Mechanism of Different Stereoisomeric Astaxanthin in Resistance to Oxidative Stress in Caenorhabditis elegans

As a potent antioxidant in human diet, astaxanthin (AST) may play important roles in alleviating oxidative stress‐driven adverse physiological effects. This study examined the effects of different stereoisomers of AST in protecting Caenorhabditis elegans from chemically induced oxidative stress. Three stereoisomers of AST investigated herein included 3S,3´S (S) AST, 3R,3´R (R) AST, and a statistical mixture (S: meso: R = 1:2:1) (M) AST. Under paraquat‐induced oxidative conditions, all three types of AST significantly enhanced survival rate of C. elegans. The accumulation levels of ROS in the worms were reduced by 40.12%, 30.05%, and 22.04% by S, R, and M AST, respectively (P < 0.05). Compared with R and M AST, S significantly enhanced the expression levels of SOD‐3. The results of RNA‐Seq analysis demonstrated that AST protected C. elegans from oxidative damage potentially by modulating genes involved in the insulin/insulin‐like growth factor (IGF) signaling (IIS) pathway and the oxidoreductase system. It is noteworthy that different stereoisomers of AST showed different effects on the expression levels of various genes related with oxidative stress. This study revealed important information on the in vivo antioxidative effects of AST stereoisomers, which might provide useful information for better utilization of AST.     

A comparison of gene expression of Listeria monocytogenes in vitro and in the soft cheese Crescenza

This study focuses on the expression analysis of virulence and stress response genes (plcA, hly, iap and sigB) in 11 different strains of Listeria monocytogenes. The first step of this study considered an in vitro analysis in synthetic laboratory medium. According to statistical analysis, significant differences emerged for genes sigB and plcA. Subsequently, a soft cheese at two temperatures (4 and 12 °C) and for different times (24 and 48 h) was evaluated. Significant expression differences emerged for the condition at 12 °C after 48 h. No association could be observed between gene expression profile and origin of the different strains.     

Stability and Antimicrobial Activity of Allyl Isothiocyanate during Long-Term Storage in an Oil-in-Water Emulsion

Abstract: This study investigated the stability and antimicrobial activity of allyl isothiocyanate (AITC) in medium chain triglyceride (MCT) or soybean oil (SBO) dispersed in an oil-in-water (o/w) system during long-term storage. Oil type, content, and oxidative stability affect the stability and antimicrobial activity of AITC during storage. High oil content is favorable for AITC stability in the emulsion. Notably, AITC with MCT is more stable than AITC with SBO with the same oil content. Consequently, AITC with MCT is more effective than AITC with SBO in inhibiting G(−) bacteria (E. coli O157:H7, Salmonella enterica, and Vibrio parahaemolyticus) and G(+) bacteria (Staphylococcus aureus and Listeria monocytogenes).     

Construction of Listeria monocytogenes Mutants with In‐Frame Deletions in the Phosphotransferase Transport System (PTS) and Analysis of Their Growth under Stress Conditions

Listeria monocytogenes is a foodborne pathogen that is difficult to eliminate due to its ability to survive under different stress conditions such as low pH and high salt. To better control this pathogen in food, it is important to understand its survival mechanisms under these stress conditions. LMOf2365_0442, 0443, and 0444 encode for phosphotransferase transport system (PTS) permease (fructose‐specific IIABC components) that is responsible for sugar transport. LMOf2365_0445 encodes for glycosyl hydrolase. These genes were induced by high pressure and inhibited under salt treatments; therefore, we hypothesized that genes encoding these PTS proteins may be involved in general stress responses. To study the function of these genes, deletion mutants of the PTS genes (LMOf2365_0442, LMOf2365_0443, and LMOf2365_0444) and the downstream gene LMOf2365_0445 were created in L. monocytogenes strain F2365. These deletion mutants were tested under different stress conditions. The growth of ?LMOf2365_0445 was increased under nisin (125 μg/mL) treatments compared to the wild‐type (P < 0.01). The growth of ?LMOf2365_0442 in salt (brain–heart infusion medium with 5% NaCl) was significantly increased (P < 0.01), and ?LMOf2365_0442 showed increased growth under acidic conditions (pH 5.0) compared to the wild‐type (P < 0.01). The results from phenotypic arrays demonstrated that some of these mutants showed slightly slower growth under different carbon sources and basic conditions. The results indicate that deletion mutants ?LMOf2365_0442 and ?LMOf2365_0445 were more resistant to multiple stress conditions compared to the wild‐type, suggesting that they may contribute to the general stress response in L. monocytogenes. An understanding of the growth of these mutants under multiple stress conditions may assist in the development of intervention strategies to control L. monocytogenes in food.     

Analysis of the role of the Listeria monocytogenes F0F1-ATPase operon in the acid tolerance response

As little is known about the genes involved in the induction of an acid tolerance response in Listeria monocytogenes, the role of the F₀F₁-ATPase was analyzed as a consequence of its role in the acid tolerance of a number of other bacteria and its conserved nature. It was found that acid adapted cells treated with N,N′-dicyclohexylcarbodiimide (DCCD) exhibited greatly enhanced sensitivity to low pH stress. Degenerate primers were designed to amplify and sequence a portion of the atpD gene. Subsequently, a PCR product from atpA to atpD was identified. While we were unable to create a deletion in the atpA gene, the plasmid pORI19 was inserted in a region between atpA and atpG to reduce, rather than eliminate, expression of the downstream genes. As expected this mutant displayed enhanced resistance to neomycin and exhibited slower growth than the wild type strain. This mutant could still induce an acid tolerance response and remained susceptible to DCCD treatment, but its relative acid sensitivity was difficult to assess as a consequence of its slow growth.     

Predictive Modeling for Growth of Non‐ and Cold‐adapted Listeria monocytogenes on Fresh‐cut Cantaloupe at Different Storage Temperatures

The aim of this study was to determine the growth kinetics of Listeria monocytogenes, with and without cold‐adaption, on fresh‐cut cantaloupe under different storage temperatures. Fresh‐cut samples, spot inoculated with a 4‐strain cocktail of L. monocytogenes (～3.2 log CFU/g), were exposed to constant storage temperatures held at 10, 15, 20, 25, or 30 °C. All growth curves of L. monocytogenes were fitted to the Baranyi, modified Gompertz, and Huang models. Regardless of conditions under which cells grew, the time needed to reach 5 log CFU/g decreased with the elevated storage temperature. Experimental results showed that there were no significant differences (P > 0.05) in the maximum growth rate k (log CFU/g h^?1) and lag phase duration λ (h) between the cultures of L. monocytogenes with or without previous cold‐adaption treatments. No distinct difference was observed in the growth pattern among 3 primary models at various storage temperatures. The growth curves of secondary modeling were fitted on an Arrhenius‐type model for describing the relationship between k and temperature of the L. monocytogenes on fresh‐cut cantaloupe from 10 to 30 °C. The root mean square error values of secondary models for non‐ and cold‐adapted cells were 0.018, 0.021, and 0.024, and 0.039, 0.026, and 0.017 at the modified Gompertz, Baranyi, and Huang model, respectively, indicating that these 3 models presented the good statistical fit. This study may provide valuable information to predict the growth of L. monocytogenes on fresh‐cut cantaloupes at different storage conditions.     

Discrimination of the Lactobacillus acidophilus group using sequencing, species-specific PCR and SNaPshot mini-sequencing technology based on the recA gene

BACKGROUND: To clearly identify specific species and subspecies of the Lactobacillus acidophilus group using phenotypic and genotypic (16S rDNA sequence analysis) techniques alone is difficult. The aim of this study was to use the recA gene for species discrimination in the L. acidophilus group, as well as to develop a species‐specific primer and single nucleotide polymorphism primer based on the recA gene sequence for species and subspecies identification. RESULTS: The average sequence similarity for the recA gene among type strains was 80.0%, and most members of the L. acidophilus group could be clearly distinguished. The species‐specific primer was designed according to the recA gene sequencing, which was employed for polymerase chain reaction with the template DNA of Lactobacillus strains. A single 231‐bp species‐specific band was found only in L. delbrueckii. A SNaPshot mini‐sequencing assay using recA as a target gene was also developed. The specificity of the mini‐sequencing assay was evaluated using 31 strains of L. delbrueckii species and was able to unambiguously discriminate strains belonging to the subspecies L. delbrueckii subsp. bulgaricus. CONCLUSION: The phylogenetic relationships of most strains in the L. acidophilus group can be resolved using recA gene sequencing, and a novel method to identify the species and subspecies of the L. delbrueckii and L. delbrueckii subsp. bulgaricus was developed by species‐specific polymerase chain reaction combined with SNaPshot mini‐sequencing. Copyright © 2012 Society of Chemical Industry     

Hepatic mRNA expression for genes related to somatotropic axis,glucose and lipid metabolisms,and inflammatory response of periparturient dairy cows treated with recombinant bovine somatotropin

Objectives of this experiment were to evaluate the effects of recombinant bovine somatotropin (rbST) treatment of periparturient dairy cows on hepatic mRNA expression for genes related to the somatotropic axis, insulin, glucose, and lipid metabolism, inflammation, and oxidative stress. Holstein cows were enrolled in the experiment at 253 ± 3 d of gestation and assigned to 1 of 3 treatments: untreated control (n = 53), 87.5 mg of rbST (n = 56; rbST87.5), and 125 mg of rbST (n = 57; rbST125). Cows in the rbST87.5 and rbST125 treatments received weekly injections of rbST from ?21 to 28 d relative to calving. A subsample of cows (control = 20, rbST87.5 = 20, rbST125 = 20) was randomly selected for collection of liver samples according to expected calving date, BCS, and previous lactation 305-d mature equivalent milk yield. Only cows that had liver sampled at ?21 ± 3, ?7 ± 3, and 7 ± 3 d relative to calving were used in the current experiment. Blood, sampled weekly from ?28 to 21 d relative to calving, was used to determine the concentrations of growth hormone, insulin-like growth factor 1, insulin, cortisol, fatty acids, β-hydroxybutyrate, glucose, haptoglobin, and tumor necrosis factor-α. Liver samples were used to determine hepatic mRNA expression of 50 genes. Treatment with rbST increased growth hormone concentrations during the postpartum period (control = 9.0 ± 0.7, rbST87.5 = 15.3 ± 1.0, rbST125 = 18.5 ± 1.3 ng/mL) and increased insulin-like growth factor 1 concentrations during the prepartum period (control = 107.4 ± 7.2, rbST87.5 = 126.9 ± 6.6, rbST125 = 139.4 ± 6.9 ng/mL). Control cows had greater postpartum concentrations of β-hydroxybutyrate (control = 776.4 ± 64.0, rbST87.5 = 628.4 ± 59.7, rbST125 = 595.4 ± 60.9 µmol/L) than rbST cows. The rbST87.5 and rbST125 treatments upregulated the hepatic mRNA expression for somatotropic axis genes (GHR, GHR1A, IGF1, IGFBP3, and SOCS2) on d ?7 relative to calving and upregulated the mRNA expression for SOCS2 on d 7. On d ?7, rbST87.5 and rbST125 treatments increased mRNA expression for genes involved in hepatic lipid transport (ANGPTL4, APOA5, APOB100, and SCARB1) and downregulated mRNA expression for PPARD, which is involved in lipid storage. On d 7, rbST tended to upregulate the mRNA expression for genes involved in gluconeogenesis (PCK1) and fatty acid β-oxidation (ACOX1), and downregulated the mRNA expression for genes involved in inflammation (TNFRSF1A, ICAM1, CXCL1, MYD88, HIF1A, IL1RN, NFKBIA, and SOCS3) and oxidative stress (XBP1). Administration of rbST during the periparturient period may improve liver function and health by increasing hepatic capacity for gluconeogenesis and lipid transport and by reducing inflammation and oxidative stress.     

Consumer Sensory Analysis of High Flavonoid Transgenic Tomatoes

Tomatoes have ameliorative effects on cardiovascular disease and cancer. In this study, metabolic engineering of flavonoids was utilized to improve the nutritional value of tomatoes by increasing flavonol and anthocyanin content. Total flavonol content was significantly increased in both the peel and flesh using the onion chalcone isomerase (CHI) gene. The Delila (Del) and Rosea1 (Ros1) genes from the snapdragon Antirrhinum majus were concomitantly expressed to produce an anthocyanin‐rich tomato which was purple in color. Sensory evaluation by a panel of 81 untrained consumers revealed no significant difference in liking of color or texture between CHI, Del/Ros1, and wild‐type tomatoes. Consumers reported marginal but significantly higher preference for the flavor and overall liking of CHI tomatoes over Del/Ros1 and wild‐type tomatoes. This study is the first to report the results of sensory tests of transgenic tomatoes by a consumer panel representing the general consuming public.     

Pathogenicity of Foodborne, Environmental and Clinical Isolates of Listeria Monocytogenes in Mice

Foodborne, environmental and clinical isolates of Listeria monocytogenes and other Listeria species were screened for pathogenicity in immunocompromised mice. Of 218 isolates of L. monocytogenes, 203 were pathogenic and 15 were nonpathogenic. All non-monocytogenes species were nonpathogenic. Pathogenic isolates were hemolytic for sheep blood. In contrast, many nonpathogenic isolates were weakly hemolytic, but were CAMP positive. Lethal doses (LD₅₀) of pathogenic isolates were 5-480 cells for immunocompromised mice and 7.2 × 10⁵ to 8.4 × 10⁷ for nonimmunocompromised mice; whereas LD₅₀s for nonpathogenic isolates were > 10⁸ cells in both immunocompromised and nonimmunocompromised mice. Selected test isolates of L. monocytogenes were serotyped; the most common serotypes were 1/2b, 1/2a, 3a and 4b. The initial source and serotype of the isolate appeared not to be related to pathogenicity in immunocompromised mice. However, hemolytic activity was related to pathogenicity.     

Detection of Virulence Genes and Growth Potential in Listeria monocytogenes Strains Isolated from Ricotta Salata Cheese

Ricotta Salata is a traditional ripened and salted whey cheese made in Sardinia (Italy) from sheep's milk. This product is catalogued as ready‐to‐eat food (RTE) since it is not submitted to any further treatment before consumption. Thus, foodborne pathogens, such as Listeria monocytogenes, can represent a health risk for consumers. In September 2012, the FDA ordered the recall of several batches of Ricotta Salata imported from Italy linked to 22 cases of Listeriosis in the United States. This study was aimed at evaluating the presence and virulence properties of L. monocytogenes in 87 samples of Ricotta Salata produced in Sardinia. The ability of this product to support its growth under foreseen packing and storing conditions was also evaluated in 252 samples. Of the 87 samples 17.2% were positive for the presence of L. monocytogenes with an average concentration of 2.2 log₁₀ cfu/g. All virulence‐associated genes (prfA, rrn, hlyA, actA, inlA, inlB, iap, plcA, and plcB) were detected in only one isolated strain. The Ricotta Salata samples were artificially inoculated and growth potential (δ) was assessed over a period of 3 mo. The value of the growth potential was always >0.5 log₁₀ cfu/g under foreseen packing and storing conditions. This study indicates that Ricotta Salata supports the L. monocytogenes growth to levels that may present a serious risk to public health, even while stored at refrigeration temperatures.     

Development and application of a new multiplex real‐time PCR assay for simultaneous identification of Brucella melitensis,Cronobacter sakazakii and Listeria monocytogenes in raw milk and cheese

Brucella melitensis, Cronobacter sakazakii and Listeria monocytogenes are important foodborne pathogens in milk and milk products, which are responsible for a variety of diseases that pose serious hazards to public health and food safety. The objective of this study was to develop a novel multiplex RTi‐PCR for the detection of B. melitensis, C. sakazakii and L. monocytogenes and to characterise the potential risk of these pathogens in raw milk and cheese. The raw milk (n = 25) and cheese samples (n = 20) were analysed by multiplex RTi‐PCR assay and detected for quantification of the three pathogens. In this study, B. melitensis, C. sakazakii and L. monocytogenes were simultaneously identified using BMEII0466, mms operon and hly as target genes, respectively. The multiplex RTi‐PCR assay that was developed showed good sensitivity and selectivity for the pathogenic microorganisms (r² = 0.986–0.997). Multiplex RTi‐PCR results showed that most of the samples were contaminated with the pathogens screened.     

Antibacterial Activity of Ozone‐Depolymerized Crawfish Chitosan

ABSTRACT: Antimicrobial activities of chitosan samples with different molecular weights (1333, 432, 201, 131, and 104 kDa) prepared by ozone treatment were examined against 2 Gram‐positive bacteria (Listeria monocytogenes and Staphylococcus aureus) and 2 Gram‐negative bacteria (Escherichia coli and Pseudomonas fluorescen) to investigate the effect of chitosan's molecular weight and concentration on the inhibition of bacterial growth. Antimicrobial activity of chitosan varied depending on the molecular weight, concentration of chitosan, and type of microorganism. Generally, the effectiveness of the chitosans significantly increased with increasing chitosan concentration, regardless of molecular size and types of bacteria. Chitosan with molecular weights ranging from 104 to 201 kDa showed relatively greater antimicrobial activity against L. monocytogenes, S. aureus, and P. fluorescen; whereas for E. coli, intermediate molecular weight chitosan was more effective in growth inhibition than lower or higher molecular weight chitosan particularly at 0.1% concentration.     

Prevalence of Virulence Genes in Extended‐Spectrum β‐lactamases (ESBLs)‐Producing Salmonella in Retail Raw Chicken in China

Extended‐spectrum β‐lactamases (ESBLs)‐producing Salmonella is a tremendous hazard to food safety and public health. The objective of this study was to determine the prevalence of 30 virulence genes (avrA, sipA, sseC, marT, rhuM, siiE, pipA, pipD, envR, gogB, gtgA, sodC1, sseI, irsA, sopE2, spvC, rck, spvR, fhuA, msgA, pagK, srfj, stkc, fimA, lpfD, pefA, stcC, steB, stjB, and tcfA) in 156 ESBLs‐producing Salmonella isolates that belonged to 21 serotypes. These isolates were recovered from retail raw chicken samples collected from 5 provinces and 2 national cities in China between 2007 and 2012. The results indicated that 154 (98.7%) ESBLs‐producing Salmonella isolates carried at least 1 virulence gene, 138 (88.5%) simultaneously carried at least 5 virulence genes, 107 (68.6%) carried 10 or more, and 20 (12.8%) carried 15 or more virulence genes. The most frequently detected virulence genes were marT (n = 127, 81.4%), siiE (n = 126, 80.8%), msgA (n = 121, 77.6%), and sipA (n = 121, 77.6%). Significant difference was identified between detection percentages of virulence genes of rhuM, pipD, envR, sopE2, pagK, lpfD, steB, and stjB in S. Indiana, S. Thompson, S. Enteritidis, S. Typhimurium, S. Shubra, S. Edinburg, and S. Agona isolates. Distribution of virulence genes were significantly influenced by sampling districts (P < 0.01), especially for sodC1 and pipD, and then were msgA and sopE2. The heatmap showed the frequencies of virulence genes in ESBLs‐producing isolates from retail chickens in southern, central, and northern regions of China were completely different from each other. Based on our findings, ESBLs‐producing Salmonella of retail chicken origin were common carriers of multiple virulence genes and were regionally distributed.     

Prevalence,characterization, and genetic diversity of Listeria monocytogenes isolated from foods of animal origin in North East India

Listeria monocytogenes is a pathogenic microorganism infects man mostly through food. A total of 1615 samples of foods of animal origin and water were collected from retail meat shops of North-Eastern India and processed. Sixty-three (3.9%) samples were positive for L. monocytogenes. Animal origin foods showing the highest prevalence was chevon (9.8%) followed by beef (8.9%), chicken (8.5%), pork (2.8%) and milk (1.8%). The prevalence rate in water from retail meat shops was 10%. Recovered L. monocytogenes were distributed into 3 serogroups, of which 74.6% fit in to 1/2a, 3a serogroup, 17.5% to 1/2b, 3b and 7.9 % to 4b, 4d, 4e serogroups. Thirty-five isolates out of 63 possessed all the tested four virulence genes. RAPD- and ERIC -PCR based analyses jointly revealed a discriminative genetic profile for the L. monocytogenes. On the whole, the occurrence of L. monocytogenes in foods of animal origin of North Eastern India displays public health hazard.     

Growth Potential of Listeria Monocytogenes and Staphylococcus Aureus on Fresh‐Cut Tropical Fruits

The objective of this study was to evaluate the fate of Staphylococcus aureus, Listeria monocytogenes, and natural microbiota on fresh‐cut tropical fruits (pitaya, mango, papaya and pineapple) with commercial PVC film at different storage temperature (5, 13, and 25 °C). The results showed that S. aureus, L. monocytogenes, and natural microbiota increased significantly on fresh‐cut tropical fruits at 25 °C. Both pathogen and natural microbiota were able to grow on fresh‐cut tropical fruits at 13 °C. The maximum population of L. monocytogenes was higher than that of S. aureus on fresh‐cut tropical fruits. L. monocytogenes and S. aureus could survive without growth on fresh‐cut pitaya, mango, and papaya at 5 °C. The population of L. monocytogenes declined significantly on fresh‐cut pineapple at all temperature, indicating composition of fresh‐cut pineapple could inhibit growth of L. monocytogenes. However, S. aureus was still able to grow on fresh‐cut pineapple at storage temperature. Thus, this study suggests that 4 kinds of fresh‐cut tropical fruits (pitaya, mango, papaya, and pineapple) should be stored at low temperature to extend shelf life as well as to ensure the safety of fresh‐cut fruits.     

Requirement of glutathione for Sod1 activation during lifespan extension

It has been shown that the activation of cytosolic superoxide dismutase (Sod1) in Saccharomyces cerevisiae is only dependent on Ccs1, which is responsible for insertion of copper into the enzyme catalytic center, and that glutathione (GSH) is not necessary for this process. In this work, we addressed an important role of GSH in Sod1 activation by a Ccs1‐dependent mechanism during oxidative stress and its role in yeast lifespan. Exponential cells of Saccharomyces cerevisiae, treated or not with 0.5 mM menadione for 1 h, were used for evaluation of the effect of a mild oxidative stress pre‐treatment on chronological lifespan. The results showed that menadione induced a lifespan extension in the wild‐type (WT) strain but this adaptive response was repressed in gsh1 and in sod1 strains. Interestingly, menadione treatment increased SOD1 and CCS1 gene expression in both WT and gsh1 strains. However, while these strains showed the same Sod1 activity before treatment, only the WT presented an increase of Sod1 activity after menadione exposure. Glutathionylation seems to be essential for Sod1 activation since no increase in activity was observed after menadione treatment in grx1 and grx2 null mutants. Our results suggest that GSH and glutathionylation are fundamental to protect Sod1 sulfhydryl residues under mild oxidative stress, enabling Sod1 activation and lifespan extension. Copyright © 2010 John Wiley & Sons, Ltd.