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Oenococcus oeni plays a key role in inducing malolactic fermentation in wine. Acid stress is often encountered under wine conditions. However, the lack of systematic studies of acid resistance mechanisms limits the downstream fermentation applications. In this study, the acid responses of O. oeni were investigated by combining transcriptome, metabolome, and genome-scale metabolic modeling approaches. Metabolite profiling highlighted the decreased abundance of nucleotides under acid stress. The gene-metabolite bipartite network showed negative correlations between nucleotides and genes involved in ribosome assembly, translation, and post-translational processes, suggesting that stringent response could be activated under acid stress. Genome-scale metabolic modeling revealed marked flux rerouting, including reallocation of pyruvate, attenuation of glycolysis, utilization of carbon sources other than glucose, and enhancement of nucleotide salvage and the arginine deiminase pathway. This study provided novel insights into the acid responses of O. oeni, which will be useful for designing strategies to address acid stress in wine malolactic fermentation.

Alicyclobacillus acidoterrestris causes the spoilage of pasteurized acidic fruit juice, seriouslydecreasing quality and posing a significant safety concern. We previously discovered that acid adaptation could induce stress adaptive responses of A. acidoterrestris, however, the underlying mechanisms of this induction have not been fully elucidated. In this work, the effects of acid adaptation (pH = 3.0, 1 h) on intracellular pH (pH

Acid-resistant bacteria are more and more widely used in industrial production due to their unique acid-resistant properties. In order to survive in various acidic environments, acid-resistant bacteria have developed diverse protective mechanisms such as sensing acid stress and signal transduction, maintaining intracellular pH homeostasis by controlling the flow of H

Aroma compounds in fruits and related products are known to occur in free and glycosidically bound forms. In this study, free and bound volatiles in juices and wines of two kiwifruit cultivars, Actinidia deliciosa ‘Hayward’ and A. chinensis ‘Hort16A’, were analyzed by solid-phase microextraction-gas chromatography-mass spectrometry. Results showed that 67 free and 79 bound volatiles were found in juices and wines. Alcohols were the most abundant free volatiles, while the most representative bound volatiles were terpenoids. β-Damascenone and 4-methyl-3-penten-2-one released from aroma precursors were found in ‘Hort16A’ juice for the first time. Fruity, floral, and sweet flavors appeared to be the characteristic aroma in kiwifruit juices and wines. Principal component analysis showed that kiwifruit wines had a great aroma potential. Cluster analysis revealed that juices and wines had different profile of free volatiles, while similar bound volatile compositions were found. Moreover, abundant bound volatiles were discovered in these two kiwifruit pomace for the first time, indicating their potential applications in food and cosmetics industry.

Overuse and misuse of antibiotics have resulted in wide antibiotic resistance in bacteria. In this study, surface-enhanced Raman spectroscopy (SERS) was used to rapidly characterize the responses of lactic acid bacteria (LAB) to antibiotics targeting the bacterial cell wall. Lactobacillus delbrueckii subsp. bulgaricus (Lb. bulgaricus) ATCC 11842 was used to represent LAB strains that are widely used in the food industry. Penicillin G, ampicillin and vancomycin were used to treat Lb. bulgaricus ATCC 11842 at different toxic levels for 30, 60 and 90 min, respectively. SERS spectra of Lb. bulgaricus after each treatment were collected to characterize the responses of Lb. bulgaricus. The collected spectra were analyzed by principal component analysis (PCA) and partial least squares regression (PLSR). Results suggested that all three antibiotics induced significant peak changes in LAB, and the spectral changes induced by each antibiotic treatment were significantly different. Moreover, the antibiotic-induced spectral changes in Lb. bulgaricus had good correlation with its proliferation ability and could be potentially used as a base for rapid quantification of the antibiotic susceptibility of bacteria. The developed method could detect the reaction of LAB to antibiotic treatments within 3 h. Therefore, the developed SERS method has the potential to be used to rapidly discriminate and quantify the antibiotic susceptibility of LAB. Moreover, such a rapid and direct method could contribute to microbial antibiotic susceptibility detection research.

For the first time, thinned young kiwifruits (TYK) have been investigated as a source of deserted agriculture resource and evaluated for their phenolic characteristics along with antioxidant capacity. Two batches of TYK in different maturity stages (TYK1 and TYK2) and mature kiwifruits (MK) were sourced from three New Zealand cultivars. Comparing with MK, the TYK, especially TYK1, exhibited significantly higher contents of total phenolics (7.92 fold), total flavonoids (17.56 fold) and total flavanols (10.11 fold) as well as elevated and phenolic-correlated antioxidant capacities. A novel UPLC-QqQ-MS/MS method with low limit of detection (0.010-0.240 mg/L) and high accuracy (RSDs 6.79%) was established to quantify 15 phenolics in the kiwifruit extracts. Epicatechin was the prevailing antioxidant in both TKY and MK, with the highest level of 2989.23 μg/g FDW present in TYK1 of 'Zesy002' cultivar. These results provide good evidence for the development and utilization of TYK of these cultivars in food and nutraceuticals.

To investigate the impact of inorganic and organic nitrogen on volatile compounds of cider, various levels (60, 150 and 240 mg N/L) of diammonium hydrogen phosphate (DAP) and amino acids mixture were added to apple juice before fermentation, respectively. The qualitative and quantitative of volatile substances were carried out by GC-MS. Further, sensory and electronic nose analysis was performed to analyze the aroma profile of cider. Both DAP and amino acids mixture promoted fermentation, significantly affected volatile compositions and increased perceived aromas of cider. Adding 60 mg N/L DAP significantly increased volatile compounds content, especially medium-chain fatty acid ethyl esters (MCFAEEs) and fatty acids (FAs), whereas higher alcohols and acetate esters concentrations were remarkably increased with 150 mg N/L amino acids supplementation. Partial least-squares (PLS) regression analysis suggested that MCFAEEs contributed the most to tropical fruity aroma. The study provides a theoretical foundation for high-quality cider brewing.

Alicyclobacillus acidoterrestris has been regarded as the main hazardous factor causing the spoilage of commercially pasteurized fruit juice due to its unique thermo-acidophilic properties. p-Coumaric acid, primarily isolated from plants and having high biological activity, is supposed to be a viable food additive. Herein, the antibacterial activity and mechanism of p-coumaric acid against A. acidoterrestris and its application in apple juice were investigated. The results showed that p-coumaric acid had an active inhibition against A. acidoterrestris vegetative cells, and both minimum inhibition concentration (MIC) and minimum bactericidal concentration (MBC) were 0.2 mg/mL. MIC of p-coumaric acid against spores was also 0.2 mg/mL, while MBC was1.6 mg/mL. Treatment of p-coumaric acid caused significant damage of cell integrity, decrease of intracellular ATP, hyperpolarization of cell membrane, degradation of whole cell protein, and malformation of cell morphology. Agarose gel retardation and fluorescence spectroscopy assays revealed that p-coumaric acid could interact with DNA and intercalate into the groove of DNA double helix to disturb normal cellular functions. Furthermore, the addition of p-coumaric acid at MIC caused 2.07 log drop in A. acidoterrestris vegetative cells and effectively inhibited the germination of spores in apple juice storage at 20 °C for 7 days. The low-temperature storage (4 °C) could inactivate A. acidoterrestris cells in apple juice, but the addition of p-coumaric acid further accelerated degradation of vegetative cells and the inactivation time was shortened from 5 days to 3 days compared with control. Importantly, the addition of p-coumaric acid could increase total phenolic content and ABTS radical scavenging activity and had no remarkable influence on pH, total soluble solids, color and sensory qualities of apple juice, regardless of storage temperature. These results revealed that p-coumaric acid could be expected to serve as an alternative or supplement agent for inhibiting the development of A. acidoterrestris in fruit juice.

Alicyclobacillus acidoterrestris can survive pasteurization and is implicated in pasteurized fruit juice spoilage. However, the mechanisms underlying heat responses remain largely unknown. Herein, gene transcription changes of A. acidoterrestris under heat stress were detected by transcriptome, and an integrated analysis with proteomic and physiological data was conducted. A total of 911 differentially expressed genes (DEGs) was observed. The majority of DEGs and differentially expressed proteins (DEPs) were exclusively regulated at the mRNA and protein level, respectively, whereas only 59 genes were regulated at both levels and had the same change trends. Comparative analysis of the functions of the specifically or commonly regulated DEGs and DEPs revealed that the heat resistance of A. acidoterrestris was primarily based on modulating peptidoglycan and fatty acid composition to maintain cell envelope integrity. Low energy consumption strategies were established with attenuated glycolysis, decreased ribosome de novo synthesis, and activated ribosome hibernation. Terminal oxidases, cytochrome bd and aa

Background During winemaking, malolactic fermentation (MLF) is usually induced by Oenococcus oeni owing to its high resistance to wine stress factors. To ensure a controlled and efficient MLF process, starter cultures are inoculated in wine. In previous studies, O. oeni strains with sub-lethal acid or ethanol stresses showed higher freeze-drying vitality and better MLF performance. To explore the mechanisms involved, influences of acid and ethanol stresses on O. oeni SD-2a were investigated in this study to gain a better understanding of the cross-protection responses. Results The results showed that acid and ethanol stresses both caused damage to cell membranes and decreased cellular adenosine triphosphate concentration. At the same time, acid stress increased the uptake of glutathione, while ethanol stress led to cell depolarization. The results of comparative proteomic analysis highlighted that heat shock protein was induced with almost all acid and ethanol stresses. In addition, the expression of stress-relevant genes (hsp20, clpP, trxA, ctsR, recO, usp) increased greatly with ethanol and acid stress treatments. Finally, the viability of O. oeni was improved with acid and ethanol pretreatments after freeze-drying. Conclusions This study demonstrated that acid and ethanol stresses had mixed influences on O. oeni SD-2a. Some physiological and molecular changes would contribute to a more stress-tolerant state of O. oeni, thereby improving the viability of lyophilized cells. © 2020 Society of Chemical Industry.

Persimmon tannin (PT), a primary physiological component of young persimmon, exhibits antibacterial activity against several microorganisms, but its effect on methicillin-resistant Staphylococcus aureus (MRSA) remains unclear. In this study, we found that PT from young astringent persimmon fruit had an active inhibition against several MRSA isolated from retail pork and the minimum inhibitory concentrations were 1000 μg/mL. Treatment of PT suppressed cell proliferation, caused decreases in membrane potential and intracellular ATP concentration, destroyed whole cell protein integrity, and induced S phase cell cycle arrest. Electron microscopy demonstrated that PT can change the normal morphology of MRSA and cause severe damage on cell wall and cell membrane. These results indicated that PT presented antimicrobial activity against foodborne MRSA and could be potentially developed as an alternative or supplemental agent to control the spread and infection of MRSA in foods.

The aim of this study was to screen potential probiotic lactic acid bacteria from Chinese spontaneously fermented non-dairy foods by evaluating their probiotic and safety properties. All lactic acid bacteria (LAB) strains were identified by 16S rRNA gene sequencing. The in vitro probiotic tests included survival under low pH and bile salts, cell surface hydrophobicity, auto-aggregation, co-aggregation, antibacterial activity, and adherence ability to cells. The safety properties were evaluated based on hemolytic activity and antibiotic resistance profile. The salt tolerance, growth in litmus milk, and acidification ability were examined on selected potential probiotic LAB strains to investigate their potential use in food fermentation. A total of 122 strains were isolated and identified at the species level by 16S rRNA gene sequencing and included 62 Lactobacillus plantarum, 40 Weissella cibaria, 12 Lactobacillus brevis, 6 Weissella confusa, and 2 Lactobacillus sakei strains. One W. cibaria and nine L. plantarum isolates were selected based on their tolerance to low pH and bile salts. The hydrophobicity, auto-aggregation, co-aggregation, and antagonistic activities of these isolates varied greatly. All of the 10 selected strains showed multiple antibiotic resistance phenotypes and no hemolytic activity. The highest adhesion capacity to SW480 cells was observed with L. plantarum SK1. The isolates L. plantarum SK1, CB9, and CB10 were the most similar strains to Lactobacillus rhamnosus GG and selected for their high salt tolerance and acidifying activity. The results revealed strain-specific probiotic properties were and potential probiotics that can be used in the food industry.

The aim of this study was to evaluate the antibiotic susceptibility of lactic acid bacteria (LAB) and coagulase‐negative staphylococci (CNS) strains isolated from naturally fermented smoked pork produced in Hunan, China. A total of 48 strains were isolated by selective medium and identified at the species level by 16S rRNA gene sequencing as follows: Staphylococcuscarnosus (23), Lactobacillus plantarum(12), Lactobacillus brevis(10), Lactobacillus sakei(1), Weissella confusa(1), and Weissella cibaria(1). All strains were typed by RAPD‐PCR, and their susceptibility to 15 antibiotics was determined and expressed as the minimum inhibitory concentration (MIC) using agar dilution method. High resistance to penicillin G, streptomycin, gentamycin, vancomycin, chloramphenicol, norfloxacin, ciprofloxacin, kanamycin, and neomycin was found among the isolates. All the strains were sensitive to ampicillin, while the susceptibility to tetracycline, oxytetracycline, erythromycin, lincomycin, and roxithromycin varied. The presence of relevant resistance genes was investigated by PCR and sequencing, with the following genes detected: str(A), str(B), tet(O), tet(M), ere(A), and catA. Eleven strains, including 3 S. carnosus, 6 L. plantarum, and 2 L. brevis, harbored more than 3 antibiotic resistance genes. Overall, multiple antibiotic resistance patterns were widely observed in LAB and S. carnosusstrains isolated from Hunan smoked pork. Risk assessment should be carried out with regard to the safe use of LAB and CNS in food production. We evaluated the antibiotic resistance of lactic acid bacteria and coagulase‐negative staphylococci strains isolated from Chinese naturally fermented smoked pork. Our results may provide important data on establishing breakpoint standards for LAB and CNS and evaluating the safety risk of these strains for commercial use.

Phloridzin has been an attractive target for the development of preparation methodologies due to its various applications as pharmaceutical ingredient and food additive. In this study, an enzymatic synthesis approach using engineered P. pastoris GS115 was developed to produce phloridzin. Four factors, identifying as key factors affecting the phloridzin yield, were optimized by response surface methodology as follows: induction time 121.8 h, pH 7.0, methanol 0.75%, and a rotation speed 194 rpm. The maximal yield of phloridzin reached 41.59 mg/L (95.31 μM) and the specific bioconversion rate of substrate was up to 98.50% with 1L working volume in a 5L bioreactor under the optimal conditions. Phloridzin with purity of 93.98% and recovery of 78.14% was achieved just using affinity chromatography and preparative high performance liquid chromatography ( Pre -HPLC). These results confirm the enzymatic synthesis developed provides an efficient alternative to traditional approach for the preparation of phloridzin in vitro .

Principal component analysis (PCA) and partial least squares (PLS) regression were applied to investigate the effect of glutathione-enriched inactive dry yeast (g-IDY) on the amino acids and volatile components of kiwi wine. Results indicated that the addition of g-IDY had positive effect on most amino acids of kiwi wine, especially glutamine and glycine. In case of pure juice fermentation, the concentrations of ethyl decanoate, 2-methylbutyric acid, trans-2-nonenal and hexyl butyrate had notably positive correlation with the addition of g-IDY. PLS regression indicated that the amino acids were highly interrelated to the volatile compositions, and glycine had the strongest positive impact on the concentrations of esters and total volatile components. This might explain the similar effect of g-IDY on the amino acids and volatile components of kiwi wine. Besides, PLS regression showed that E-nose was a good method to predict volatile compositions of kiwi wine, especially esters.

Fresh “Xuxiang” kiwifruit juice with or without pomace was used for fermentation with different levels of glutathione‐enriched inactive dry yeast (g‐IDY). After alcohol fermentation, the glutathione (GSH) content, color parameters, phenolic composition, and antioxidant activities of kiwi wine were determined. The results suggested that the addition of g‐IDY to kiwi juice increased GSH, glutathione disulfide, total GSH concentrations, and the total phenolic content of kiwi wine. No obvious effect on individual phenolic content was found in wine with g‐IDY. The color parameters of wines fermented with pomace were remarkably improved with addition of 100 mg/L and 300 mg/L g‐IDY. The supply of g‐IDY improved the antioxidant activities of kiwi wine fermented with pomace. Principal component analysis suggested that with the same supplies of g‐IDY, wine fermented with pomace was better than wine fermented through pure juice in all measurements. PRACTICAL APPLICATIONS: The improvements in color and antioxidant capacity of kiwi wine have always been the focus of kiwi wine brewing. Constant improvement in methods which contribute to high quality of kiwi wine provides crucial support for wine industry and even agriculture development. The aim of this study is to investigate the impacts of glutathione‐enriched inactive dry yeast (g‐IDY) on quality of kiwi wine. These results could provide feasible advices about g‐IDY application to kiwi wine for improving the sensory and nutritional properties.

Oenococcus oeni can exert its function in hostile wine conditions during the malolactic fermentation process. Therefore, it is an important microbial resource for exploring resistance genes. Hsp20 is an important small heat shock protein from O. oeni. The conserved consensus motif “A-x-x-x-x-G-x-L” of Hsp20 announced its role as a member of the small heat shock protein family. The hsp20 gene from O. oeni SD-2a was cloned to create the recombinant plasmid pTriEx-Hsp20. The recombinant plasmid was transformed into Escherichia coli BL21(DE3) competent cells, and the Hsp20 protein was induced by isopropyl-β-d-thiogalactoside (IPTG). The hsp20 gene from O. oeni SD-2a was successfully expressed, and a 20-kDa fusion protein was identified by SDS-PAGE. The purified Hsp20 protein was obtained using Ni-affinity chromatography. Additionally, BL21(DE3)/Hsp20 and BL21(DE3)/Ctrl were treated at high temperatures of 42 and 52 °C, at pH values of 2.0–12.0, under oxidative shock with 0.1% (v/v) and 0.2% (v/v) H2O2, and under an osmotic shock of 430 and 860 mM NaCl to compare the effects of heterologous expression of the Hsp20 protein from O. oeni SD-2a for stress resistance. Notably, Hsp20 overexpression showed enhanced resistance than the control strain did when confronted with different elevated stress conditions. The results demonstrated heterologous expression of the hsp20 gene from O. oeni SD-2a significantly improved the resistance of the host E. coli bacteria against stress conditions.

The aim of this study was to evaluate probiotic properties of ten Streptococcus thermophilus strains (st1 to st10) isolated from pickles in China. These strains all had β-galactosidase activity, which laid foundation for studying their probiotic properties. In this study, the bile salt hydrolase activity, lysozyme resistance, tolerance to simulated gastric juice, bile salt tolerance, and bacterial adhesion capacity to the Caco-2 cells of these selected strains were detected in vitro conditions. The results indicated that the bile salt hydrolase activities of st2, st6, and st9 were higher than that for other strains. St10 showed the greatest lysozyme resistance (> 80% survival), followed by st9, st8, st7, st5, and st6. As for the tolerance to simulated gastric juice, st5 possessed the highest survival rate (35%), followed by st6 (30%). St6 was the best performer in both bile salt tolerance and bacterial adhesion capacity to the Caco-2 cells. The results of fluorescence microscope and electron microscope further confirmed previous studies and more intuitively demonstrated the st6 strain's tolerance to harsh environments. Overall, these strains were expected to possess beneficial properties and have the potentiality to be probiotics.

Persimmon tannin (PT) exhibits antibacterial activity against methicillin-resistant Staphylococcus aureus (MRSA) isolated from retail pork. The involved molecular mechanisms were investigated for the first time using transcriptome and metabolome in this study. Results showed that subinhibitory concentration of PT (0.5 mg/ml) induced significant changes in MRSA at both transcriptional and metabolic levels, as 370 genes and 19 metabolites were differentially expressed. Bioinformatic analysis revealed that the varying genes and metabolites were mainly involved in pathways of membrane transport, amino acids, carbohydrate, and energy metabolism. The highlighted changes were those related to osmotic regulation, intracellular pH regulation, amino acid synthesis and metabolism, glycolysis, TCA cycle and iron metabolism, suggesting the multifaceted effects including cell membrane damage, amino acids limitation, energy metabolism disorder and iron deprivation induced by PT. The results provided insight into the anti-MRSA mechanism of PT, which is useful for PT’s development and application in food safety.

The β-glucosidase derived from microorganisms has attracted worldwide interest for their industrial applications, but studies on β-glucosidases from Oenococcus oeni are rare. In this paper, catalytic mechanism of a novel β-glucosidase BGL0224 of Oenococcus oeni SD-2a was explored for the first time by kinetic parameters determination, fluorescence spectroscopy and quenching mechanism analysis, molecular dynamics simulation. The results indicated that BGL0224 had universal catalytic effect on different types of glycoside substrates, but the catalytic efficiencies were different. Fluorescence quenching analysis results suggested that the quenching processes between BGL0224 and seven kinds of substrates were predominated by the static quenching mechanism. A reasonable three-dimensional model of BGL0224 was obtained using the crystal structure of E.coli BglA as a template. The analysis results of molecular simulation (RMSD, Rg, RMSF and hydrogen bonding) showed that the composite system ‘BGL0224-pNPG’ was very stable after 40 ns. The catalytic process of BGL0224 acting on ‘p-Nitrophenyl β- d -glucopyranoside’ conformed to the double displacement mechanism. Two glutamic acid residues ‘Glu178 and Glu377’ played a vital role in the whole catalytic process. Overall, this study gave specific insights on the catalytic mechanism of BGL0224, which was of great significance for developing its potential applications in food industry.

β-glucosidase is widely used in food industry for its glycosidic hydrolysis properties. In this paper, performance of a novel β-glucosidase BGL0224 from Oenococcus oeni for aroma enhancement of Cabernet Sauvignon wines was explored for the first time. The main projects included electronic nose, gas chromatography-mass spectrometry and sensory analysis. Principal component analysis indicated that electronic nose had the ability to discriminate different treatments by responding specifically to volatile compounds. Adding BGL0224 before alcoholic fermentation significantly increased the “aromatic index” of Cabernet Sauvignon wines. Compared with the control group, adding BGL0224 increased more than 15 volatile aroma compounds of Cabernet Sauvignon wines. Furthermore, with the addition of BGL0224 before alcoholic fermentation, the concentrations of medium-chain fatty acid ethyl esters, long-chain fatty acid ethyl esters and terpenes in wines were 3004.19 ± 36.14 μg/L, 4090.37 ± 94.07 μg/L and 210.21 ± 8.98 μg/L respectively, which were all significantly higher than other treatments. Sensory analysis showed that BGL0224 increased “tropical fruity”, “sweet fruity” and “floral notes” characteristics while inhibited “citrus” characteristics of Cabernet Sauvignon wines. Overall, this study provided support for industrial application of BGL0224 by evaluating its performance for aroma enhancement of Cabernet Sauvignon wines.

The effects of selenium (Se) addition methods on antioxidant activity and flavor properties of fermented Pleurotus eryngii (P. eryngii) using Lactobacillus plantarum (L. plantarum) inoculated and natural fermentation were investigated. After fermentation, the Se-enrichment rates in fruiting bodies of Se-added fermented P. eryngii were all more than 50%. Se addition, especially in the form of Se-enriched L. plantarum inoculation, had a significantly positive effect on total phenolic content and DPPH radical scavenging activity. Non-volatiles analysis revealed that the highest ration of lactic acid to acetic acid and the highest umami intensity were observed in P. eryngii fermented by inoculating Se-enriched L. plantarum (Lp-Se). Principal components analysis and cluster analysis of volatiles clearly separated Se-treated and plain experiments, which mainly due to dissimilarities in alcohols, aldehydes and ketones. Additionally, Lp-Se obtained the highest alcohols, especially 1-octen-3-ol with mushroom flavor. In short, Se-enriched L. plantarum inoculation could produce high-quality fermented P. eryngii.

A novel method was developed to rapidly concentrate, detect, and differentiate bacteria in skimmed milk using surface enhanced Raman scattering (SERS) mapping on 4-mercaptophenylboronic acid (4-MPBA) functionalized silver (Ag) dendrites. The 4-MPBA functionalized Ag dendritic SERS substrate was used to capture the bacterial cells and enhance the bacterial signal. Salmonella, a significantly important food pathogen, was used as the representative strain to optimize and evaluate the developed method. The capture efficiency for Salmonella enterica subsp enterica BAA1045 (SE1045) was 84.92 ± 3.25% at 106 CFU/mL and as high as 99.65 ± 3.58% at 103 CFU/mL. Four different strains, two gram-negative and two gram-positive, can be clearly distinguished by their SERS spectra using principle component analysis. A mapping technique was utilized to automatically collect 400 spectra over an area of 60 μm × 60 μm to construct a visual image for a sensitive and statistically reliable detection within 30 min. Using this method, we were able to detect as low as 103 CFU/mL bacterial cells in 50 mM NH4HCO3 solution and 102 CFU/mL cells in both 1% casein and skimmed milk. Our results demonstrate the feasibility of using SERS mapping method coupled with 4-MPBA functionalized Ag dendrites for rapid and sensitive bacteria detection in complex liquid samples.

Oenococcus oeni (O. oeni) strains are critically important for winemaking. They bring microbial stabilization, de-acidification and sensory improvement through malolactic fermentation (MLF) to almost all red wines and some white wines. In this study, 23 strains of O. oeni isolated from China were evaluated for use as MLF starter cultures. Strains were characterized and screened in wine-like media with increasing selective pressure, such as low pH and high ethanol concentration. Their abilities to conduct MLF were affected significantly by ethanol concentration and pH. Eight strains were selected and subsequently evaluated by detecting the enzyme-encoding genes involved in biogenic amines synthesis and wine aroma modification. Seven well-performing strains were further assessed in newly alcohol-fermented Cabernet Sauvignon wine. The results show that strains O. oeni CS-4b and O. oeni ME-5b exhibited better performance than the commercial starter O. oeni 31MBR in wine. Therefore, they were selected as potential MLF starters suitable for wine with high ethanol concentration and low pH.

Polyphenols are the predominant ingredients in apple seeds. However, few data are available on the phenolic profile or antioxidant activity in apple seeds in previous researches. In this study, low-molecular-weight phenolic compounds and antioxidant activity in seeds, peels, and flesh of seven apple cultivars grown in northwest China were measured and analyzed using HPLC and FRAP, DPPH, ABTS assays, respectively. HPLC analysis revealed phloridzin as the dominant phenolic compound in the seeds with its contents being 240.45–864.42mg/100gDW. Total phenolic content (TPC) measured by the Folin–Ciocalteu assay in apple seed extracts of seven cultivars ranged from 5.74 (Golden Delicious) to 17.44 (Honeycrisp)mgGAE/gDW. Apple seeds showed higher antioxidant activity than peels or flesh; antioxidant activity in seeds varied from 57.59 to 397.70μM Trolox equivalents (TE)/g FW for FRAP, from 37.56 to 64.31μM TE/g FW for DPPH, and from 220.52 to 708.02μM TE/g FW for ABTS. TPC in apple seeds was significantly correlated with all three assays. Principal component analysis (PCA) indicated that Honeycrisp was characterized with high contents of total polyphenols and phloridzin. Our findings suggest that phenolic extracts from apple seeds have good commercial potential as a promising antioxidant for use in food or cosmetics.

β-Glucosidases play an important role in food industry. Oenococcus oeni are typical lactic acid bacteria that initiate malolactic fermentation of wines. 35 β-glucosidases from O. oeni were selected and their conserved domains and evolutionary relationships were further explored in this study. The homology analysis results indicated that 35 β-glucosidases were basically derived from GH1 and GH3 family. A novel β-glucosidase was successfully expressed and characterized. The recombinant protein, referred to as BGL0224, consisted of a total 480 amino acids with an apparent molecular weight of 55.15 kDa and was classified as GH1 family. It achieved the highest activity at pH 5.0 and 50 °C. The activity and stability were significantly increased when 12% ethanol was supplemented to the enzyme. Using p-NPG as substrate, the Km, Vmax and Kcat of BGL0224 were 0.34 mM, 382.81 U/mg and 351.88 s−1, respectively. In all, BGL0224 has good application prospects in food industry.

Ethanol acclimations have been proven to be beneficial for enhancing the viability of lyophilized Oenococcus oeni. To explore the relevant mechanisms, cell integrity, protein patterns, morphology, exopolysaccharides (EPS) content of lyophilized O. oeni SD-2a with ethanol acclimations were investigated in this research. After freeze-drying, the improvement of cell integrity was found in all ethanol acclimated cells. Comparative proteomics combined with bioinformatics revealed the important role of central carbohydrate metabolism, polysaccharides production, cell envelope biosynthesis and stress responses related proteins with ethanol acclimations in freeze-dried O. oeni SD-2a. Extracellular polymeric substances were observed in mild ethanol (5% and 8%) adapted cells using scanning electron microscope. In addition, mild ethanol (5% and 8%) adapted cells showed higher EPS content. Transcriptional analysis showed that citrate lysis gene over-expressed in all ethanol acclimated freeze-dried cells. Therefore, citrate was added in culture medium and it is highlighted that the addition of citrate was useful for the growth of freeze-dried O. oeni SD-2a. These results provide new insights of molecular mechanisms regulated by ethanol acclimation and improve our understanding of biological characteristics involved in freeze-dried O. oeni.

This study provided phenotypic and molecular analysis of the antibiotic resistance within coagulase-negative staphylococci and lactic acid bacteria isolated from naturally fermented Chinese cured beef. A total of 49 strains were isolated by selective medium and identified at the species level by 16S rRNA gene sequencing as follows: Staphylococcus carnosus (37), Lactobacillus plantarum (6), Weissella confusa (4), Lactobacillus sakei (1), and Weissella cibaria (1). All strains were typed by random amplified polymorphic DNA fingerprinting, and their antibiotic resistances profiles to 15 antibiotics were determined as the MIC by using the agar dilution method. All the tested strains were sensitive to ampicillin, and most of them were also sensitive to penicillin, gentamycin, neomycin, norfloxacin, and ciprofloxacin with low MICs. High resistance to streptomycin, vancomycin, erythromycin, roxithromycin, lincomycin, and kanamycin was widely observed, while the resistant levels to tetracycline, oxytetracycline, and chloramphenicol varied. The presence of corresponding resistance genes in resistant isolates was investigated by PCR, with the following genes detected: tet(M) gene in 9 S. carnosus strains and 1 W. confusa strain; erm(F) gene in 10 S. carnosus strains; ere(A) gene in 6 S. carnosus strains; ere(A) gene in 4 S. carnosus strains and 1 L. plantarum strain; and str(A) gene and str(B) gene in 3 S. carnosus strains. The results indicated that multiple antibiotic resistances were common in coagulase-negative staphylococci and lactic acid bacteria strains isolated from naturally fermented Chinese cured beef. Safety analysis and risk assessment should be performed for application in meat products.

Malolactic fermentation (MLF), usually induced by Oenococcus oeni (O. oeni), is an important process to improve wine quality. Acid acclimation has been proven to be useful for enhancing the viability of lyophilized O. oeni. To explain the involved mechanisms, cell integrity, morphology and protein patterns of lyophilized O. oeni SD-2a were investigated with acid acclimation. After lyophilization, improvement of cell integrity and more extracellular polymeric substances (EPS) were observed in acid acclimated cells. Combined with GO and KEGG analysis, different abundant proteins were noticeably enriched in the carbohydrate metabolism process, especially amino sugar and nucleotide sugar metabolism. The most significant result was the over-expression of proteins participating in cell wall biosynthesis, EPS production, ATP binding and the bacterial secretion system. This result indicated the important role of acid acclimation on cell envelope properties. In addition, protein response to stress and arginine deiminase pathway were also proven to be over-expressed.

The phenolics profile of two new kiwifruit cultivars, Zespri® SunGold and Zespri® Sweet Green, were characterized and quantified for the first time using cyclic voltammetry, an electrochemical method, combined with HPLC. Results from the cyclic voltammetry revealed high correlations with those obtained from the spectrophotometry and HPLC methods, providing evidence to support the application of cyclic voltammetry as a rapid method in determining the phenolic profile and reducing power of kiwifruit extracts. Catechol-containing phenolics were identified as the major phenolic sub-class in the skins while flavonoids and phenolic acids were abundant in flesh of the tested cultivars. Epicatechin was the predominant phenolic compound and contributor to antioxidant capacity in all samples. Results also showed that SunGold and Sweet Green (both flesh and skin) exhibited significantly higher phenolic contents and antioxidant activities comparing with the well-established commercial ‘Hayward’ cultivar, indicating their commercial value and potential applications in food and nutraceuticals.

Here we presented a simple, rapid and label-free surface-enhanced Raman spectroscopy (SERS) based mapping method for the detection and discrimination of Salmonella enterica and Escherichia coli on silver dendrites. The sample preparation was first optimized to maximize sensitivity. The mapping method was then used to scan through the bacterial cells adsorbed on the surface of silver dendrites. The intrinsic and distinct SERS signals of bacterial cells were used as the basis for label-free detection and discrimination. The results show the developed method is able to detect single bacterial cells adsorbed on the silver dendrites with a limit of detection as low as 10(4) CFU mL(-1), which is two orders of magnitude lower than the traditional SERS method under the same experimental condition. The time needed for collecting a 225 points map was approximately 24 minutes. Moreover, the developed SERS mapping method can realize simultaneous detection and identification of Salmonella enterica subsp. enterica BAA1045 and Escherichia coli BL21 from a mixture sample using principle component analysis. Our results demonstrate the great potential of the label-free SERS mapping method to detect, identify and quantify bacteria and bacterial mixtures simultaneously.

This study explored the feasibility of degrading aflatoxin B1 (AFB1) in contaminated peanut meal by extrusion cooking. Peanut meal contaminated with AFB1 (35.8 ± 1.5 μg/kg) was extrusion-cooked in a twin-screw extruder and AFB1 was quantified by ELISA. The effects of barrel temperature, material moisture content, feed rate, and screw speed as well as their interactions on the reduction rate of AFB1 in peanuts meal were evaluated by Response surface methodology (RSM) to optimize the extrusion conditions. Under the optimal conditions (barrel temperature of 150 °C, material moisture content of 40 g/100 g, feed rate of 17 g/min, and screw speed of 152 rpm), 77.6 ± 2.2% of AFB1 was degraded. The barrel temperature and material moisture have significant influences (P

Oenococcus oeni (O. oeni) induces malolactic fermentation to improve wine quality. However, the molecular basis of mechanisms involved in its freeze-drying resistance remains unclear. In this work, O. oeni SD-2a without freeze-drying (No-FD), with freeze-drying (FD) and freeze-drying with monosodium glutamate (MSG) (FD-MSG) were investigated. Flow cytometry results showed severe cell damage in FD cells and less membrane transmissibility in FD-MSG cells. Meanwhile, extracellular polymeric substances (EPS) were detected in FD and FD-MSG cells by scanning electron microscopy. Bioinformatic analysis revealed that varying proteins were involved in carbon, lipid and nucleic acids metabolism, stress response, oxidoreductase activity and signal sensing. Among the identified proteins, the highlighted proteins were those involved in polysaccharides production and signal sensing for freeze-drying resistance and cyclopropane fatty acid metabolism for MSG addition.

The aim of this study was to evaluate the antibiotic susceptibility of lactic acid bacteria (LAB) and coagulase-negative staphylococci (CNS) strains isolated from naturally fermented smoked pork produced in Hunan, China. A total of 48 strains were isolated by selective medium and identified at the species level by 16S rRNA gene sequencing as follows: Staphylococcus carnosus (23), Lactobacillus plantarum (12), Lactobacillus brevis (10), Lactobacillus sakei (1), Weissella confusa (1), and Weissella cibaria (1). All strains were typed by RAPD-PCR, and their susceptibility to 15 antibiotics was determined and expressed as the minimum inhibitory concentration (MIC) using agar dilution method. High resistance to penicillin G, streptomycin, gentamycin, vancomycin, chloramphenicol, norfloxacin, ciprofloxacin, kanamycin, and neomycin was found among the isolates. All the strains were sensitive to ampicillin, while the susceptibility to tetracycline, oxytetracycline, erythromycin, lincomycin, and roxithromycin varied. The presence of relevant resistance genes was investigated by PCR and sequencing, with the following genes detected: str(A), str(B), tet(O), tet(M), ere(A), and catA. Eleven strains, including 3 S. carnosus, 6 L. plantarum, and 2 L. brevis, harbored more than 3 antibiotic resistance genes. Overall, multiple antibiotic resistance patterns were widely observed in LAB and S. carnosus strains isolated from Hunan smoked pork. Risk assessment should be carried out with regard to the safe use of LAB and CNS in food production.We evaluated the antibiotic resistance of lactic acid bacteria and coagulase-negative staphylococci strains isolated from Chinese naturally fermented smoked pork. Our results may provide important data on establishing breakpoint standards for LAB and CNS and evaluating the safety risk of these strains for commercial use.

The accumulation of selenium (Se) by Lactobacillus delbrueckii ssp. bulgaricus (Lb) and Streptococcus thermophilus (St) at the different cultivation conditions, including initial pH, inoculum dose (%), and temperature (°C), was investigated in this work. Se enrichment efficiency was optimized using the Design-Expert software for response surface methodology on a basis of single-factor experiment. The antioxidant activities of Se-enriched Lactic acid bacteria (LAB) were also investigated. The qualitative analysis of Se-enriched LAB was performed by FT-IR spectra. The cell morphology and chemical element components were measured by a scanning electron microscope (SEM) equipped with energy-dispersive X-ray spectroscopy. The results indicated that the optimum initial pH, inoculum doses, and temperatures of Lb and St were 5.96, 6.73%, 33.24 °C, and 6.37, 6%, 40 °C, respectively. Under the optimal conditions, the ratios of Se enrichment reached 94.34% for Lb and 97.05% for St. Furthermore, Se-enriched LAB enhanced scavenging rates on DPPH, ABTS free radical, and also heightened reducing activity. The FT-IR results showed that the two Se-enriched strains had similar characteristic absorption peaks, which were further demonstrated that both Se biomasses had the same carbonyl, carboxyl, and hydroxyl groups. Elemental selenium nanoparticles were verified around cell surfaces of Se-enriched LAB, which implied that both strains had detoxification ability when grown in liquid media containing selenite.