Lactobacillus Sanfranciscensis Research Articles

Development of species-specific PCR primers and polyphasic characterization of Lactobacillus sanfranciscensis isolated from Korean sourdough

Lactobacillus sanfranciscensis is a bacterium used in sourdough that provides desirable properties such as better flavor and texture to the sourdough bread. Here, the intra-species diversity of L. sanfranciscensis strains isolated from Korean sourdough was studied using genotypic (multiplex-RAPD-PCR: multiplex-Randomly Amplified Polymorphic DNA-polymerase chain reaction) and phenotypic (VITEK2 Compact system) analyses. For this, a novel species-specific set of PCR primers was developed to identify L. sanfranciscensis using the recently published genome database. The primers were able to detect L. sanfranciscensis isolated from Korean sourdough with 100% accuracy. Genotyping and phenotyping analyses at the strain level demonstrated that Korean sourdough possesses various biotypes of L. sanfranciscensis strains. These strains were clustered into 5 subtypes (genotyping) or 7 subtypes (phenotyping). In summary, this strategy to construct novel primers reduced the chance of cross amplification and was able to identify the desired strain. The various strains isolated in this study can be used to develop a sourdough starter after the analysis of their fermentation characteristics.

Sourdough microbial community dynamics: An analysis during French organic bread-making processes

Natural sourdoughs are commonly used in bread-making processes, especially for organic bread. Despite its role in bread flavor and dough rise, the stability of the sourdough microbial community during and between bread-making processes is debated. We investigated the dynamics of lactic acid bacteria (LAB) and yeast communities in traditional organic sourdoughs of five French bakeries during the bread-making process and several months apart using classical and molecular microbiology techniques. Sourdoughs were sampled at four steps of the bread-making process with repetition. The analysis of microbial density over 68 sourdough/dough samples revealed that both LAB and yeast counts changed along the bread-making process and between bread-making runs. The species composition was less variable. A total of six LAB and nine yeast species was identified from 520 and 1675 isolates, respectively. The dominant LAB species was Lactobacillus sanfranciscensis, found for all bakeries and each bread-making run. The dominant yeast species changed only once between bread-making processes but differed between bakeries. They mostly belonged to the Kazachstania clade. Overall, this study highlights the change of population density within the bread-making process and between bread-making runs and the relative stability of the sourdough species community during bread-making process.

Isolation and molecular identification of lactic acid bacteria and Bifidobacterium spp. from faeces of the blue-fronted Amazon parrot in Brazil.

In Brazil, the blue-fronted Amazon parrot (Amazona aestiva) is a common pet. The faecal microbiota of these birds include a wide variety of bacterial species, the majority of which belong to the Gram-positive lactic acid bacteria (LAB) clade. The aim of this study was to investigate differences in the diversity and abundance of LAB and Bifidobacterium spp. in the cloacae between wild and captive birds and to select, identify and characterise LAB for consideration as a parrot probiotic. Cloacal swabs were collected from 26 wild and 26 captive birds. Bacterial DNA was extracted, and the 16S rRNA genes were amplified. The numbers of PCR-positive Enterococcus, Pediococcus, and Lactobacillus species isolated from wild and captive birds were significantly different (P<0.05). Enterococcus was the most frequently isolated genus, followed by Pediococcus, Lactobacillus, Lactococcus and Bifidobacterium. Enterococcus faecium, Pediococcus pentosaceus, Lactococcus lactis, Lactobacillus coryniformis, Lactobacillus sanfranciscensis and Bifidobacterium bifidum were the most frequently isolated species from all birds. This study increases our understanding of the faecal microbiota, and may help to improve the nutrition and habitat management of captive and wild parrots. The bacterial population identified in the faecal microbiota of clinically healthy wild and captive parrots can serve as a database to analyse variations in the gut microbiota of pathogen-infected parrots and to develop probiotics specific to these genera.

Antifungal properties of selected lactic acid bacteria and application in the biological control of ochratoxin A producing fungi during cocoa fermentation

Thirteen Lactic acid bacteria strains isolated from fermenting cocoa and seven reference strains were used in order to assess their antifungal properties towards three ochratoxin A (OTA) producing fungi (Aspergillus carbonarius, Aspergillus niger and Aspergillus ochraceus). Furthermore, two of the isolates strains (A19 and A21) identified as belonging to the genus of Pediococcus as well as Lactobacillus plantarum B4496, Lactobacillus brevis 207 and Lactobacillus sanfranciscensis BB12 showed interesting in vitro broad antifungal activities towards the three ochratoxin-producing fungi with inhibition percentages ranging from 15% to 66.7%. Treatment of cell-free supernatant at 100°C affected antifungal activity suggesting that the main compounds responsible for this activity were of proteic nature, and hence could be bacteriocins. Application of isolate A19 in cocoa fermentation as starter inhibited the growth of each of the OTA-producing species. At the end of fermentation in boxes inoculated with A19, A. niger was not detectable while A. carbonarius concentration was found to be 2 Log CFU/g of wet beans. The assessment of the ochratoxin produced during fermentation of cocoa inoculated with A. carbonarius indicated that the use of isolate A19 as starter could reduce their level of growth so as to have only a toxin production of 0.0012 ± 0.0005 μg/kg after 40 days of storage, while this was 2.45 ± 0.35 μg/kg of fermented and dried cocoa beans in the absence of A19. This work is a contribution for the application of biological control of OTA-producing fungi during cocoa production.

Increasing ATP conservation in maltose consuming yeast, a challenge for industrial organic acid production in non-aerated reactors

Anaerobic fermentation processes are economically attractive for industry, as costs for aeration and stirring are greatly reduced. An example of such a process is (bio)ethanol production by Saccharomyces cerevisiae, in which a low but positive ATP gain leads to elevated product yields. However, for other fermentative pathways, such as in an engineered homolactic strain of S. cerevisiae, the net ATP gain for the fermentation of glucose to lactate is null due to the requirement of ATP for product export. Therefore, increasing the conservation of ATP is of major importance for such 'zero-ATP pathways'. One opportunity to increase ATP conservation arises when disaccharides are used as a substrate. In many industrial microorganisms, disaccharides are cleaved by hydrolysis, which results in the dissipation of energy that is available in this cleaving reaction. However, phosphorolytic cleavage could be used to replace the hydrolysis of disaccharides, thereby increasing the ATP yield. In this study, growth of S. cerevisiae on maltose was used as a model. All known native maltose metabolism genes were removed and replaced by a maltose phosphorylase (Lactobacillus sanfranciscensis) and a single overexpressed copy of the native MAL11 maltose transporter. Because maltose phosphorylase cleaves maltose into glucose and β-glucose-1-phosphate, additionally a β-phosphoglucomutase (Lactococcus lactis) was co-expressed in this strain. Anaerobic maltose-limited chemostat cultures showed that replacement of maltose hydrolysis by phosphorolysis increased the biomass yield of the mutant strain by 26% over the wild type strain, demonstrating the potential of phosphorolysis to improve ATP conservation of disaccharide metabolism in industrial microorganisms [1].

Use of sourdough fermentation and mixture of wheat, chickpea, lentil and bean flours for enhancing the nutritional, texture and sensory characteristics of white bread

This study aimed at investigating the addition of legume (chickpea, lentil and bean) flours to wheat flour bread. Type I sourdough containing legumes or wheat–legume flours were prepared and propagated (back slopped) in laboratory, according to traditional protocols that are routinely used for making typical Italian breads. Based on kinetic of acidification and culture-dependent data, the wheat–legume sourdough was further characterized and selected for bread making. As determined by RAPD-PCR and partial sequencing of 16S rDNA gene analyses, lactic acid bacteria in wheat–legume sourdough included Lactobacillus plantarum, Lactobacillus sanfranciscensis, Leuconostoc mesenteroides, Lactobacillus fermentum, Weissella cibaria, Lactobacillus pentosus, Lactobacillus coryneformis, Lactobacillus rossiae, Lactobacillus brevis, Lactobacillus parabuchneri and Lactobacillus paraplantarum. Two breads containing 15% (w/w) of legume (chickpea, lentil and bean) flours were produced using selected wheat–legume sourdough (WLSB) and traditional wheat sourdough (WSB). Compared to wheat yeasted bread (WYB), the level of total free amino acids (FAA) was higher in WSB and WLSB. Phytase and antioxidant activities were the highest in WLSB. Compared to bread WYB, the addition of legume flours decreased the in vitro protein digestibility (IVPD) (WYB versus WSB). However, the dough fermentation with WSLB favored an increase of IVPD. According to the levels of carbohydrates, dietary fibers and resistant starch, WSB and WLSB showed lower values of hydrolysis index (HI) compared to WYB. As showed by texture and image analyses and sensory evaluation of breads, a good acceptability was found for WSB and, especially, WLSB breads.

The genome of the Lactobacillus sanfranciscensis temperate phage EV3

BackgroundBacteriophages infection modulates microbial consortia and transduction is one of the most important mechanism involved in the bacterial evolution. However, phage contamination brings food fermentations to a halt causing economic setbacks. The number of phage genome sequences of lactic acid bacteria especially of lactobacilli is still limited. We analysed the genome of a temperate phage active on Lactobacillus sanfranciscensis, the predominant strain in type I sourdough fermentations.ResultsSequencing of the DNA of EV3 phage revealed a genome of 34,834 bp and a G + C content of 36.45%. Of the 43 open reading frames (ORFs) identified, all but eight shared homology with other phages of lactobacilli. A similar genomic organization and mosaic pattern of identities align EV3 with the closely related Lactobacillus vaginalis ATCC 49540 prophage. Four unknown ORFs that had no homologies in the databases or predicted functions were identified. Notably, EV3 encodes a putative dextranase.ConclusionsEV3 is the first L. sanfranciscensis phage that has been completely sequenced so far.

An integrated technological approach to the selection of lactic acid bacteria of flour origin for sourdough production

Several lactic acid bacteria (LAB) were evaluated in situ for their potential in sourdough fermentation. The strains belonged to Lactobacillus plantarum, Lactobacillus sakei, Lactobacillus sanfranciscensis, Leuconostoc citreum, Leuconostoc mesenteroides, Leuconostoc pseudomesenteroides and Weissella cibaria. LAB were used, in individual inocula, to carry out the fermentation of γ-ray treated (sterile) flour and untreated commercial flour, in order to evaluate their performances both in the absence and presence of the native microbiota of flour. The pH and total titratable acidity (TTA) showed a strong and fast acidification of the experimental sourdough determined by W. cibaria and Ln. citreum strains. All strains were followed during fermentation by plate count. Randomly amplified polymorphic DNA (RAPD)-PCR analysis applied on the colonies isolated from the highest dilution of samples confirmed the dominance of the added strains in all sourdoughs prepared with sterile and non-sterile flour. The analysis of organic acids, performed by high-performance liquid chromatography (HPLC), confirmed that some W. cibaria and Ln. citreum strains showed an optimal fermentation quotient. The volatile organic compound (VOC) composition resulting from the gas chromatography coupled with mass spectrometry (GC/MS) analysis of sourdough headspace recognised 51 chemical compounds including acids, alcohols, aldehydes, esters, ketones, lactones, acetate, alkane, and phenol, most of which are of LAB origin and are relevant for the final bread. After baking, the breads were evaluated for the height of the central slices, colour of crust and crumb, hardness and number and distribution of alveolus. The combination of these results indicated that strains Ln. citreum PON10079 and PON10080 and W. cibaria PON10030 and PON10032 are suitable cultures to use in industrial production.

1,8-Bis(dimethylamino)naphthalene/9-aminoacridine: A new binary matrix for lipid fingerprinting of intact bacteria by matrix assisted laser desorption ionization mass spectrometry

The effectiveness of a novel binary matrix composed of 1,8-bis(dimethylamino)naphthalene (DMAN; proton sponge) and 9-aminoacridine (9AA) for the direct lipid analysis of whole bacterial cells by matrix assisted laser desorption ionization mass spectrometry (MALDI MS) is demonstrated. Deprotonated analyte signals nearly free of matrix-related ions were observed in negative ion mode. The effect of the most important factors (laser energy, pulse voltage, DMAN/9AA ratio, analyte/matrix ratio) was investigated using a Box–Behnken response surface design followed by multi-response optimization in order to simultaneously maximize signal-to-noise (S/N) ratio and resolution. The chemical surface composition of single or mixed matrices was explored by X-ray photoelectron spectroscopy (XPS). Moreover, XPS imaging was used to map the spatial distribution of a model phospholipid in single or binary matrices.The DMAN/9AA binary matrix was then successfully applied to the analysis of intact Gram positive (Lactobacillus sanfranciscensis) or Gram negative (Escherichia coli) microorganisms. About fifty major membrane components (free fatty acids, mono-, di- and tri-glycerides, phospholipids, glycolipids and cardiolipins) were quickly and easily detected over a mass range spanning from ca. 200 to ca. 1600m/z. Moreover, mass spectra with improved S/N ratio (compared to single matrices), reduced chemical noise and no formation of matrix-clusters were invariably obtained demonstrating the potential of this binary matrix to improve sensitivity.

In vitro assessment of functional properties of lactic acid bacteria isolated from faecal microbiota of healthy dogs for potential use as probiotics

Lactic acid bacteria were isolated and identified in the faeces of Chinese Crested and Yorkshire terrier pups and their probiotic features were investigated in vitro. Thirty seven isolates were identified as Lactobacillus or Enterococcus. Out of these isolates, 31 were lactic acid bacteria (LAB) and belonged to the species Lactobacillus reuteri (16/37; 43.3%), Lactobacillus animalis (7/37; 18.9%), Lactobacillus acidophilus (3/37; 8.1%), Lactobacillus sanfranciscensis (2/37; 5.4%), Lactobacillus murinus (2/37; 5.4%), and Lactobacillus paraplantarum (1/37; 2.7%), while six other LAB isolates were Enterococcus spp. (6/37; 16.2%). Strains were tested for resistance to gastric acidity (pH 2.5 for 3 h) and bile salts (0.3% ox gall), cell surface hydrophobicity by microbial adhesion to solvents, antagonism against pathogenic bacteria (Staphylococcus aureus, Enterococcus faecalis, Bacillus cereus, Pseudomonas aeruginosa, Escherichia coli, Salmonella enterica serovar Typhimurium and Listeria monocytogenes), production of hydrogen peroxide, and antibiotic susceptibility. Thirty four strains were highly resistant to acidic conditions with slight (18 strains) to moderate (16 strains) growth inhibition by bile salts. Seven isolates had highly hydrophobic cellular surfaces and 28 strains exhibited strong antagonism against the bacterial pathogens tested, although 8 isolates tested against Leptospira interrogans had no effect on pathogen growth. All isolates produced low rates of hydrogen peroxide. Based on these results, two Lactobacillus strains showed promising probiotic-related features and merit investigation as probiotics for dogs.

Microbial ecology of sourdough fermentations: Diverse or uniform?

Sourdough is a specific and stressful ecosystem inhabited by yeasts and lactic acid bacteria (LAB), mainly heterofermentative lactobacilli. On the basis of their inocula, three types of sourdough fermentation processes can be distinguished, namely backslopped ones, those initiated with starter cultures, and those initiated with a starter culture followed by backslopping. Typical sourdough LAB species are Lactobacillus fermentum, Lactobacillus paralimentarius, Lactobacillus plantarum, and Lactobacillus sanfranciscensis. Typical sourdough yeast species are Candida humilis, Kazachstania exigua, and Saccharomyces cerevisiae. Whereas region specificity is claimed in the case of artisan backslopped sourdoughs, no clear-cut relationship between a typical sourdough and its associated microbiota can be found, as this is dependent on the sampling, isolation, and identification procedures. Both simple and very complex consortia may occur. Moreover, a series of intrinsic and extrinsic factors may influence the composition of the sourdough microbiota. For instance, an influence of the flour (type, quality status, etc.) and the process parameters (temperature, pH, dough yield, backslopping practices, etc.) occurs. In this way, the presence of Lb. sanfranciscensis during sourdough fermentation depends on specific environmental and technological factors. Also, Triticum durum seems to select for obligately heterofermentative LAB species. Finally, there are indications that the sourdough LAB are of intestinal origin.

Antifungal Lipids Produced by Lactobacilli and Their Structural Identification by Normal Phase LC/Atmospheric Pressure Photoionization–MS/MS

Lactobacillus hammesii converts linoleic acid into an antifungal hydroxy fatty acid. High speed counter-current chromatography (HSCCC) using a hexane/ethyl acetate/methanol/water solvent system [3.5:1.5:3:2 (v/v/v/v)] allowed isolation of the antifungal hydroxy fatty acid. A method was developed for characterization of antifungal hydroxy fatty acids using normal phase liquid chromatography combined with atmospheric pressure photoionization-tandem mass spectrometry (LC/APPI-MS/MS). The position of unsaturations and hydroxyl groups was determined directly from crude lipid extracts and their hydroxylated derivatives. The antifungal compounds were identified as a racemic mixture of 10-hydroxy-cis-12-octadecenoic and 10-hydroxy-trans-12-octadecenoic acid. Additionally, HSCCC and LC/APPI-MS/MS methods were used to elucidate the pathway of conversion of linoleic acid by Lactobacillus sanfranciscensis , Lactobacillus plantarum , and L. hammesii to hydroxy fatty acids and conjugated linoleic acid. This study links previously reported 10-hydroxy-12-octadecenoic acid producing Lactobacillus strains to antifungal activities.

The lactic acid bacteria and yeast microbiota of eighteen sourdoughs used for the manufacture of traditional Italian sweet leavened baked goods

The lactic acid bacteria and yeast microbiota of eighteen sourdoughs used for the manufacture of some traditional Italian sweet leavened baked goods were studied through culture-dependent method and pyrosequencing. Flours used for back slopping and sourdoughs were also biochemically characterized. Principal component analysis was applied to explore eventual correlations between process parameters applied during back slopping, some flour nutrients, profile of microbiota, and biochemical characteristics of sourdoughs. The median values of the cell density of lactic acid bacteria and yeasts were 8.05 and 7.03logCFU/g, respectively. As shown by culture-dependent method, Lactobacillus sanfranciscensis was identified in all the sourdoughs, except for Panaredda and Torcolo di San Costanzo. For eleven sourdoughs, all the lactic acid bacteria isolates were allotted to this species. For Buccellato di Lucca, Mbriagotto, Pandoro, and Nadalin sourdoughs, at least 80% of the isolates was allotted to this species. Other lactic acid bacteria isolated with a relatively high frequence were Lactobacillus plantarum and Leuconostoc citreum. Pyrosequencing confirmed and complemented the culture-dependent approach, detecting L. sanfranciscensis also in Panaredda and Torcolo di San Costanzo sourdoughs. Saccharomyces cerevisiae was identified in all the sourdoughs, except for Mbriagotto, Ciambella di Mosto and Pandolce Genovese. These latter sourdoughs harbored strains of Candida humilis, whereas five sourdoughs combined the presence of both yeast species. Positive correlations were found between time of back slopping and cell density and main metabolites of lactic acid bacteria. Percentage of sourdough used as inoculum was mainly correlated with the cell density of yeasts and the concentration of ethanol.This study provided a comprehensive and comparative approach to highlight the dominant microbiota of Italian sourdoughs, which could be exploited further to guarantee a highly reproducible quality of the Italian sweet goods studied, while preserving their traditional trait.

Effects of the Peptide Pheromone Plantaricin A and Cocultivation with Lactobacillus sanfranciscensis DPPMA174 on the Exoproteome and the Adhesion Capacity of Lactobacillus plantarum DC400

This study aimed at investigating the extracellular and cell wall-associated proteins (exoproteome) of Lactobacillus plantarum DC400 when cultivated on modified chemically defined medium (CDM) supplemented with the chemically synthesized pheromone plantaricin A (PlnA) or cocultured with L. plantarum DPPMA20 or Lactobacillus sanfranciscensis DPPMA174. Compared to monoculture, two-dimensional gel electrophoresis (2-DE) analysis showed that the exoproteome of L. plantarum DC400 was affected by PlnA and cocultivation with strains DPPMA20 and, especially, DPPMA174. The highest similarity of the 2-DE maps was found between DC400 cells cultivated in monoculture and in coculture with strain DPPMA20. Almost all extracellular proteins (22 spots) and cell wall-associated proteins (40 spots) which showed decreased or increased levels of synthesis during growth in CDM supplemented with PlnA and/or in coculture with strain DPPMA20 or DPPMA174 were identified. On the basis of the sequences in the Kyoto Encyclopedia of Genes and Genomes database, changes to the exoproteome concerned proteins involved in quorum sensing (QS), the transport system, stress response, carbohydrate metabolism and glycolysis, oxidation/reduction processes, the proteolytic system, amino acid metabolism, cell wall and catabolic processes, and cell shape, growth, and division. Cultivation with PlnA and cocultivation with strains DPPMA20 and, especially, DPMMA174 markedly increased the capacity of L. plantarum DC400 to form biofilms, to adhere to human Caco-2 cells, and to prevent the adhesion of potential intestinal pathogens. These phenotypic traits were in part related to oversynthesized moonlighting proteins (e.g., DnaK and GroEL, pyruvate kinase, enolase, and glyceraldehyde-3-phosphate dehydrogenase) in response to QS mechanisms and interaction with L. plantarum DPPMA20 and, especially, L. sanfranciscensis DPPMA174.

The antimicrobial peptide pheromone Plantaricin A increases antioxidant defenses of human keratinocytes and modulates the expression of filaggrin, involucrin, β‐defensin 2 and tumor necrosis factor‐α genes

Plantaricin A (PlnA) is a peptide with antimicrobial and pheromone activities. PlnA was synthesized chemically and used as a pure peptide or synthesized biologically using Lactobacillus plantarum DC400 co-cultured with Lactobacillus sanfranciscensis DPPMA174. Cell-free supernatant (CFS) was used as a crude PlnA preparation. As estimated using the 3-(4,5-dimethyl-2-yl)-2,5-diphenyltetrazolium bromide and the 2',7'-dichlorofluorescein diacetate assays, both PlnA preparations increased the antioxidant defenses of human NCTC 2544 keratinocytes. PlnA (10μg/ml) had a higher activity than hyaluronic acid or 125μg/ml α-tocopherol. Effects on the transcriptional regulation of filaggrin (FLG), involucrin (IVL), hyaluronan synthase (HAS2), human β-defensin-2 (HBD-2) and tumor necrosis factor-alpha (TNF-α) genes were assayed. Compared with the control, expression of the FLG gene in NCTC 2544 cells increased in cells treated with hyaluronic acid, 1 or 10μg/ml PlnA. Compared with the control, the level of IVL gene expression increased in NCTC 2544 cells treated with 10μg/ml PlnA. No significant difference was found between the level of the HAS2 gene expressed by control cells and cells treated with PlnA. Compared with chemically synthesized PlnA, the up-regulation of the HBD-2 gene by CFS was higher. Compared with the control, expression of TNF-α decreased in NCTC 2544 cells after treatment with 1 or 10μg/ml of chemically synthesized PlnA. In contrast, the level of TNF-α was highest in the presence of 10μg/ml CFS-PlnA. These findings suggest that the PlnA was positively sensed by human keratinocytes, promoting antioxidant defenses, barrier functions and antimicrobial activity of the skin.

Functional and Technological Aspects of Sourdough Fermentation with Lactobacillus sanfranciscensis

In recent years, there has been a growing interest in sourdough bread, which is a traditional product with its lasting production since ancient times. Studies related to the characterizations of sourdough lactic acid bacteria, their contribution to consumer health, dough structure, bread texture and shelf-life have been carried out. In this review, one of the key lactic acid bacteria of sourdough, Lactobacillus sanfranciscensis, is discussed from several aspects. The importance of San Francisco bread for consumer health is also emphasized. In addition to the contribution of L. sanfranciscensis to the textural properties and shelf-life of sourdough bread; the health benefits, flavour and antimicrobial substances, rheological characteristics of sourdough fermented by L. sanfranciscensis are reviewed. The functional aspects of sourdough fermented by L. sanfranciscensis are also demonstrated such as offering alternative bakery products for coeliac patients, diabetics or consumers having allergy to the bakery products fermented with baker’s yeast (Saccharomyces cerevisiae) that are known as anti-S. cerevisiae antibodies.

Effect of sourdough fermentation on the quality of Chinese Northern-style steamed breads

The effect of spontaneous and lactic acid bacteria (LAB) fermentation (Lactobacillus plantarum (LP), Lactobacillus sanfranciscensis (LS), Lactobacillus brevis (LBr) and Lactobacillus delbrueckii subsp bulgaricus (LB) strains) on the specific volume, crumb texture, volatiles profile and other aspects of steamed bread quality were studied. The addition of sourdough starter significantly decreased dough water absorption, development time and stability while increasing steam bread specific volume. The fermentation with LAB starters reached the desired pH 4 five times faster than the spontaneous starters. Steamed bread leavened with LS and LB starters had the softest crumb texture whereas LP and LBr starters produced the lowest setback viscosity. From the total of 77 compounds identified in sourdough steamed bread, ethanol and 3-methyl-1-butanol were the most abundant. All sourdough steamed breads had 22 compounds in common, which represented about 90% of the total peak area. But unique volatile compounds were present in sourdough steamed bread with different starters.

Complex microbiota of a Chinese “Fen” liquor fermentation starter (Fen-Daqu), revealed by culture-dependent and culture-independent methods

Daqu is a traditional fermentation starter that is used for Chinese liquor production. Although partly mechanized, its manufacturing process has remained traditional. We investigated the microbial diversity of Fen-Daqu, a starter for light-flavour liquor, using combined culture-dependent and culture-independent approaches (PCR–DGGE). A total of 190 microbial strains, comprising 109 bacteria and 81 yeasts and moulds, were isolated and identified on the basis of the sequences of their 16S rDNA (bacteria) and 26S rDNA and ITS regions (fungi). DGGE of DNA extracted from Daqu was used to complement the culture-dependent method in order to include non-culturable microbes. Both approaches revealed that Bacillus licheniformis was an abundant bacterial species, and Saccharomycopsis fibuligera, Wickerhamomyces anomalus, and Pichia kudriavzevii were the most common yeasts encountered in Fen-Daqu. Six genera of moulds (Absidia, Aspergillus, Mucor, Rhizopus, Rhizomucor and Penicillium) were found. The potential function of these microorganisms in starters for alcoholic fermentation is discussed. In general the culture-based findings overlapped with those obtained by DGGE by a large extent. However, Weissella cibaria, Weissella confusa, Staphylococcus saprophyticus, Enterobacter aerogenes, Lactobacillus sanfranciscensis, Lactobacillus lactis, and Bacillus megaterium were only revealed by DGGE.

Typing of Lactobacillus sanfranciscensis isolates from traditional sourdoughs by combining conventional and multiplex RAPD–PCR profiles

In the present work, a rapid and reproducible molecular method, based on the combination of conventional and multiplex RAPD–PCR reactions, was developed for typing Lactobacillus sanfranciscensis isolates from traditional sourdoughs. At first, four random primers, two used singly and two combined with the primer RD1, were chosen on the basis of their differentiating capability and reproducibility. The four resulting profiles for each isolate were integrated into a unique profile to be statistically treated by cluster analysis. The method was validated on 58 L. sanfranciscensis isolates coming from three traditional Italian sourdoughs. This new RAPD method was useful for determining the genomic diversity within the L. sanfranciscensis species. In particular, the intraspecific diversity of this species seemed to be related to the sourdough origin.

Levansucrase and sucrose phoshorylase contribute to raffinose, stachyose, and verbascose metabolism by lactobacilli

Raffinose family oligosaccharides (RFOs) in food are considered anti-nutritional factors. This study elucidated the role of α-galactosidase (α-Gal), levansucrase, and sucrose phosphorylase for conversion of RFOs by lactobacilli. Quantification of gene expression by reverse-transcriptase quantitative PCR revealed that expression of levansucrase and sucrose phosphorylase by Lactobacillus reuteri is increased more than 100 fold when sucrose or raffinose are available. Fava bean (Vicia faba) or field pea (Pisum sativum) flours were fermented with α-Gal positive L. reuteri or α-Gal negative Lactobacillus sanfranciscensis. Isogenic strains lacking levansucrase activity, a L. reuteri ftfA mutant and a L. sanfranciscensis levS mutant, were used for comparison. During growth in pulse flours, L. sanfranciscensis accumulated melibiose and α-galactooligosaccharides (α-GOSs); the levansucrase-negative strain did not grow. L. reuteri metabolized raffinose, stachyose, and verbascose by levansucrase activity and accumulated α-GOSs as metabolic intermediates. Oligosaccharide metabolism in the levansucrase-negative mutant was slower, and accumulation of α-GOSs was not observed. The use of sorghum sourdough fermented with L. reuteri LTH5448 and bean flour in gluten-free baking converted RFOs to α-GOSs by levansucrase and invertase activities. In conclusion, the elucidation of the role levansucrase in RFO metabolism by lactobacilli allowed the conversion or hydrolysis of RFOs in food fermentations.