Characterization of the dominant bacterial communities of traditional fermentation starters for Hong Qu glutinous rice wine by means of MALDI-TOF mass spectrometry fingerprinting, 16S rRNA gene sequencing and species-specific PCRs

The main goal of this work was to investigate the bacterial flora present in the traditional fermentation starters using matrix-assisted laser desorption/ionisation time of flight (MALDI-TOF) mass spectrometry fingerprinting combined with 16S rRNA gene sequencing and species-specific PCRs. A total of 1314 bacterial strains were obtained from ten kinds of representative wine fermentation starters (5 Hong Qu and 5 Yao Qu) and classified by MALDI-TOF mass spectrometry. Sixteen different bacterial species belonging to six different genera (Bacillus, Staphylococcus, Leuconostoc, Pediococcus, Lactobacillus and Lactococcus) were identified with high accuracy by comparing them with generated spectra of MALDI Biotyper database. Comparison of 16S rRNA gene phylogeny and MALDI-TOF mass spectrometry analysis showed that MALDI-TOF mass spectrometry fingerprinting was a good complementary approach to 16S rRNA sequencing and even a more powerful tool in the accurate differentiation and classification of Bacillus species, especially for differentiating between Bacillus subtilis, Bacillus amyloliquefaciens and Bacillus methylotrophicus. MALDI-TOF mass spectrometry was also found to provide valuable information at species levels for the genus of Lactobacillus. The identification revealed that most strains isolated from traditional fermentation starters were B. amyloliquefaciens (31.81%), followed by Bacillus megaterium (16.89%), Lc. lactis subsp. lactis (8.60%) and B. subtilis (8.14%). In the culture-dependent quantitative analysis, 69.01% of the detected isolates were Bacillus species, and 30.44% of the detected isolates belonged to the class of lactic acid bacteria (LAB), including Leu. mesenteroides, Pediococcus pentosaceus, Lactobacillus plantarum, Lactobacillus paracasei, Lactobacillus brevis, Pediococcus acidilactici, Lactobacillus coryniformis and Lc. lactis subsp. lactis. This study provides detailed evaluation of bacterial communities in the traditional fermentation starters for Hong Qu glutinous rice wine using a fast and cost-effective technique—MALDI-TOF mass spectrometry fingerprinting. Information provided by this study is fundamental and useful to develop effective strategies for the selection of beneficial bacterial strains and improve of quality of Hong Qu glutinous rice wine.     

Functionality and safety of lactic bacterial strains from Korean kimchi

The functional characteristics and safety of five different putative probiotic Lactobacillus plantarum strains and one Leuconostoc citreum strain, isolated from kimchi, were studied. In view of their potential application in food biotechnology, this information was considered necessary, and included the determination of antibiotic resistance, haemolysis, gelatinase activity and biogenic amine production. In addition to a Lactobacillus fermentum strain isolated from pig faeces, two commercial probiotic strains (Lactobacillus rhamnosus GG, ATCC 53103, and Lactobacillus plantarum 299v, both originating from the human intestinal track) were included in the studies for reference purposes. With the exception of L. fermentum, which was resistant to a low concentration (MIC: 30 μg/ml) of tetracycline, all strains were susceptible to the 8 antibiotics tested (erythromycin, gentamicin, ampicillin, tetracycline, chloramphenicol, streptomycin, ciprofloxacin, and benzylpenicillin). All strains survived physiological conditions typical of the upper GIT, comprising a pH of 3.0 for 1 h, and a subsequent 2 h under conditions simulating the duodenum, which included the exposure to 10 % of bile salts. Moreover, all strains were resistant to 0.4 % phenol, and were unable to produce any detectable biogenic amines under the test conditions. These preliminary in vitro tests indicate the safety and functionality of the five selected L. plantarum strains and thus their potential as probiotic candidates.     

The potentials of Bacillus licheniformis strains for inhibition of B. cereus growth and reduction of biogenic amines in cheonggukjang (Korean fermented unsalted soybean paste)

The aim of this study was to control Bacillus cereus growth and biogenic amines formation by using a starter in cheonggukjang fermentation. We used the four selected B. licheniformis strains for cheonggukjang fermentation and investigated whether the strains are effective in reducing both B. cereus level and biogenic amines content. We confirmed that the selected strains are able to reduce B. cereus level to below l log CFU/g after cheonggukjang fermentation from the initial level of 3–4 log CFU/g (artificial contamination). Tryptamine, β-phenylethylamine, putrescine, cadaverine, tyramine, spermidine, and spermine were detected whereas histamine, serotonin, noradrenaline, and dopamine were not detected in the cheonggukjang samples. Most of biogenic amines contents were lower than those of previous studies, indicating that the strains produced the low level of biogenic amines in cheonggukjang fermentation. The acceptable levels of amino-type nitrogen, free amino acids, and isoflavones were contained in the cheonggukjang samples. These results demonstrate that the selected strains are not only effective for reduction of both B. cereus contamination and biogenic amines but also suitable for preparation of cheonggukjang with good quality.     

Changes in flavour characteristics and bacterial diversity during the traditional fermentation of Chinese rice wines from Shaoxing region

As a traditional alcoholic beverage, Chinese rice wine is quite popular for its unique flavor and high nutritional value. In this study, we investigated the changes of flavor compounds in varying stages of fermentation by HS-SPME-GC-MS and HPLC. In addition, 16s rDNA-PCR was utilized to analyze the changes of bacterial communities during fermentation of Chinese rice wine from the Shaoxing region. It was noted that the content of flavor compounds including esters, higher alcohols, amino acids and organic acids was different in varying stages of fermentation. In general, the flavor compounds of Chinese rice wine were mainly produced after pre-fermentation. The results also showed that the bacterial community structures and diversity of Chinese rice wine varied significantly during different fermentation stages, and more than 10 genera of bacteria was detected in Chinese rice wine fermentation broth. Among these specific bacteria identified in the study, Lactobacillus (the predominant genus) and Bacillus might take an active part in flavor development in Chinese rice wine. This implied that these bacteria might play significant contributions on the flavor of Chinese rice wine.     

Biomass production and small-scale testing of freeze-dried lactic acid bacteria starter strains for cassava fermentations

Based on their predominance in Gari fermentations, as well as suitable technological properties, Lactobacillus plantarum, Lactobacillus fermentum, Weissella paramesenteroides and Leuconostoc mesenteroides strains were investigated for their suitability for development as starter strains for this African traditional fermented cassava product. The strains were grown in optimized growth media in 2 L fermenters, harvested and freeze dried, and then tested in lab-scale cassava mash fermentation trials for their ability to ferment the cassava. The strains performed well and rapidly increased the titratable acidity from 1.1 to 1.3% at 24 h to 1.3–1.6% at 48 h. The benefit of including starter cultures was that it lowered the pH of the product much faster and to lower levels than in the uninoculated control fermentation. The results furthermore indicated that especially the L. plantarum-group strains could be produced as starter strains at low cost. Overall, the results of this study showed that starter strains could be easily and economically produced, and thus represent a feasible possibility for further development for application in the field.     

Evaluation of functional properties of lactobacilli isolated from Korean white kimchi

Kimchi, probably Korea's most famous traditional fermented food, is well known for its beneficial properties. Among several hundred different types of kimchi in Korea, white (baek) kimchi is prepared without chilli and is widely appreciated also by non-Koreans because of its unique mild flavour. In an approach to identify the bacteriological basis for proposed health benefits, we isolated 11 Lactobacillus strains from six samples of white kimchi, and investigated their safety and functional features. These strains represented the species Lactobacillus brevis, Lactobacillus plantarum and Lactobacillus sakei that dominated the populations within a range of 3 × 10⁶ to 4 × 10⁸ CFU/mL. Following safety assessment based on antibiotic resistance and biogenic amine production, 7 different strains were selected for further studies including evaluation of their adaptation to cabbage juice and resistance to phenol. Growth in and adaptation to the cabbage juice was favourably influenced by addition of 2% salt. Final selection was based on in vitro passage of simulated stomach duodenum conditions (SSDP model). The strains L. plantarum HAC01 and L. sakei HAC10 were administered to a diet-induced obese (DIO) mouse model receiving a high-fat (HF) diet to assess their functionality in-vivo. Animal groups receiving the viable strains showed significantly lower body weight and total weight gain during 8 weeks compared to the high-fat control group. This study provides preliminary information on the use of in-vitro and in-vivo features for safety and functionality evaluation of Lactobacillus strains from white kimchi. These “first-level” criteria for strain selection may serve as model, thereby facilitating potentially new probiotic developments.     

Starter culture fermentation of Chinese sauerkraut: Growth,acidification and metabolic analyses

Chinese sauerkraut is a kind of traditional and typical fermented food. Four lactic acid bacteria (LAB) strains, Leuconostoc mesenteroides NCU1426, Lactococcus lactis NCU1315, Lactobacillus plantarum NCU1121 and Lactobacillus casei NCU1222 isolated from Chinese sauerkraut, were used in single starter cultures. Microbiological changes and pH values were monitored during fermentation. Metabolic substrates and products during the fermentation were examined using high performance liquid chromatography (HPLC) technology. Results have shown that Leu. mesenteroides and Lc. lactis grew faster, produced lactic acid earlier and were poorly acid-resistant, whereas Lb. plantarum and Lb. casei produced much more lactic acid throughout fermentation and showed better acid-tolerance. Two Lactococcus had outstanding performance in sucrose utilization while the other two Lactobacillus were likely to use glucose and fructose during fermentation. Unexpectedly, Leu. mesenteroides and Lc. lactis showed weak citric acid metabolism in fermentation. All the four LAB strains were able to utilize malic acid in early fermentation. In conclusion, these LAB strains have shown notable differences in growth and fermentative properties during starter culture fermentation of Chinese sauerkraut, probably resulting from LAB fermentative function and a mixture of complex substrates.     

Occurrence of antilisterial structural bacteriocins genes in meat borne lactic acid bacteria

The ability to inhibit the growth of Listeria cells and the presence of bacteriocin encoding genes was examined in 115 LAB strains isolated from Argentinean vacuum-packaged beef and different traditional fermented sausages. Lactobacillus (L) sakei, Lactobacillus (L) curvatus and Enterococcus (E) faecium showed a great inhibition of all Listeria strains evaluated while Pediococcus (P) acidilactici and Lactobacillus (L) plantarum demonstrated a limited or absent antilisterial activity. Both L. curvatus and L. sakei carried the sppA, sppQ and sapA structural genes, encoding for sakacin P, sakacin Q and curvacin A bacteriocins, respectively. Whilst L. curvatus exhibited a higher occurrence of these genes, L. sakei strains were more effective at inhibiting Listeria (L) strains, Listeria monocytogenes UC8159 and Listeria innocua 7 being the most sensitive to these bacteriocins. Among analyzed E. faecium strains, the wide distribution of entA, entB and entP genes accounted for the high antilisterial activity particularly observed against L. monocytogenes FBUNT. The structural gene plantEF was mostly present in Lactobacillus plantarum strains and no pedA gene was found in P. acidilactici evaluated strains. The antilisterial potential of L. sakei and E. faecium offers great possibilities for the meat industry as biopreservative cultures, although more studies are needed in order to conclude about this issue.     

Biogenic amine inhibition and quality protection of Harbin dry sausages by inoculation with Staphylococcus xylosus and Lactobacillus plantarum

This study was conducted to evaluate the effects of inoculation with Staphylococcus xylosus, Lactobacillus plantarum, or a mixture of strains (L. plantarum + S. xylosus) on the formation of biogenic amines (BAs) and quality characteristics in Harbin dry sausage. Microbial analysis shows that total aerobic bacteria and lactic acid bacteria (LAB) counts were higher in the inoculated sausages, especially in those inoculated with a mixture of strains, but the growth of enterobacteriaceae was inhibited (P < 0.05). A sharp decrease in the pH value of the sausages was observed, and the moisture content and water activity were significantly decreased during fermentation. Sausages inoculated with a mixture of bacterial strains had the lowest pH, moisture content and water activity (P < 0.05). Inoculation of dry sausages with S. xylosus or L. plantarum, especially a mixture of strains (L. plantarum + S. xylosus), significantly delayed lipid oxidation, improved sensory characteristics, and inhibited BA accumulation. Six types of BAs (cadaverine, putrescine, tryptamine, 2-phenylethylamine, histamine, and tyramine) were inhibited by the presence of L. plantarum and S. xylosus, and a mixture of them had the most inhibitive effect. Correlation analysis showed that the BA concentrations correlated well with enterobacteriaceae counts, and some BAs were negatively correlated with LAB counts. These results demonstrate that S. xylosus and L. plantarum could be used as starter cultures in Harbin dry sausage production to inhibit BA accumulation and improve quality characteristics.     

Preliminary evaluation of biogenic amines content in Chilean young varietal wines by HPLC

Biogenic amines are organic nitrogenous compounds produced in wines mainly during malolactic fermentation. Its presence is a health risk for sensitive individuals and it could negatively affect the wine quality. The objective of this work was to determine the biogenic amines content in Chilean young varietal wines. For that, a liquid chromatographic method with pre-column derivatization was optimized and validated to achieve a reliable quantification of the most important biogenic amines in wines, i.e. histamine, tyramine, spermine, spermidine, putrescine, cadaverine and phenylethylamine. Using a face-centered central composite design derivatization and chromatographic conditions were optimized. Derivatization with dansyl chloride showed an optimum yield for all biogenic amines at 60 °C during 60 min. These derivatives were optimally separated with a C₁₈ YMC-Pack ODS-A column set a 40 °C using a mobile phase composed of acetonitrile and water under gradient conditions. An effective separation and quantification of the biogenic amines was achieved in 40 min using ultraviolet detection at 254 nm. Validation was established according to ICH guidelines, calibrations in matrix fitted a linear regression model with R² > 0.997. Repeatability (n = 6) and intermediate precision (n = 3) in matrix showed RSD values <2.34% and <3.16%, respectively. This method was applied to determine for the first time the biogenic amines content in Chilean wines. Twenty-seven samples of young varietal wines, i.e. Cabernet Sauvignon (n = 9), Merlot (n = 8) and Carménère (n = 10) were analyzed finding a biogenic amines content from 18.12 to 39.84 mg L^-1, where putrescine showed the highest concentration in all grape varieties. No significant difference was found between grapes varieties regarding total biogenic amines content (P = 0.7842, df = 26). Along with putrescine, histamine and tyramine were the most abundant biogenic amines but similar levels were also observed with spermidine.     

Induction of plantaricin MG under co-culture with certain lactic acid bacterial strains and identification of LuxS mediated quorum sensing system in Lactobacillus plantarum KLDS1.0391

A bacteriocin named plantaricin MG was produced by Lactobacillus plantarum KLDS1.0391 which was isolated from “jiaoke”, a traditional, naturally fermented cream from Inner Mongolia in China. Bacteriocin production was increased significantly when L. plantarum KLDS1.0391 co-cultured with certain lactic acid bacteria (LAB) strains. Among 76 strains belonging to the genera Lactobacillus, Lactococcus, Leuconostoc and Streptococcus, four strains namely Lactobacillus helveticus KLDS1.9207, Enterococcus faecium KLDS4.0352, Lactobacillus reuteri KLDS1.0737 and Enterococcus faecalis KLDS4.0313 were shown to induce bacteriocin production of L. plantarum KLDS1.0391. Cell numbers of L. plantarum KLDS1.0391 were greatly enhanced when co-cultured with four bacteriocin-inducing strains. Bacteriocin production was not induced by autoclaved cultures and cell-free supernatants (CFS) of inducing strains, indicating that living cells of inducing strains might be necessary for enhancement of bacteriocin production. The presence of plnD and luxS genes was detected by polymerase chain reaction (PCR), and the existence of plNC8HK gene was detected by single oligonucleotide nested PCR (SON-PCR).     

Bacterial dynamics during the spontaneous fermentation of cassava dough in gari production

The microbial dynamics and diversity during solid substrate fermentation of cassava, a case study of gari production in West Africa, was investigated. The 16S rDNA gene sequence analysis of the PCR-Denaturing Gradient Gel Electrophoresis (PCR–DGGE) analysis of microbial community DNA and Pulsed Field Gel Electrophoresis (PFGE) of selected isolates as well as culturing techniques using different selective media were used to monitor the bacterial dynamics during cassava fermentation. The V3 variable region of the 16S gene was analyzed and the closest relatives of Lactobacillus plantarum, Lactobacillus fermentum, Lactobacillus pentosus, Lactobacillus acidophilus and Lactobacillus casei were identified by sequencing of the DGGE band amplimers. The DGGE amplimers also revealed the succession and dynamics of LAB; there was a progressive increase in their population proportional to the fermentation period. The analysis of the PFGE band patterns showed that five diverse species of LAB were involved in the fermentation. The representative isolate of each of the PFGE clusters was phenotypicaly identified as L. plantarum, L. fermentum and Leuconostoc mesenteroides by the API 50 CHL sugar fermentation profile. These combinations of parameters identified heterofermentative LAB as bacteria that initiated the fermentation, reduced the pH below four and increased the acidity of the fermentation mash. Information such as this is relevant for the development of starter cultures and predictability of the process for traditional fermented foods and to aid their intermediate and large scale production.     

Development of propidium monoazide combined with real-time quantitative PCR (PMA-qPCR) assays to quantify viable dominant microorganisms responsible for the traditional brewing of Hong Qu glutinous rice wine

The main disadvantage of DNA-based quantification methods is their inability to distinguish between viable and non-viable cells, as dead cells may also retain a significant amount of DNA. To overcome this limitation, propidium monoazide (PMA) treatment was applied to effectively prevent amplification of DNA from dead cells. The objectives of this study were to develop and evaluate an accurate detection method that combines PMA staining with real-time qPCR to quantify viable dominant microorganisms during the traditional brewing of Hong Qu glutinous rice wine. Firstly, three methodological factors (concentration of PMA, incubation time and photoactivation time) were optimized for representative strains of different microbial species. For the tested microbial strains (Monascus purpureus, Saccharomyces cerevisiae and Lactobacillus plantarum), a treatment of 25 μM PMA for 20 min incubation followed by 10 min photoactivation was considered optimal to achieve a compromise between minimal impact on intact cells and maximal signal reduction for the compromised cells. Compared with qPCR without PMA treatment, results of PMA-qPCR showed that the optimized PMA pretreatment can effectively exclude the dead microbial cells without any negative effects on the intact cells. The concentrations of viable cells calculated by PMA-qPCR were comparable (the same order of magnitude was obtained) with those obtained by culture-based method. Finally, the developed PMA-qPCR methods were successfully applied to monitor the total bacteria, LAB, fungi, yeasts, M. purpureus, S. cerevisiae and L. plantarum during the traditional brewing of Hong Qu glutinous rice wine. Both of the total bacterial and fungal populations increased significantly at the early brewing stage, but they decreased at the later brewing stage. PMA-qPCR results revealed that the predominant microorganisms varied in different brewing phases. L. plantarum and S. cerevisiae grew significantly and became the predominant bacterial and fungal species at the final stage of traditional brewing process, respectively. This study standardized the PMA-qPCR parameters for dominant microorganisms associated with the traditional brewing of Hong Qu glutinous rice wine, which would show great value in scientifically understanding of the brewing mechanism and provide useful information for the selection of beneficial microbial strains to improve wine quality.     

Diversity of lactic acid bacteria and yeasts in traditional sourdoughs collected from western region in Inner Mongolia of China

Twenty-eight traditional sourdough samples for Chinese steamed breads were collected from the western region in Inner Mongolia of China. The values of pH and the total titratable acidity (TTA) of sourdoughs varied from 2.78–5.13 to 8.3–15.5 mL, respectively. In all samples, the number of lactic acid bacteria (LAB) ranged from 6.56 to 8.72 Log cfu/g, while the amount of yeasts ranged from 4.13 to 6.37 Log cfu/g. A total of 108 LABs and 85 yeasts were identified using 16S rRNA gene, 26S rRNA gene sequencing and species-specific PCR. All LAB isolates were identified as 17 species, and all the yeasts were identified as 11 species. Diversity of lactic acid bacteria and yeasts were analyzed by denaturing gradient gel electrophoresis (DGGE). Both the results of identification and DGGE profiles analysis indicated that Lactobacillus plantarum group and Saccharomyces cerevisiae were the predominant microflora in our samples. Further studies by species-specific PCR revealed that all the isolates in Lactobacillus plantarum group were Lactobacillus plantarum. Interestingly, Lactobacillus guizhouensis, Lactobacillus rossiae, Rhodotorula mucilaginosa and Pichia farinosa isolated by cultured-based method were not detected by DGGE, whereas Lactobacillus zeae was only found in DGGE profiles.     

Reduction in vitro of the minor Fusarium mycotoxin beauvericin employing different strains of probiotic bacteria

The interaction between the minor Fusarium mycotoxins BEA and 13 bacterial strains characteristic of the gastrointestinal tract as Bifidobacterium longum, Bifidobacterium bifidum, Bifidobacterium breve, Bifidobacterium adolescentes, Lactobacillus rhamnosus, Lactobacillus casei-casei, Lactobacillus plantarum, Eubacterium crispatus, Salmonella fecalis, Salmonella termofilus, Lactobacillus ruminis, Lactobacillus casei and Lactobacillus animalis was studied.The fermentations were carried out in the liquid medium of MRS during 4, 12, 16, 24 and 48 h at 37 °C, under anaerobic conditions.Levels of BEA in the fermentation liquid, on the cell walls and on the internal part of the cells were determined using liquid chromatography coupled to the mass spectrometry detector (LC-MS/MS). Results showed that the bacteria reduced the concentration of the BEA present in the medium, part of the mycotoxin was adsorbed by cell wall and part internalized by the bacteria. All the bacteria analyzed in this study showed a significant BEA reduction during the fermentation process, in particular the mean diminution resulted variable from 66 to the 83%.     

Occurrence of biogenic amines in beers from Chilean market

The presence of tryptamine, phenylethylamine, putrescine, cadaverine, histamine, tyramine, spermidine and spermine was determined for the first time in beers commercialized in Chilean market. Dansylated amines were separated on a Zorbax-XDB C₁₈ column using a binary mobile phase composed of acetonitrile and 20 mmol L⁻¹ ammonium formate, pH 5.5. Chromatographic conditions were optimized using an experimental design, and validation was carried out following ICH recommendations. Calibration data fitted a linear regression model with R² > 0.996. Repeatability (n = 6) and intermediate precision (n = 3) in matrix showed RSD values lower than 5.00% and 5.21%, respectively. Recoveries at three different concentrations ranged from 75.50 to 96.48%. The proposed method was applied to determine the biogenic amines content in 101 beer samples produced by macro- (n = 65) and microbreweries (n = 36) using low (n = 63) and high fermentations (n = 38). Biogenic amines content in beer samples ranged from 0.53 to 85.04 mg L⁻¹, from which beers produced by microbreweries (19.13 ± 16.92 mg L⁻¹) showed a significant higher biogenic amines content (P = 0.0021) than beers produced by macrobreweries (9.65 ± 4.50 mg L⁻¹). Putrescine was the principal biogenic amine found in all kind of beers regardless origin, and the type of fermentation and brewery. Only 2 samples presented relevant levels of histamine and tyramine, but both below the limits reported by the European Food Safety Authority (EFSA). Thus, it can be concluded that beers commercialized in Chile are not a serious toxicological risk (e.g. hypertensive crisis) regarding the type and content of biogenic amines.     

Prevention of bread mould spoilage by using lactic acid bacteria with antifungal properties

The ability of lactic acid bacteria (LAB) to inhibit Aspergillus, Fusarium, and Penicillium, the main contaminants in bread, was evaluated. Only four strains (Lactobacillus plantarum CRL 778, Lactobacillus reuteri CRL 1100, and Lactobacillus brevis CRL 772 and CRL 796) from 95 strains tested displayed antifungal activity. The major antifungal compounds were acetic and phenyllactic acids. The fermentation quotient (FQ = 2.0) and the leaven volume (80 cm³) of doughs with LB and yeasts were higher than doughs without LB. The inclusion of antifungal LAB strains in the starter culture allowed a reduction in the concentration of calcium propionate by 50% while still attaining a shelf life similar to that of traditional bread containing 0.4% CP.     

Reduction of biogenic amines and aflatoxins in Doenjang samples fermented with various Meju as starter cultures

The objective of this study was to determine the contents of biogenic amines (BAs) and aflatoxin levels in laboratory-made starter culture soybean fermented samples of Doenjang. Various starter cultures of Meju, made by using different microbial strains, were used for the production of Doenjang samples under standardized fermentation/manufacturing conditions, and the contents of BAs and aflatoxins were analyzed. Determination of nine different BAs and total aflatoxin was carried out by high performance liquid chromatography and immunoassay-based total aflatoxin detection kit, respectively. It was found that the amount of BAs in all laboratory-made starter culture Doenjang samples was within the limit of human consumption safety level. Also the level of total aflatoxin was under the limit, according to the standard parameters of FAO/WHO.     

Antimicrobial phenolic extracts able to inhibit lactic acid bacteria growth and wine malolactic fermentation

The purpose of this study was to determine whether phenolic extracts with antimicrobial activity may be considered as an alternative to the use of sulfur dioxide (SO₂) for controlling malolactic fermentation (MLF) in winemaking. Inhibition of the growth of six enological strains (Lactobacillus hilgardii CIAL-49, Lactobacillus casei CIAL-52, Lactobacillus plantarum CIAL-92, Pediococcus pentosaceus CIAL-85, Oenococcus oeni CIAL-91 and O. oeni CIAL-96) by phenolic extracts (n = 54) from different origins (spices, flowers, leaves, fruits, legumes, seeds, skins, agricultural by-products and others) was evaluated, being the survival parameter IC₅₀ calculated. A total of 24 extracts were found to significantly inhibit the growth of at least two of the LAB strains studied. Some of these extracts were also active against two acetic acid bacteria (Acetobacter aceti CIAL-106 and Gluconobacter oxydans CIAL-107). Transmission electron microscopy of the bacteria cells after incubation with the phenolic extract confirmed damage of the integrity of the cell membrane. Finally, to test the technological applicability of the extracts, the eucalyptus extract was added (2 g/L) to an industrially elaborated red wine, and the progress of the MLF was evaluated by means of residual content of malic acid. Addition of the extract significantly delayed the progress of both inoculated and spontaneous MLF, in comparison to the control wine (no antimicrobial agent added), although not as effective as K₂S₂O₅ (30 mg/L). These results demonstrated the potential applicability of phenolic extracts as antimicrobial agents in winemaking.     

A new fear in wine: Isolation of Staphylococcus epidermidis histamine producer

Biogenic amines (BAs) are undesirable compounds in all foods and beverages because they may induce food-borne intoxications when consumed at high concentrations. The aim of this study was to identify the organism responsible for the synthesis of histamine in a Tempranillo red wine from Ribera de Duero D.O. (Denomination of origin), Spain. Bacteria present in wines after malolactic fermentation were isolated, identified and typed. Four strains of the species Oenococcus oeni and two strains of Staphylococcus epidermidis were found. None of the O. oeni strains produced histamine, cadaverine or putrescine, but one of the S. epidermidis strains (Pa) was able to produce all three in synthetic medium and grape must, although not in wine. From the data obtained in this work, histamine present in Tempranillo wine B was produced by the S. epidermidis strain Pa. This is the first report in which the presence of S. epidermidis has been reported in wine and whose ability to produce histamine, putrescine and cadaverine has been demonstrated in grape must.