnobacteriocin B2., Conclusions: The presence of bacteriocin production in C. piscicola enhances its inhibition of L. monocytogenes., Significance and Impact of the Study: Due to the emergence of resistance, a bacteriocin negative lactic acid bacteria may be more suited for practical use as a bioprotective agent against L. monocytogenes in ready-to-eat foods.

Aims: The objective of this study was to determine the technological behaviour (implantation and biogenic amines production) of Carnobacterium divergens V41, an anti-Listeria bacteriocin producer (divercin V41), after inoculation in cold smoked salmon (CSS)., Methods and Results: Implantation of the strain was followed by multiplex-PCR during 27 days of storage at 4 degrees C, and biogenic amines were quantified by HPLC. It was found that the strain was able to develop quite well in CSS among lactic wild flora. Divercin V41 (400 AU ml-1) was produced in CSS, and the biogenic amine content was not modified by inoculation of the bacteria., Conclusions: Carnobacterium divergens V41 is a safe, interesting, bioprotective agent., Significance and Impact of the Study: This strain could potentially be used for efficient prevention of L. monocytogenes growth in CSS.

Aim: The study was carried out to assess the agreement of API 50 CH fermentation data of food lactobacilli with their RAPD profiles to determine whether the system could be used alone as a reliable taxonomic tool for this genus., Methods and Results: API 50 CH, RAPD and DNA:DNA reassociation data for 42 lactobacilli from tef and kocho were compared with 30 type strains. Discrepancies were observed between the three methods in assigning strains of Lactobacillus plantarum, Lact. fermentum, Weissella minor and Lact. gallinarum, and Lact. fermentum, Lact. amylophilus, Lact. casei subsp. pseudoplantarum and Lact. rhamnosus. DNA reassociation data agreed well with RAPD results., Conclusions: API 50 CH profiles should be complemented with molecular genetic results for effective identification in Lactobacillus., Significance and Impact of the Study: The study suggested less dependability of metabolic data alone as an identification tool.

Acetaldehyde is a volatile flavor compound present in many fermented foods and is important in the production of red and white wines. Nine strains of the genera Lactobacillus and Oenococcus were able to metabolize acetaldehyde in a resting cell system, whereas two Pediococcus strains were not. Acetic acid and ethanol were produced from its degradation. A Lactobacillus and an Oenococcus were able to degrade SO(2)-bound acetaldehyde, as well. A coincubation of resting cells of Saccharomyces bayanus Première Cuvée and Oenococcus oeni Lo111 showed that strain Lo111 metabolized acetaldehyde produced by the yeast. The ability of malolactic bacteria to degrade free and SO(2)-bound acetaldehyde has implications for sensory and color qualities and the use of SO(2) in wine.

The production of some bacteriocins by lactic acid bacteria is regulated by induction peptides (IPs) that are secreted by a dedicated secretion system. The IP gene cbaX, for carnobacteriocin A production by Carnobacterium piscicola LV17A, and a presumptive IP gene (orf6), associated with the genetic locus for enterocin B production in Enterococcus faecium BFE 900, were fused to the signal peptide of the bacteriocin divergicin A from Carnobacterium divergens LV13 to access the general secretory pathway. The culture supernatants of C. piscicola UAL26 and Lactococcus lactis MG1363 containing either of these constructs were used to induce bacteriocin production by Bac(-) cultures of C. piscicola LV17A or E. faecium CTC492. The cbaX fusion product induced bacteriocin production by Bac(-) C. piscicola LV17A, but the orf6 fusion product did not induce bacteriocin production by E. faecium CTC492. This represents a relatively simple method of confirming the role of presumptive IPs. The transformation of C. piscicola LV17A with the CbaX gene under expression of the P32 promoter from L. lactis resulted in constitutive production of bacteriocin by either the dedicated transport apparatus or the general secretory pathway.

Major commercially available strains for induction of malolactic fermentation in wine were examined for arginine metabolism in a resting cell system at wine pH with the aim of evaluating their ability to excrete and utilize citrulline, a precursor of carcinogenic ethyl carbamate (urethane). All strains tested excreted citrulline from arginine degradation. Citrulline was stored intracellularly during growth in arginine rich medium and was released upon lysis of the cells. All strains were found to degrade citrulline as a sole amino acid and some of them were able to reutilize previously excreted citrulline.

Colicin V is a small, proteinaceous bacterial toxin, produced by many strains of Escherichia coli and other members of the Enterobacteriaceae, that fits the definition of class II bacteriocins of Gram-positive bacteria. Export of colicin V is dependent on specific ABC (ATP-binding cassette) secretion proteins which recognize a double-glycine-type leader peptide on the immature colicin V bacteriocin. Replacement of the colicin V leader peptide by a signal peptide from the signal sequence-dependent bacteriocin divergicin A allowed expression of colicin V in lactic acid bacteria. This system may serve as a model for the heterologous expression of other small bacteriocins active against Gram-negative bacteria and other antibacterial peptides from lactic acid bacteria.