Effect of Different Sterilization Methods on the Microbial and Physicochemical Changes in Prunus mume Juice during Storage.

This study evaluated the pasteurization (P), ozone (O3), ultrasonic (US), and high-hydrostatic-pressure (HHP) sterilization approaches for processing of Prunus mume regarding browning factors and microorganisms, compared with non-sterilization (control check, CK) treatment. The microorganisms (total bacterial count and fungi and yeast count) in the juice were identified after different sterilization techniques, while the quality parameter changes (degree of browning, color measurements, total phenolic content, reducing sugar, ascorbic acid, 5-hydroxymethyl furaldehyde (5-HMF), amino acid nitrogen, total soluble solids (TSS), pH value) were investigated. The results indicate that P and HHP treatment reduced non-enzymatic browning while substantially impacting the color measurements, TSS, and pH, while the sterilization effect was remarkable, with a rate exceeding 90%. Furthermore, the Prunus mume juices treated with P and HHP sterilization were used as the objects, and the CK group was used as the control group. They were placed at 4 °C, 25 °C and 37 °C, respectively, and stored in dark for 15 d. Sampling and determination were carried out on 0, 3, 6, 9, 12, and 15 d, respectively. M-&-Y (molds and yeasts) were not detected in the late storage period, and no obvious microbial growth was observed during storage, indicating that P and HHP treatments could ensure the microbial safety of Prunus mume juice. P- and HHP- treated Prunus mume juice has better quality and low temperature storage is beneficial for maintaining the quality of Prunus mume juice. Therefore, P treatment or HHP treatment combined with low temperature storage could achieve a more ideal storage effect. Overall, this study conclusively established that P and HHP methods were suitable for sterilizing Prunus mume juice. These techniques minimally affected overall product quality while better maintaining the quality parameters than the untreated juice samples and those exposed to O3 and US treatment.

Fluorescence detection of Staphylococcus aureus using vancomycin functionalized magnetic beads combined with rolling circle amplification in fruit juice.

Staphylococcus aureus is a common foodborne pathogen that can cause a suppurative infection after eating contaminated food. Detection of S. aureus plays an important role in the food industry. In this study, a strategy for the detection of S. aureus using magnetic separation (MS) technology combined with rolling circle amplification (MS-RCA) was proposed. The strategy used antibiotics to capture bacteria and employed RCA products as signal output probes. Vancomycin (Van), as a commonly used antibiotic, can recognize peptidoglycan on the cell wall of Gram-positive bacteria and can effectively identify target bacteria. Therefore, we prepared BSAylated-Van functionalized magnetic beads (Van-MBs) for the pre-enrichment of S. aureus. To ensure the selectivity of this method, we used biotin-pig IgG to bind S. aureus. In addition, to amplify the output signal of the MS-RCA strategy, we introduced streptavidin (SA) and successfully obtained the Van-MBs@S. aureus@biotin-pig IgG@SA@biotin-RCA probe complex and used the biotin-avidin-system (BAS) by combining magnetic separation technology and RCA technology to realize the enrichment and specific detection of S. aureus. Furthermore, by optimizing the experimental conditions such as the magnetic separation time and the amount of Van-MBs, the detection performance of this method was improved. Under the optimal conditions, the detection limit of this method for S. aureus was 3.3 × 102 CFU/mL in fruit juice, and it was less affected by other bacteria.

Antibacterial activity of Ricinus communis plant extract against antibiotic resistant Helicobacter pylori and Gluconobacter oxydans isolated from fresh apple juices samples.

Bacteria were isolated from samples of Fresh Apple juices from shops of three different localities of Lahore. Analysis of samples from Liberty, Anarkali and Yateem khana Markets show different levels of contamination. There were pathogenic and non-pathogenic bacteria in all samples and were identified by the morphological and biochemical tests. Most of the plasmids of pathogenic bacteria were 4kb in their molecular size. Ribotyping of 16S ribosomal RNA gene sequencing was done to confirm Helicobacter pylori strain and Gluconobacter oxydans. The highest sensitivity of 210mm was shown by Enterobacter sp. against Aztheromysine disk (15µg) while Micrococcus sp. was highly resistant against all of the Antibiotics applied. The antibiotic resistance of pathogenic bacteria was also checked against Ricinus communis plant's extracts, all isolated bacterial pathogens were resistant but only, E.coli was inhibited at 300µl of the extracts. Presence of pathogenic bacteria in Apple juice samples was due to contamination of sewage water in drinking water while some of these pathogenic bacteria came from Apple's tree and other from store houses of fruits.

Inhibition of LPMOs by Fermented Persimmon Juice.

Fermented persimmon juice, Kakishibu, has traditionally been used for wood and paper protection. This protective effect stems at least partially from inhibition of microbial cellulose degrading enzymes. The inhibitory effect of Kakishibu on lytic polysaccharide monooxygenases (LPMOs) and on a cocktail of cellulose hydrolases was studied, using three different cellulosic substrates. Dose dependent inhibition of LPMO activity by a commercial Kakishibu product was assessed for the well-characterized LPMO from Thermoascus aurantiacus TaAA9A, and the inhibitory effect was confirmed on five additional microbial LPMOs. The model tannin compound, tannic acid exhibited a similar inhibitory effect on TaAA9A as Kakishibu. It was further shown that both polyethylene glycol and tannase can alleviate the inhibitory effect of Kakishibu and tannic acid, indicating a likely mechanism of inhibition caused by unspecific tannin-protein interactions.

Lactiplantibacillus plantarum CCMA 0743 and Lacticaseibacillus paracasei subsp. paracasei LBC-81 metabolism during the single and mixed fermentation of tropical fruit juices.

Fruit juices have shown promising results as new probiotic carriers. This study aimed to evaluate acerola, jelly palm, and passion fruit juices as substrates for fermentation using Lactiplantibacillus plantarum CCMA 0743 and Lacticaseibacillus paracasei LBC-81 in single and mixed cultures. First, the juices were evaluated as substrate and selected based on bacterial growth performance during fermentation. Afterward, the impact of fermentation on sugars, organic acids, and bioactive compounds was also appraised. Phytochemical modification of three different juices fermented by lactic acid bacteria at 37 °C/24 h was evaluated. After 18 h of fermentation, passion fruit juice showed higher cell viable counts of single and mixed L. plantarum CCMA 0743 culture, above 9.00 Log CFU/mL, and pH between 4.07 and 4.10. Sugars consumption and organic acid production were influenced by juice composition and culture used. The mixed culture reduced the total sugars in the passion fruit juice by approximately 53.0% (8.51 g/L). Lactic acid was the main product of the sugars fermentation, with higher concentrations detected in passion fruit juice (8.39-11.23 g/L). Bioactive compounds were analyzed on the selected substrate. The fermentative process reduced antioxidant activity and carotenoid content. However, single L. plantarum CCMA 0743 culture increased the yellow flavonoid content of passion fruit juice by approximately 3.0 µg/mL. L. plantarum CCMA 0743 showed high and suitable cell, viable counts, to claimed probiotic products, increasing bioactive compounds in passion fruit juice. Therefore, this strain and passion fruit substrate showed attractive potential to produce alternative and functional fermented fruit beverages.

Development of a pulse-induced electrochemical biosensor based on gluconamide for Gram-negative bacteria detection.

Pathogenic bacteria can cause the outbreaks of disease and threaten human health, which stimulates the development of advanced detection techniques. Herein, a specific and sensitive electrochemical biosensor for Gram-negative bacteria was established based on the conductive polymer with artificial muscle properties.  The effective recognition was achieved through the specific carbohydrate-carbohydrate interaction between gluconamide and lipopolysaccharide.  The application of impulse voltage enhances the efficiency of recognition and shortens the detection time through the temporary deformation of the electrode surface, with a limit of detection (LOD)  of 1 × 100 CFU/mL and a linear range of 1 × 100 - 1 × 106 CFU/mL for Escherichia coli (E. coli). In addition  to the merits of low cost, high efficiency, and rapidity,  the developed label-free electrochemical biosensor can also be applicable for other Gram-negative bacteria, owning promising potential in the application of portable devices and paving a potential way for the construction of electrochemical biosensors.

An innovative strategy to rapidly inactivate 8.2-log Enterococcus faecalis in fresh pineapple juice using cold atmospheric plasma.

Enterococcus faecalis is a life-threatening bacterium that resists high levels of antibiotics or chemical preservatives. In this study, we aimed to investigate the inactivation of E. faecalis in fresh pineapple juice (FPJ) with two different cold atmospheric plasmas (CAP) reinforced by H2O2/H2O cold vapor: a plasma jet and a surface dielectric barrier discharge (SDBD). CAP treatments for 300 s with plasma jet and 420 s with SDBD caused an 8.2 log reduction of E. faecalis. The concentration of reactive oxygen and nitrogen species induced in FPJ by plasmas was also evaluated. In terms of quality attributes of FPJ, no noticeable color changes (ΔE) were observed. Furthermore, a trifle of loss of organic content such as sugars and organic acids was observed after treatments. These results suggest that our rapid CAP strategy effectively inactivated E. faecalis in FPJ with no change of color and negligible effects on other physicochemical properties.

One-pot synthesis of magnetic self-assembled carrageenan-ε-polylysine composites: A reusable and effective antibacterial agent against Alicyclobacillus acidoterrestris.

Epsilon-polylysine (EPL) is a cationic polymer with broad antibacterial activity. Base on this property, anionic carrageenan (CG) was self-assembled with EPL and introduced into the one-pot coprecipitation process to fabricate the magnetic Fe3O4-CG-EPL composites. To demonstrate the successful synthesis of composites, characterization techniques including XRD, FT-IR, TEM, XPS and VSM were employed. The prepared composites exhibited effective antibacterial activity against Alicyclobacillus acidoterrestris with the minimum inhibitory concentration of 10.0 mg mL-1. Investigations into deciphering the antibacterial mechanism demonstrated that the presence of Fe3O4-CG-EPL caused irreversible damage to the cell membrane and serious leakage of intracellular protein, resulting in the inactivation of bacteria. The aim of this work is to develop a new effective control method for A. acidoterrestris in food field.

Fermentation of Jamaican Cherries Juice Using Lactobacillus plantarum Elevates Antioxidant Potential and Inhibitory Activity against Type II Diabetes-Related Enzymes.

Jamaican cherry (Muntinga calabura Linn.) is tropical tree that is known to produce edible fruit with high nutritional and antioxidant properties. However, its use as functional food is still limited. Previous studies suggest that fermentation with probiotic bacteria could enhance the functional properties of non-dairy products, such as juices. In this study, we analyze the metabolite composition and activity of Jamaican cherry juice following fermentation with Lactobacillus plantarum FNCC 0027 in various substrate compositions. The metabolite profile after fermentation was analyzed using UPLC-HRMS-MS and several bioactive compounds were detected in the substrate following fermentation, including gallic acid, dihydrokaempferol, and 5,7-dihydroxyflavone. We also found that total phenolic content, antioxidant activities, and inhibition of diabetic-related enzymes were enhanced after fermentation using L. plantarum. The significance of its elevation depends on the substrate composition. Overall, our findings suggest that fermentation with L. plantarum FNCC 0027 can improve the functional activities of Jamaican cherry juice.

Selenium bio-enrichment of Mediterranean fruit juices through lactic acid fermentation.

This work was carried out to elaborate selenium (Se) bio-enriched fermented Mediterranean fruit juices. To this purpose, pomegranate and table red grape juices were added with sodium selenite (Na2SeO3) and fermented by Levilactobacillus brevis CRL 2051 and Fructobacillus tropaeoli CRL 2034 individually or combined. To better evaluate the effect of selenite addition and starter strain inoculums on the total bacterial community of the fruit juices, fermentation trials were performed with raw and pasteurized fruit juices. No statistical significant differences were observed for total mesophilic microorganisms (TMM) and rod-shaped lactic acid bacteria (LAB) levels among raw and pasteurized juices inoculated with the starter strains, while significant differences between those juices with and without selenite were registered. LAB cocci, Pseudomonadaceae and yeasts were detected only for the raw juice preparations. The dominance of L. brevis CRL 2051 and F. tropaeoli CRL 2034 was confirmed by randomly amplified polymorphic DNA (RAPD)-PCR analysis. After fermentation, pH dropped for all inoculated trials and control raw juices. The soluble solid content (SSC) levels of the raw juices were higher than the corresponding pasteurized trials. The thermal treatment affected consistently yellowness of grape juice trials and redness of pomegranate juices. No microbial Se accumulation was registered for pomegranate juices, while F. tropaeoli CRL 2034 accumulated the highest amount of Se (65.5 µg/L) in the grape juice. For this reason, only trials carried out with raw grape juices were investigated by metagenomics analysis by Illumina MiSeq technology. Non-inoculated grape juices were massively fermented by acetic acid bacteria while Fructobacillus and Lactobacillus (previous genus name of Levilactobacillus) represented the highest operational taxonomy units (OTUs) relative abundance % of the trials inoculated with the starter strains as confirmed by this technique.

Effect of lactic acid fermentation of watermelon juice on its sensory acceptability and volatile compounds.

Fermentation increases food shelf-life but is characterized by changes that affect product's perception. Watermelon juice was fermented with Lactobacillus plantarum (WJ-LP), L. rhamnosus (WJ-LR), L. casei (WJ-LC), L. brevis (WJ-LB) and Pediococcus pentosaceus (WJ-PP). Their sensory characteristics and volatile compounds were investigated by consumers and Headspace Solid Phase Microextraction integrated with Gas Chromatography-Mass Spectrometry, respectively. WJ-PP and WJ-LB were most liked and perceived with 'watermelon-flavor', 'natural taste', 'sweet' and 'watermelon-color' while WJ-LC, WJ-LP and WJ-LR were least liked and perceived as 'sour', 'bitter', 'off-flavor', 'aftertaste' and 'intense-flavor'. Fifty-four volatiles were identified. After fermentation, alcohols, ketones, monoterpenes, acids, and furans increased while aldehydes and alkanes decreased. Lactic acid fermentation introduced 4-decanone and 2,3-butanedione in WJ-LB, WJ-LC, WJ-LP and WJ-LR, however, heptanal, 2-heptenal, 2,6-nonadienal, 2-decenal, and 2,4-decadienal in WJ-LC, heptanal, 2-hexenal, 2-heptenal, 2,6-nonadienal, 2-decenal and octanal in WJ-LR and 2,6-dimethyl-2,6-octadiene in WJ-LP disappeared. Juice sensory profiles were associated with their volatile compounds.

Antibacterial Activity of Caffeic Acid Combined with UV-A Light against Escherichia coli O157:H7, Salmonella enterica Serovar Typhimurium, and Listeria monocytogenes.

The aim of this study was to evaluate the antibacterial activity of caffeic acid (CA), which is a natural polyphenol, combined with UV-A light against the representative foodborne bacteria Escherichia coli O157:H7, Salmonella enterica serovar Typhimurium, and Listeria monocytogenes. Data regarding the inactivation of these bacteria and its dependence on CA concentration, light wavelength, and light dose were obtained. E. coli O157:H7 and Salmonella Typhimurium were reduced to the detection limit when treated with 3 mM CA and UV-A for 3 J/cm2 and 4 J/cm2, respectively, and 5 J/cm2 treatment induced 3.10 log reduction in L. monocytogenes. To investigate the mechanism for inactivation of Salmonella Typhimurium and L. monocytogenes, measurement of polyphenol uptake, membrane damage assessment, enzymatic activity assay, and transmission electron microscopy (TEM) were conducted. It was revealed that CA was significantly (P < 0.05) absorbed by bacterial cells, and UV-A light allowed a higher uptake of CA for both pathogens. Additionally, CA plus UV-A treatment induced significant (P < 0.05) cell membrane damage. In the enzymatic activity assay, the activities of both pathogens were reduced by CA, and a greater reduction occurred by use of CA plus UV-A. Moreover, transmission electron microscopy (TEM) images indicated that CA plus UV-A treatment notably destroyed the intercellular structure. In addition, antibacterial activity was also observed in commercial apple juice, which showed results similar to those obtained from phosphate-buffered saline (PBS), resulting in a significant (P < 0.05) reduction for all three pathogens without any changes in color parameters (L*, a*, and b*), total phenolic compounds, and DPPH (2,2-diphenyl-1-picrylhydrazyl) free radical scavenging activity. IMPORTANCE Photodynamic inactivation (PDI), which involves photoactivation of a photosensitizer (PS), is an emerging field of study, as it effectively reduces various kinds of microorganisms. Although there are several PSs that have been used for PDI, there is a need to find naturally occurring PSs for safer application in the food industry. Caffeic acid, a natural polyphenol found in most fruits and vegetables, has recently been studied for its potential to act as a novel photosensitizer. However, no studies have been conducted regarding its antibacterial activity depending on treatment conditions and its antibacterial mechanism. In this study, we closely examined the effectiveness of caffeic acid in combination with UV-A light for inactivating representative foodborne bacteria in liquid medium. Therefore, the results of this research are expected to be utilized as basic data for future application of caffeic acid in PDI, especially when controlling pathogens in liquid food processing.

Quality attributes optimization of orange juice subjected to multi-frequency thermosonication: Alicyclobacillus acidoterrestris spore inactivation and applied spectroscopy ROS characterization.

This is the first time to investigate the synergistic inactivation effect and mechanism of multifrequency ultrasound (MTUS) on A. acidoterrestris (AAT) vegetative cells and spores, nutrients and enzymes of orange juice. The optimized results of MTUS (using Box Behnken design- surface responsemethodology) and further comparison with different mode of ultrasound (mono-and multi-frequency) revealed that 20/40 kHz, 24 min and 64 °C were the best optimum results. The AAT spores and vegetative cells were inactivated by 2 and 4 logs, respectively, without deteriorating orange juice contents. In addition, AAT inactivation indicated an inversely proportional relationship with ROS production. FT-IR and UV-Vis spectroscopy characterization confirmed the existence of ROS in treated orange juice and LF-NMR analysis confirmed the inactivation of AAT spores. The findings illustrated the successfully used dual-frequency ultrasound technology for fruit beverages, promoting beneficial changes in physical properties without any significant effects on the quality of ascorbic acid.

Isolation and identification of guaiacol producing Alicyclobacillus fastidiosus strains from orchards in Poland.

The genus Alicyclobacillus comprises a group of Gram-positive, thermo-acidophilic bacteria that are capable of producing highly resistant endospores during unfavorable environmental conditions. The members of this genus inhabit natural environments, including hot springs and soils. The main reason behind the spoilage of final commercial fruit products by Alicyclobacillus is the contamination of fruits with soil at the time of harvesting. Some of the Alicyclobacillus species, including Alicyclobacillus acidoterrestris, are categorized as spoilage bacteria due to their ability to produce off-flavor compounds (e.g., guaiacol and halophenols) that adversely affect the taste and aroma of beverages. In our study, Alicyclobacillus species were isolated from Polish orchard soils and fruits and were subjected to 16S rDNA sequencing. The results of the analysis showed that the isolated strains belonged to A. acidoterrestris and Alicyclobacillus fastidiosus species. All the three isolated strains of A. fastidiosus (f1, f2, f3) exhibited similar morphological and biochemical properties as the strain described in the literature. However, these isolated strains were able to produce guaiacol at temperatures of 20°C, 25°C, and 45°C. Thus, the strains of A. fastidiosus discovered in the present study can be included in the group of spoilage species as they possessed the gene responsible for the production of guaiacol.

Cloudy Apple Juice Fermented by Lactobacillus Prevents Obesity via Modulating Gut Microbiota and Protecting Intestinal Tract Health.

Obesity and hyperglycemia are two serious chronic diseases that are increasing in incidence worldwide. This research aimed to develop a fermented cloudy apple juice with good hyperglycemia intervention activities. Here, cloudy apple juice (CAJ), cloudy apple juice rich in polyphenols (CAJP) and fermented cloudy apple juice rich in polyphenols (FCAJP) were prepared sequentially, and then the effects of the three apple juices on weight, lipid level, gut microbiota composition and intestinal tract health were evaluated for obese mice induced by a high-fat diet. The research findings revealed that the FCAJP showed potential to inhibit the weight gain of mice, reduce fat accumulation, and regulate the blood lipid levels of obese mice by decreasing the ratio of the Firmicutes/Bacteroidotas, improving the Sobs, Ace, and Chao indexes of the gut microbiota and protecting intestinal tract health. In addition, the FCAJP augmented the abundance of Akkermansia and Bacteroides, which were positively related to SCFAs in cecal contents. This study inferred that FCAJP could be developed as a healthy food for preventing obesity and hyperglycemia.

High pressure homogenization inactivation of Escherichia coli and Staphylococcus aureus in phosphate buffered saline, milk and apple juice.

High pressure homogenization (HPH) offers new opportunities for food pasteurization/sterilization. Escherichia coli and Staphylococcus aureus suspended in phosphate buffered saline (PBS) buffer, milk and apple juice at initial concentration of ~106 log10 CFU per ml were subjected to HPH treatments up to 200 MPa with inlet temperatures at 4-40°C. After HPH at 200 MPa with the inlet temperature at 40°C, the count of E. coli suspended in PBS, milk and apple juice reduced by 3·42, 3·67 and 3·19 log10 CFU per ml respectively while the count of S. aureus decreased by 2·21, 1·02 and 2·33 log10 CFU per ml respectively suggesting that S. aureus was more resistant. The inactivation data were well fitted by the polynomial equation. Milk could provide a protective effect for S. aureus against HPH. After HPH at 200 MPa with the inlet temperature at 20°C, the cell structure of E. coli was destroyed, while no obvious damages were found for S. aureus.

Anti-listerial properties of chemical constituents of Eruca sativa (rocket salad): From industrial observation to in vitro activity.

The frequency of foodborne outbreaks epidemiologically associated with Listeria monocytogenes in fresh produce has increased in recent years. Although L. monocytogenes may be transferred from the environment to vegetables during farming, contamination of food products most commonly occurs in food processing facilities, where L. monocytogenes has the ability to establish and persist on processing equipment. The current study was undertaken to collect data on the occurrence of L. monocytogenes and the identity of the endogenous microbiota in a fresh produce processing facility, for which information has remained scarce. L. monocytogenes was not detected in the facility. Experiments simulating conditions in the processing environment were performed, including examination of bacterial growth in nutrients based on vegetables (salad juice) compared to in other types of nutrients (fish, meat). Results showed that the endogenous microbiota (dominated by Pseudomonas) grew well in iceberg lettuce and rocket salad juice at low temperatures, while growth inhibition of L. monocytogenes was observed, particularly in rocket salad juice. The anti-listerial activity in rocket salad juice was retained in a polar chromatographic fraction containing several metabolites. Characterization of this active fraction, using LC-MS/MS, led to identification of 19 compounds including nucleosides and amino acids. Further work is necessary to determine the molecular mechanism responsible for the inhibitory activity of rocket salad constituents. The study nevertheless suggests that the available nutrients, as well as a low temperature (3 °C) and the in-house bacterial flora, may influence the prevalence of L. monocytogenes in fresh produce processing facilities.

Coating Lacticaseibacillus rhamnosus GG in Alginate Systems: an Emerging Strategy Towards Improved Viability in Orange Juice.

Fruit juices are successfully proposed as suitable probiotic vehicles, but researchers' efforts should be developed to avoid effects of bacteria overgrowing on sensory and nutritional cues of final products and to preserve viability of probiotic bacteria during storage. In the present study, encapsulation of Lacticaseibacillus rhamnosus GG strain in alginate systems was performed through ionotropic gelation technology. The alginate systems were optimized by using Box-Behnken Design to investigate the influence of three independent variables at three different levels: particle mean size and polydispersity index. The optimized probiotic-loaded alginate particles were added to orange juice samples. The viability of the probiotic strain, both as free and microencapsulated, was evaluated in orange juice stored at 5°C for 35 days. Morphology and size of probiotic-loaded alginate particles were found suitable for incorporation into juice. TEM analysis revealed that unloaded systems were clustered as nanoparticles (CL_NP), while the loaded sample appeared as a coated system (Coated_LGG). Microbiological evaluation revealed that the encapsulation assured the survival of Coated_LGG, with a reduction of less than 1-unit log in cellular density after 35 days of refrigerated storage in orange juice. Results indicated that the encapsulated bacteria did not affect the macroscopic properties neither the microbiological characteristic of orange juice; thus, it can be proposed as functional food.

The responses of Salmonella enterica serovar Typhimurium to vanillin in apple juice through global transcriptomics.

Salmonella enterica serovar Typhimurium can survive some extreme environment in food processing, and vanillin generally recognized as safe is bactericidal to pathogens. Thus, we need to explore the responses of S. Typhimurium to vanillin in order to apply this antimicrobial agent in food processing. In this study, we exposed S. Typhimurium to commercial apple juice with/without vanillin (3.2 mg/mL) at 45 °C for 75 min to determine the survival rate. Subsequently, the 10-min cultures were selected for transcriptomic analysis. Using high-throughput RNA sequencing, genes related to vanillin resistance and their expression changes of S. Typhimurium were identified. The survival curve showed that S. Typhimurium treated with vanillin were inactivated by 5.5 log after 75 min, while the control group only decreased by 2.3 log. Such a discrepancy showed the significant antibacterial effect of vanillin on S. Typhimurium. As a result, 265 differentially expressed genes (DEGs) were found when coping with vanillin, among which, 225 showed up-regulation and 40 DEGs were down-regulated. Treated with vanillin, S. Typhimurium significantly up-regulated genes involved in cell membrane, acid tolerance response (ATR) and oxidative stress response, cold shock cross-protection, DNA repair, virulence factors and some key regulators. Firstly, membrane-related genes, including outer membrane (bamE, mepS, ygdI, lolB), inner membrane (yaiY, yicS) and other proteins (yciC, yjcH), were significantly up-regulated because of the damaged cell membrane. Then, up-regulated proteins associated with arginine synthesis (ArgABCDIG) and inward transportation (ArtI, ArtJ, ArtP and HisP), participated in ATR to pump out the protons inside the cell in this scenario. Next, superoxide stress response triggered by vanillin was found to have a significant up-regulation as well, which was controlled by SoxRS regulon. Besides, NADH-associated (nuoA, nuoB, nuoK, nadE, fre and STM3021), thioredoxin (trxA, trxC, tpx and bcp) and glutaredoxin (grxC and grxD) DEGs led to the increase of the oxidative stress response. Cold shock proteins such as CspA and CspC showed an up-regulation, suggesting it might play a role in cross-protecting S. Typhimurium from vanillin stress. Furthermore, DEGs in DNA repair and virulence factors, including flagellar assembly, adhesins and type III secretion system were up-regulated. Some regulators like fur, rpoE and csrA played a pivotal role in response to the stress caused by vanillin. Therefore, this study sounds an alarm for the risks caused by stress tolerance of S. Typhimurium in food industry.

Use of non-Saccharomyces yeasts in cider fermentation: Importance of the nutrients addition to obtain an efficient fermentation.

The isolation of autochthonous yeast species presents a good strategy to select new microorganisms for developing an adequate inoculum to carry out fermentations and generate representative products of the cider production zone. However, non-Saccharomyces yeasts have been considered to have low capacity to carry out a complete fermentation as Saccharomyces cerevisiae. In this work, five autochthonous yeasts from a cider fermentation process were isolated and identified as Saccharomyces cerevisiae, Kluyveromyces marxianus, Pichia membranaefaciens, P. kluyveri and Zygosaccharomyces rouxii. A series of fermentations were developed at laboratory level, using each species individually and it was observed that only S. cerevisiae was able to finish the process. K. marxianus consumed less than 50% of the sugars; P. kluyveri and Z. rouxii consumed less than 70% and P. membranaefaciens consumed more than 90% but the yield (ethanol produced for sugar consumed (YP/S)) was 0.39. Nevertheless, the addition of magnesium, zinc and nitrogen increased the fermentative capacity of almost all species: K. marxianus, Z. rouxii and P. kluyveri, showed an increase in ethanol production when nutrients were added, obtaining more than 80 g/L of ethanol, and showing that those nutrients are necessary to complete the fermentation. This work describes the potential use of different non-Saccharomyces species to carry out fermentation of apple juice and highlights the importance of certain nutrients to enable an efficient alcoholic fermentation and the generation of desirable volatile compounds for cider production.