Papers by francesca valerio
International Journal of Food Microbiology, 2009
A fully automated electrophoretic method for separating cell-wall proteins was applied to 102 iso... more A fully automated electrophoretic method for separating cell-wall proteins was applied to 102 isolates from olive phylloplane and brine previously phenotypically identified as Lactobacillus plantarum , Leuconostoc mesenteroides subsp. mesenteroides (60), Leuconostoc spp. (12) and Enterococcus faecium (19). The high sensitivity of this technique allowed to discriminate isolates and to distinguish closely related strains within the same species. L. plantarum was characterized by proteins at 70.4 and 163.9 kDa, while 3 proteins (20.4, 52.9, 68. kDa) were typical for E. faecium. No characteristic proteins were found for Leuconostoc isolates except for that at 31.8 kDa mainly related to the ecological origin since only isolates from brine showed this protein. However, Leuc. mesenteroides subsp. mesenteroides and Leuconostoc spp. were differentiated by additional proteins. Sequence analysis of the 16S rRNA gene of selected strains was in agreement with the phenotypic identification and allowed to identify all Leuconostoc spp. isolates as Leuc. mesenteroides. Cellwall protein fingerprinting performed by microfluidic technology completed the phenotypic characterization of olive-related lactic acid bacteria allowing their strain typing.
Systematic and Applied Microbiology, 2009
Thirty samples of Italian durum wheat semolina and whole durum wheat semolina, generally used for... more Thirty samples of Italian durum wheat semolina and whole durum wheat semolina, generally used for the production of Southern Italy's traditional breads, were subjected to microbiological analysis in order to explore their lactic acid bacteria (LAB) diversity and to find strains with antifungal activity. A total of 125 presumptive LAB isolates (Gram-positive and catalase-negative) were characterized by repetitive extragenic palindromic-PCR (REP-PCR) and sequence analysis of the 16S rRNA gene, leading to the identification of the following species: Weissella confusa, Weissella cibaria, Leuconostoc citreum, Leuconostoc mesenteroides, Lactococcus lactis, Lactobacillus rossiae and Lactobacillus plantarum. The REP-PCR results delineated 17 different patterns whose cluster analysis clearly differentiated W. cibaria from W. confusa isolates. Seventeen strains, each characterized by a different REP-PCR pattern, were screened for their antifungal properties. They were grown in a flour-based medium, comparable to a real food system, and the resulting fermentation products (FPs) were tested against fungal species generally contaminating bakery products, Aspergillus niger, Penicillium roqueforti and Endomyces fibuliger. The results of the study indicated a strong inhibitory activity -comparable to that obtained with the common preservative calcium propionate (0.3% w/v) -of ten LAB strains against the most widespread contaminant of bakery products, P. roqueforti. The screening also highlighted the unexplored antifungal activity of L. citreum, L. rossiae and W. cibaria (1 strain), which inhibited all fungal strains to the same or a higher extent compared with calcium propionate. The fermentation products of these three strains were characterized by low pH values, and a high content of lactic and acetic acids.
Fems Microbiology Letters, 2004
The ability of lactic acid bacteria (LAB) to produce phenyllactic (PLA) and 4-hydroxy-phenyllacti... more The ability of lactic acid bacteria (LAB) to produce phenyllactic (PLA) and 4-hydroxy-phenyllactic (OH-PLA) acids, metabolites involved in food quality and preservation, has been evaluated by HPLC analysis in 29 LAB strains belonging to 12 species widely used in the production of fermented foods. Metabolite production was demonstrated for all strains of the species Lactobacillus plantarum, Lactobacillus alimentarius, Lactobacillus rhamnosus, Lactobacillus sanfranciscensis, Lactobacillus hilgardii, Leuconostoc citreum, and for some strains of Lactobacillus brevis, Lactobacillus acidophilus and Leuconostoc mesenteroides subsp. mesenteroides. Strains were distinguished by analysis of variance in three groups including 15 strains that produced both metabolites (0.16-0.46 mM PLA and 0.07-0.29 mM OH-PLA), five strains accumulating in culture only PLA (0.17-0.57 mM) and nine non-producer strains ( 6 0.10 mM PLA and 6 0.02 mM OH-PLA). Improvement of phenyllactic acid production was obtained in a selected L. plantarum strain by increasing the concentration of phenylalanine in culture and using low amounts of tyrosine.
Current Microbiology, 2008
Strains of Lactobacillus spp., isolated from sourdough and olive brines (seven strains), and thre... more Strains of Lactobacillus spp., isolated from sourdough and olive brines (seven strains), and three human isolates were screened for their antagonistic activity in coculture against the ureolytic pathogen Yersinia enterocolitica. A general reduction in the pathogen population was observed after 6 h when each Lactobacillus strain was cocultured with the pathogen at a ratio of 100:1 cfu ml−1, causing an almost complete inhibition of urease activity. Strains were also screened for their performances in in vitro tests such as adhesion ability to Caco-2 cells, tolerance to low pH, bile salts, and simulated digestion, which enabled the differences between strains to be highlighted. Three strains, L. paracasei IMPC2.1 and L. plantarum ITM21B and ITM5BG, met the main criteria for selecting effective probiotics: the ability to inhibit the pathogen Y. enterocolitica and, consequently, its urease activity (ITM21B); survival of simulated digestion (ITM21B and IMPC2.1); strong adhesion ability to enterocytes and good survival at low pH and in the presence of bile salts (ITM5BG and IMPC2.1).
Applied and Environmental Microbiology, 2006
The ability of potentially probiotic strains of Lactobacillus plantarum and Lactobacillus paracas... more The ability of potentially probiotic strains of Lactobacillus plantarum and Lactobacillus paracasei to survive on artichokes for at least 90 days was shown. The anchorage of bacterial strains to artichokes improved their survival in simulated gastrointestinal digestion. L. paracasei IMPC2.1 was further used in an artichoke human feeding study involving four volunteers, and it was shown that the organism could be recovered from stools.
International Journal of Food Microbiology, 2008
Four fermentation products (FPs) of the lactic acid bacterium Lactobacillus plantarum ITM21B were... more Four fermentation products (FPs) of the lactic acid bacterium Lactobacillus plantarum ITM21B were screened for their anti-Bacillus activity in vitro and in bread-making trials. Results of the storage tests performed with loaves prepared with an FP or calcium propionate demonstrated that after 3 days at 30°C, gross spoilage was evident in only the control loaves, which contained Bacillus subtilis at numbers of about 10 9 cfu/g. The highest inhibitory activity was shown by DM-FP obtained by growing L. plantarum in a defined medium (DM). Significantly, this medium contained an amino acceptor of the aminoacid transamination, namely α-ketoglutaric acid, and an aminoacid pool. With loaves prepared using the DM-acid mixture which simulated the DM-FP composition, the same reduction of ropy spoilage as with DM-FP was obtained after 3 days, while the efficacy of the mixture decreased after 7 days. This result suggests the potential involvement of some unknown metabolites in the inhibitory activity of DM-FP. In baked products made with flour based media (M1-FP, M2-FP, M3-FP), no ropy symptoms were noticeable after 3 days storage although a considerable Bacillus count was detected. DM-FP was as effective as calcium propionate (0.3% w/w, based on flour mass) in prolonging the Bacillus free-shelf life of yeast-leavened bread for 7 days.
International Journal of Food Microbiology, 2010
This study reports the dynamics of microbial populations adhering on the surface of debittered gr... more This study reports the dynamics of microbial populations adhering on the surface of debittered green olives cv. Bella di Cerignola in fermentation sets inoculated with the probiotic strain Lactobacillus paracasei IMPC2.1 in different brining conditions (4% and 8% (w/v) NaCl) at room temperature and 4°C. The probiotic strain successfully colonized the olive surface dominating the natural LAB population and decreasing the pH of brines to ≤ 5.0 after 30 days until the end of fermentation. The dynamics of microbial populations associated with olive surface and belonging to the different groups indicated that inoculated olives held at room temperature did not host Enterobacteriaceae at the end of fermentation. Yeast populations were present in a low number (≤ log 10 5.7 CFU/g) throughout the process. A considerable genetic diversity of LAB species colonizing the olive surface was found mainly in inoculated set brined in 8% NaCl, as indicated by the Shannon diversity index calculated for each set. Generally, strains of Lactobacillus coryniformis, L. paracasei, L. plantarum, L. pentosus, L. rhamnosus, L. brevis, L. mali, L. vaccinostercus, L. casei, Leuconostoc mesenteroides, Leuc. pseudomesenteroides, Lactococcus lactis, Weissella paramesenteroides, W. cibaria, Enterococcus casseliflavus group and E. italicus were identified during the whole process. In particular, L. pentosus was the most frequently isolated species and it showed a high strain diversity throughout fermentation in all processes except for the one held at 4°C. Also a notable incidence of Leuc. mesenteroides on olives was highlighted in this study during all fermentation. Results indicated that the human strain L. paracasei IMPC2.1 can be considered an example of a strain used in the dual role of starter and probiotic culture which allowed the control of fermentation processes and the realization of a final probiotic product with functional appeal.
Antonie Van Leeuwenhoek International Journal of General and Molecular Microbiology, 2010
The main aim of this work was the identification of genetic determinants involved in bacteriocin ... more The main aim of this work was the identification of genetic determinants involved in bacteriocin production by strain ITM317 of Pseudomonas savastanoi pv. savastanoi, besides bacteriocin characterization. The bacteriocin was observed to be a heat-sensitive, high molecular weight proteinaceous compound. We identified a transposon (Tn5)-induced mutant which had lost its ability to produce the bacteriocin. The Tn5 insertion’s responsibility for the above mutated phenotype was demonstrated by marker-exchange mutagenesis. An EcoRI DNA fragment, corresponding to the EcoRI Tn5-containing fragment of the mutant, was also cloned from the wild-type strain, and its introduction into the mutant complemented the mutation. Moreover, that fragment enabled bacteriocin production by P. s. pv. savastanoi ITM302, a strain not previously capable of doing so. DNA sequence analysis revealed that Tn5 insertion occurred in the mutant within a large ORF encoding a protein which showed similarity with proteins from the Rhs family. The DNA region including that ORF showed features which have been considered typical of the Rhs genetic elements previously identified in other bacteria but whose function is as yet unclear. The results of this study for the first time identify an Rhs-like element in P. s. pv. savastanoi, and for the first time indicate that an Rhs element is involved in bacteriocin production, also suggesting this possible function for Rhs genetic elements previously characterized in other bacteria.
Journal of Clinical Gastroenterology, 2010
We have used a simplified gnotobiotic mouse model to evaluate the effects of single bacterial spe... more We have used a simplified gnotobiotic mouse model to evaluate the effects of single bacterial species, Lactobacillus paracasei NCC2461, on the metabolic profiles of intact intestinal tissues using highresolution magic-angle-spinning 1 H NMR spectroscopy (HRMAS). A total of 24 female gnotobiotic mice were divided into three groups: a control group supplemented with water and two groups supplemented with either live L. paracasei or a γ-irradiated equivalent. HRMAS was used to characterize the biochemical components of intact epithelial tissues from the duodenum, jejunum, ileum, proximal, and distal colons in all animals and data were analyzed using chemometrics. Variations in relative concentrations of amino acids, anti-oxidant, and creatine were observed relating to different physiological properties in each intestinal tissue. Metabolic characteristics of lipogenesis and fat storage were observed in the jejunum and colon. Colonization with live L. paracasei induced region-dependent changes in the metabolic profiles of all intestinal tissues, except for the colon, consistent with modulation of intestinal digestion, absorption of nutrients, energy metabolism, lipid synthesis and protective functions. Ingestion of γ-irradiated bacteria produced no effects on the observed metabolic profiles. 1 H MAS NMR spectroscopy was able to generate characteristic metabolic signatures reflecting the structure and function of intestinal tissues. These signals acted as reference profiles with which to compare changes in response to gut microbiota manipulation at the tissue level as demonstrated by ingestion of a bacterial probiotic.
Applied and Environmental Microbiology, 2005
With the aim of developing new functional foods, a traditional product, the table olive, was used... more With the aim of developing new functional foods, a traditional product, the table olive, was used as a vehicle for incorporating probiotic bacterial species. Survival on table olives of Lactobacillus rhamnosus (three strains), Lactobacillus paracasei (two strains), Bifidobacterium bifidum (one strain), and Bifidobacterium longum (one strain) at room temperature was investigated. The results obtained using a selected olive sample demonstrated that bifidobacteria and one strain of L. rhamnosus (Lactobacillus GG) showed a good survival rate, with a recovery of about 10 6 CFU g ؊1 after 30 days. The Lactobacillus GG population remained unvaried until the end of the experiment, while a slight decline (to about 10 5 CFU g ؊1 ) was observed for bifidobacteria. High viability, with more than 10 7 CFU g ؊1 , was observed throughout the 3-month experiment for L. paracasei IMPC2.1. This strain, selected for its potential probiotic characteristics and for its lengthy survival on olives, was used to validate table olives as a carrier for transporting bacterial cells into the human gastrointestinal tract. L. paracasei IMPC2.1 was recovered from fecal samples in four out of five volunteers fed 10 to 15 olives per day carrying about 10 9 to 10 10 viable cells for 10 days.
Applied and Environmental Microbiology, 2000
Sourdough lactic acid bacteria were selected for antifungal activity by a conidial germination as... more Sourdough lactic acid bacteria were selected for antifungal activity by a conidial germination assay. The 10-fold-concentrated culture filtrate of Lactobacillus plantarum 21B grown in wheat flour hydrolysate almost completely inhibited Eurotium repens IBT18000, Eurotium rubrum FTDC3228, Penicillium corylophilum IBT6978, Penicillium roqueforti IBT18687, Penicillium expansum IDM/FS2, Endomyces fibuliger IBT605 and IDM3812, Aspergillus niger FTDC3227 and IDM1, Aspergillus flavus FTDC3226, Monilia sitophila IDM/FS5, and Fusarium graminearum IDM623. The nonconcentrated culture filtrate of L. plantarum 21B grown in whole wheat flour hydrolysate had similar inhibitory activity. The activity was fungicidal. Calcium propionate at 3 mg ml ؊1 was not effective under the same assay conditions, while sodium benzoate caused inhibition similar to L. plantarum 21B. After extraction with ethyl acetate, preparative silica gel thin-layer chromatography, and chromatographic and spectroscopic analyses, novel antifungal compounds such as phenyllactic and 4-hydroxyphenyllactic acids were identified in the culture filtrate of L. plantarum 21B. Phenyllactic acid was contained at the highest concentration in the bacterial culture filtrate and had the highest activity. It inhibited all the fungi tested at a concentration of 50 mg ml ؊1 except for P. roqueforti IBT18687 and P. corylophilum IBT6978 (inhibitory concentration, 166 mg ml ؊1 ). L. plantarum 20B, which showed high antimold activity, was also selected. Preliminary studies showed that phenyllactic and 4-hydroxy-phenyllactic acids were also contained in the bacterial culture filtrate of strain 20B. Growth of A. niger FTDC3227 occurred after 2 days in breads started with Saccharomyces cerevisiae 141 alone or with S. cerevisiae and Lactobacillus brevis 1D, an unselected but acidifying lactic acid bacterium, while the onset of fungal growth was delayed for 7 days in bread started with S. cerevisiae and selected L. plantarum 21B.
Fems Microbiology Letters, 2004
The ability of lactic acid bacteria (LAB) to produce phenyllactic (PLA) and 4-hydroxy-phenyllacti... more The ability of lactic acid bacteria (LAB) to produce phenyllactic (PLA) and 4-hydroxy-phenyllactic (OH-PLA) acids, metabolites involved in food quality and preservation, has been evaluated by HPLC analysis in 29 LAB strains belonging to 12 species widely used in the production of fermented foods. Metabolite production was demonstrated for all strains of the species Lactobacillus plantarum, Lactobacillus alimentarius, Lactobacillus rhamnosus, Lactobacillus sanfranciscensis, Lactobacillus hilgardii, Leuconostoc citreum, and for some strains of Lactobacillus brevis, Lactobacillus acidophilus and Leuconostoc mesenteroides subsp. mesenteroides. Strains were distinguished by analysis of variance in three groups including 15 strains that produced both metabolites (0.16–0.46 mM PLA and 0.07–0.29 mM OH-PLA), five strains accumulating in culture only PLA (0.17–0.57 mM) and nine non-producer strains (≤0.10 mM PLA and ≤0.02 mM OH-PLA). Improvement of phenyllactic acid production was obtained in a selected L. plantarum strain by increasing the concentration of phenylalanine in culture and using low amounts of tyrosine.
Applied and Environmental Microbiology, 2003
Phenyllactic acid (PLA) has recently been found in cultures of Lactobacillus plantarum that show ... more Phenyllactic acid (PLA) has recently been found in cultures of Lactobacillus plantarum that show antifungal activity in sourdough breads. The fungicidal activity of PLA and growth inhibition by PLA were evaluated by using a microdilution test and 23 fungal strains belonging to 14 species of Aspergillus, Penicillium, and Fusarium that were isolated from bakery products, flours, or cereals. Less than 7.5 mg of PLA ml ؊1 was required to obtain 90% growth inhibition for all strains, while fungicidal activity against 19 strains was shown by PLA at levels of <10 mg ml ؊1 . Levels of growth inhibition of 50 to 92.4% were observed for all fungal strains after incubation for 3 days in the presence of 7.5 mg of PLA ml ؊1 in buffered medium at pH 4, which is a condition more similar to those in real food systems. Under these experimental conditions PLA caused an unpredictable delaying effect that was more than 2 days long for 12 strains, including some mycotoxigenic strains of Penicillium verrucosum and Penicillium citrinum and a strain of Penicillium roqueforti (the most widespread contaminant of bakery products); a growth delay of about 2 days was observed for seven other strains. The effect of pH on the inhibitory activity of PLA and the combined effects of the major organic acids produced by lactic acid bacteria isolated from sourdough bread (PLA, lactic acid, and acetic acid) were also investigated. The ability of PLA to act as a fungicide and delay the growth of a variety of fungal contaminants provides new perspectives for possibly using this natural antimicrobial compound to control fungal contaminants and extend the shelf lives of foods and/or feedstuffs.
Applied and Environmental Microbiology, 2002
A bacteriocin produced by Pseudomonas syringae pv. ciccaronei, used at different purification lev... more A bacteriocin produced by Pseudomonas syringae pv. ciccaronei, used at different purification levels and concentrations in culture and in planta, inhibited the multiplication of P. syringae subsp. savastanoi, the causal agent of olive knot disease, and affected the epiphytic survival of the pathogen on the leaves and twigs of treated olive plants. Treatments with bacteriocin from P. syringae pv. ciccaronei inhibited the formation of overgrowths on olive plants caused by P. syringae subsp. savastanoi strains PVBa229 and PVBa304 inoculated on V-shaped slits and on leaf scars at concentrations of 10 5 and 10 8 CFU ml ؊1 , respectively. In particular, the application of 6,000 arbitrary units (AU) of crude bacteriocin (dialyzed ammonium sulfate precipitate of culture supernatant) ml ؊1 at the inoculated V-shaped slits and leaf scars resulted in the formation of knots with weight values reduced by 81 and 51%, respectively, compared to the control, depending on the strains and inoculation method used. Crude bacteriocin (6,000 AU ml ؊1 ) was also effective in controlling the multiplication of epiphytic populations of the pathogen. In particular, the bacterial populations recovered after 30 days were at least 350 and 20 times lower than the control populations on twigs and on leaves, respectively. These results suggest that bacteriocin from P. syringae pv. ciccaronei can be used effectively to control the survival of the causal agent of olive knot disease and to prevent its multiplication at inoculation sites.
The effects of wheat bran and of a Lactobacillus brevis-based bioingredient (LbBio), obtained aft... more The effects of wheat bran and of a Lactobacillus brevis-based bioingredient (LbBio), obtained after growth in flourbased medium, on quality of yeast-leavened wheat bread (WWB) were investigated. Bran was used in bread formulation by substituting a part (20 g/100 g) of white wheat flour (WBB), while LbBio was used instead of the water content (WWB + LbBio and WBB + LbBio). The use of LbBio in WWB resulted in the biological acidification of the dough due to lactic, phenyllactic and OH-phenyllactic acid contents determining a high fermentation quotient value and an improved bread texture and microbiological quality. Conversely, wheat bran reduced the specific volume and crumb hardness during storage at 25°C, and affected the antibacterial ability of LbBio during 30°C storage. Our findings demonstrated that LbBio counteracted the negative effects of bran and allowed to obtain an enriched fibre bread with specific volume and soft crumb comparable to bread without bran. Industrial relevance: Bread is a perishable food with a short microbiological and physico-chemical shelf-life. The main microbiological alteration occurring into few days after baking is the "rope spoilage" caused by spore-forming bacteria originating from raw materials. This phenomenon, often misinterpreted as a sign of unsuccessful dough leavening and not visible from outside, is more common under industrial production conditions during the hot season causing large economic losses in the warm climates of Mediterranean countries, Africa and Australia. The use of sourdough often controls this alteration even if the industrial application of this traditional process is limited by the long leavening times. In this study, an innovative procedure for the preparation of yeastleavened bread comprising the addition of a fermentation product from Lactobacillus brevis grown in a flourbased medium has been applied. The resulting fermentation product (LbBio bioingredient) acts as a sourdough acidifying the dough and improving the textural, physico-chemical and microbiological properties of the resulting bread. The application of bioingredient LbBio could represent an innovative strategy in industrial bread production to obtain acidified yeast-leavened products, thus, preventing the ropy spoilage and reducing the negative effects of bran addition.
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Papers by francesca valerio