Associations between Listeria monocytogenes genomic characteristics and adhesion to polystyrene at 8 °C.

Listeria monocytogenes remains a threat to the food system and has led to numerous foodborne outbreaks worldwide. L. monocytogenes can establish itself in food production facilities by adhering to surfaces, resulting in increased resistance to environmental stressors. The aim of this study was to evaluate the adhesion ability of L. monocytogenes at 8 °C and to analyse associations between the observed phenotypes and genetic factors such as internalin A (inlA) genotypes, stress survival islet 1 (SSI-1) genotype, and clonal complex (CC). L. monocytogenes isolates (n = 184) were grown at 8 °C and 100% relative humidity for 15 days. The growth was measured by optical density at 600 nm every 24 h. Adherent cells were stained using crystal violet and quantified spectrophotometrically. Genotyping of inlA and SSI-1, multi-locus sequence typing, and a genome-wide association study (GWAS) were performed to elucidate the phenotype-genotype relationships in L. monocytogenes cold adhesion. Among all inlA genotypes, truncated inlA isolates had the highest mean adhered cells, ABS595nm = 0.30 ± 0.15 (Tukey HSD; P < 0.05), while three-codon deletion inlA isolates had the least mean adhered cells (Tukey HSD; P < 0.05). When SSI-1 was present, more cells adhered; less cells adhered when SSI-1 was absent (Welch's t-test; P < 0.05). Adhesion was associated with clonal complexes which have low clinical frequency, while reduced adhesion was associated with clonal complexes which have high frequency. The results of this study support that premature stop codons in the virulence gene inlA are associated with increased cold adhesion and that an invasion enhancing deletion in inlA is associated with decreased cold adhesion. This study also provides evidence to suggest that there is an evolutionary trade off between virulence and adhesion in L. monocytogenes. These results provide a greater understanding of L. monocytogenes adhesion which will aid in the development of strategies to reduce L. monocytogenes in the food system.

The efficacy of different sanitizers against MS2 bacteriophage introduced onto plastic or stainless steel surfaces.

The virucidal activities of 11 prepared disinfectant solutions (active ingredients of household sanitizers) and 10 household sanitizers against bacteriophage MS2 on plastic and stainless steel surfaces were studied. Among the prepared sanitizers, 70-90% ethanol and ethanol-based disinfectants resulted in 1-2.5 log PFU/mL reductions on both surfaces. The 70% isopropanol and isopropanol-based formula reduced MS2 by 0.7-1.5 log PFU/mL on both surfaces. Other disinfectants, containing 0.1% benzalkonium chloride (BAC), 0.5% hydrogen peroxide, or 4% acetic acid, showed significant (P < 0.05) lower log reductions (-0.17-0.55 log PFU/mL) compared with other treatments. At room temperature, the virucidal activities of 70% ethanol on plastic (1.46-1.64 log PFU/mL reductions) and stainless steel (0.84-0.93 log PFU/mL reductions) surfaces were not significantly (P > 0.05) affected by the treatment time (30-600 s). However, 85% ethanol-treated groups showed significant (P < 0.05) higher log reductions in 60 and 600 s treated groups (1.69-2.24 log PFU/mL) compared with those in 30 s treated groups (0.92-1.32 log PFU/mL). Their virucidal activities were further examined at low temperatures (4 and 8 °C). We observed that the surface inactivation efficacies were not affected by the low temperatures. In addition, the virucidal activities of household sanitizers revealed that sanitizers with 1.84% (pH = 12.5, ∼17,500 ppm free-chlorine concentrations) or 3% (pH = 13.1, ∼38,100 ppm free-chlorine concentrations) sodium hypochlorite (NaClO) reduced 4.15-6.23 log PFU/mL MS2 on hard surfaces after 60 s contact time. Furthermore, an approximately 1.5 log PFU/mL reduction was observed in groups treated by sanitizer H (active ingredients: 58% ethanol + 0.1% quaternary ammonium compound). Household products with BAC or organic acid resulted in -0.28-0.33 log reductions on two surfaces after 30 or 60 s treatment. Therefore, the use of ethanol and NaClO-based products should be considered as a potential surface decontamination strategy in the food industry.

Ethanol Concentration of Kombucha Teas in British Columbia, Canada.

ABSTRACT: Kombucha is a sweetened tea beverage fermented by bacterial and yeast cultures. Sweeteners, such as glucose, sucrose, fructose, and others are converted by yeasts into ethanol and then by Acetobacter and other bacterial species into a weak acetic acid solution that is diluted, flavored, and packaged into glass or aluminum cans for consumer consumption. Naturally, fermented kombucha contains 0 to 3% alcohol by volume (ABV). However, kombucha containing ethanol is concerning for pregnant women and young children for whom low levels of ethanol consumption (<3% ABV) create adverse medical outcomes. In the province of British Columbia (BC), Canadian beverages containing >1% ABV are regulated as liquor. This study assessed ethanol concentrations in kombucha collected from processors and purchased at retail venues in BC. Ethanol values were compared with the place of manufacture (country or province) and place of purchase (grocery stores, restaurants, farmers' markets, recreational centers, and processors). Ethanol (n = 684) levels were measured by using a headspace gas chromatography-mass spectrometry method with a detection limit of 0.0002% ABV for ethanol. Overall, teas contained mean and median ethanol of 0.77 and 0.62% ABV, respectively, ranging from nondetectable up to 3.62% ABV. Four kombucha teas (0.6%) made by BC processors tested over 3% ABV, and 31.5% of samples contained ethanol that exceeded the BC regulatory limits for nonalcoholic beverages of 1% ABV. Kombucha manufactured in BC had significantly higher mean ethanol values (1.16% ABV) in comparison to all other places of manufacture. Similarly, mean ethanol tea values obtained from BC processors (1.2% ABV) and restaurants (1.01% ABV) were significantly higher than those obtained at other retail venues. This study demonstrates the potential for alcohol harm to at-risk populations consuming kombucha teas sold in BC.

Prevalence and toxigenic profiles of Bacillus cereus isolated from dried red peppers, rice, and Sunsik in Korea.

Bacillus cereus is a spore-forming foodborne pathogen responsible for diarrheal and emetic types of food poisoning. Intoxication is caused by various enterotoxins or by emetic toxin. Because of its widespread presence and the ability to form heat-stable endospores in a relatively short time, B. cereus has been difficult to control. In this study, 21 rice and 36 Sunsik (a mixture of powdered raw grains) samples were examined for the prevalence of B. cereus. A multiplex PCR assay was used to evaluate the distribution of 10 different toxigenicity-related genes among 1,082 B. cereus strains isolated from dried red peppers (919 isolates), rice (98 isolates), and Sunsik (65 isolates). The results suggest that (i) the examined foods were free of the emetic toxin but not free of enterotoxins and (ii) the distribution of enterotoxigenic genes was significantly different among the B. cereus isolates from various sources.

Cudrania tricuspidata Extract Protects against Reflux Esophagitis by Blocking H2 Histamine Receptors.

Cudrania tricuspidata has been used in East Asia as a folk medicine for symptoms such as inflammation, allergy, and gastritis. Administration of C. tricuspidata extract to pylori-ligated rat stomachs reduces gastric acid secretion and alleviates esophagus damage caused by gastric reflux. Therefore, in this study we aimed to investigate whether C. tricuspidata extracts inhibit reflux esophagitis by blocking H2 histamine receptor (H2R). Dimaprit, a H2R specific agonist, induced intracellular cyclic adenosine monophosphate (cAMP) production in U937 cells. Pretreatment with C. tricuspidata extracts significantly blocked dimaprit-induced cAMP production in a concentration-dependent manner. To extracted C. tricuspidata with different ethanol concentrations to determine the optimum method. We found that the 70% ethanol extract showed the most potent H2R antagonistic effect against dimaprit-induced cAMP production. However, water extract did not show any H2R blocking effect. These findings suggest that C. tricuspidata extracted using ethanol specifically inhibits gastric acid secretion and reduces esophageal injury by blocking H2R in a competitive manner. Therefore, C. tricuspidata extracts may be used in food or medicine to prevent H2R-related diseases, such as gastric hyperacidity and reflux esophagitis.

Oral Administration of Lactobacillus curvatus HY7601 and Lactobacillus plantarum KY1032 with Cinnamomi Ramulus Extract Reduces Diet-Induced Obesity and Modulates Gut Microbiota.

Obesity is a major health issue worldwide, and is associated with many diseases including type 2 diabetes. In this study, we evaluated the anti-obesity effects of combinations of two lactic acid bacteria (LAB), Lactobacillus curvatus HY7601 and Lactobacillus plantarum KY1032, and Cinnamomi Ramulus (CR) extract, and explored the mechanism through which they modulate gut microbiota using diet-induced obese mice. Male C57BL/6J mice were randomly divided into five groups that received a high-fat diet (HFD), HFD and LAB (HFD+LAB), HFD and CR extract (HFD+CR), HFD with LAB and CR extract (HFD+LAB+CR), or normal diet for 10 weeks. The mice in the HFD+LAB+CR group showed significant reductions in body weight gain, in particular epididymal fat and liver, blood leptin levels, and an increase in the levels of blood adiponectin. In addition, the LAB and CR extract altered the gut microbiota, mainly increasing the alpha diversity. These results demonstrate that a mixture of two LAB (Lactobacillus curvatus HY7601 and Lactobacillus plantarum KY1032) and CR extracts alleviate HFD-induced obesity, and has potential of being used as a strategy for the treatment of obesity.

Lactobacillus plantarum HY7714 Restores TNF-α Induced Defects on Tight Junctions.

In addition to intestinal balance, probiotics are known to have beneficial effects on skin inflammation, metabolic diseases, and emotions. Previously, we have reported the skin anti-aging effects of Lactobacillus plantarum HY7714 (HY7714) in a clinical trial. To prove the protective skin effects of HY7714 through the intestinal tight junction (TJ), we investigated the effects of HY7714 on the intestines through tumor necrosis factor (TNF)-α induced TJ defects in Caco-2 cells. Specifically, 24 h treatment with HY7714 restored the decreased expression of zonula occludens-1, occludin, and claudin-1 compared to the TNF-α-treated groups (P<0.05). It also attenuated the level of pro-inflammatory cytokines interleukin-6, 8, and 1ß. Further, increases in the mRNA levels of Elk-1, nuclear factor-κB, and myosin light chain kinase expression induced by TNF-α were recovered by HY7714. These findings imply that HY7714 improves intestinal barrier integrity and is a potential therapeutic agent for dysfunctions derived from TJ defects.

Prevalence and genetic diversity of Enterobacter sakazakii in ingredients of infant foods.

Various food samples in Korea were examined for the presence of Enterobacter sakazakii. Dried shrimp had the highest contamination rate among the examined dried fish products. E. sakazakii isolates were confirmed using an API 20E kit and a polymerase chain reaction (PCR) targeting the rDNA operons. The isolates were subtyped by pulsed-field gel electrophoresis (PFGE) using XbaI to elucidate the genetic diversity of the organisms. Ten pulsotypes were identified using PFGE and 22 types were identified from the random amplified polymorphism DNA (RAPD) assay. An antibiotic resistance test was performed by disk diffusion assay using eight antibiotics: nalidixic acid, ciprofloxacin, chloramphenicol, ampicillin, tetracycline, gentamicin, kanamycin, and cephalothin. Most of the E. sakazakii isolates were resistant to ampicillin or cephalothin but susceptible to the other antibiotics. The analysis of E. sakazakii isolates using PFGE, RAPD, and the antibiotic resistance test identified 18 composite types from 113 isolates, suggesting diverse sources of contamination.

Prevalence, genetic diversity, and antibiotic resistance patterns of Campylobacter jejuni from retail raw chickens in Korea.

Campylobacter jejuni is a frequently detected food-borne pathogenic bacterium. Clinical cases are mostly sporadic but campylobacteriosis can have serious consequences, such as the development of Guillain-Barré syndrome as well as diarrheal diseases. We examined 265 retail raw chickens from Korean markets for the presence of C. jejuni using the US Food and Drug Administration standard cultural method and multiplex polymerase chain reaction (mPCR). The mPCR-confirmed C. jejuni isolates were subtyped by pulsed field gel electrophoresis (PFGE) and flaA-typing for investigating the genetic diversity of the microorganism in retail raw chickens. Restriction enzymes SmaI and DdeI were used for PFGE and flaA-typing, respectively. Campylobacter spp. were found in 181 samples (68.3%) and C. jejuni in 100 samples (37.74%). For C. jejuni, 73 pulsotypes and 30 flaA types were detected. Antibiotic resistance tests performed by disk diffusion assay indicated that most C. jejuni isolates were resistant to tetracycline, nalidixic acid, and ciprofloxacin, and 87 composite types were revealed by PFGE, flaA-typing, and the antibiotic resistance tests. Our results show that the genetic diversity of C. jejuni isolates is very high and the correlation between genotype and antibiotic resistance was low even though many bacteria showed multi-drug resistance.

Prevalence and genetic diversity of Bacillus cereus in dried red pepper in Korea.

Bacillus cereus is a foodborne spore-forming bacterial pathogen that is ubiquitous in the natural environment. Infections with this pathogen manifest as diarrheal or emetic types of food poisoning. In this study, 140 samples of dried red pepper purchased in Korea were assayed for the presence of B. cereus according to the U.S. Food and Drug Administration standard culture method. A multiplex PCR assay was developed for the rapid confirmation of B. cereus as an alternative to conventional biochemical confirmation tests. The genetic diversity of B. cereus isolates was investigated using a random amplified polymorphic DNA (RAPD) assay. B. cereus was found in 84.3% of the dried red pepper samples, with an average concentration of 1.9 x 10(4) CFU/g. B. cereus could be detected and distinguished from B. thuringiensis in the multiplex PCR assay by using the BCFW1 plus BCrevnew and the K3 plus K5 primer sets designed to detect the gyrB gene of B. cereus and B. thuringiensis and the cry gene of B. thuringiensis. A RAPD assay using the OPG 16 and MUP 3 primers was used to successfully distinguish among isolates, thus elucidating the genetic diversity of B. cereus isolates. The discriminating ability of the OPG 16 primer (142 types) was about threefold higher than that of MUP 3 (52 types) in the RAPD assay.

Effects of naturally occurring compounds on fibril formation and oxidative stress of beta-amyloid.

Beta-amyloid (betaA)-induced oxidative toxicity on neuronal cells is a principal route in Alzheimer's disease (AD), and its toxicity occurs after fibril formation. Inhibitory or promoting effects of naturally occurring compounds on betaA fibril formation were evaluated. Among 214 tested compounds, curcuminoids, flavone type flavonoids, and naphthoquinones were shown to be potent inhibitors of betaA fibrilization. The addition of the curcuminoids, curcumin, demethoxycurcumin, and bisdemethoxycurcumin strongly inhibited betaA fibril formation. Flavonoids such as quercetin, rhamnetin, and fisetin strongly inhibited betaA fibril formation. Limonoids, cinnamic acids, and catechins enhanced fibril formation in vitro. Anthothecol possessed the most enhancing activity on fibril formation of the compounds tested. On the other hand, it was found that curcuminoids showed cytotoxicity with the 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazolium bromide assay and did not protect HT22 murine neuroblastoma cells from betaA(25-35) insult. Two flavone type flavonoids, morin and quercetin, exhibited no cytotoxicity and strongly protected HT22 murine neuroblastoma cells from betaA(25-35) oxidative attack. Conclusively, morin or quercetin could be a key molecule for the development of therapeutics for AD.

Oral administration of Lactobacillus plantarum HY7714 protects hairless mouse against ultraviolet B-induced photoaging.

Ultraviolet (UV) irradiation alters multiple molecular pathways in the skin, thereby inducing skin damage, including photoaging. In recent years, probiotics have gained interest due to their beneficial effects on skin health, such as inhibiting atopic dermatitis and improving skin immunity or inflammation. However, little is known about the effects of probiotics on UVBinduced photoaging. In this study, we evaluated the effect of Lactobacillus plantarum HY7714 against UVB-induced photoaging in human dermal fibroblasts and hairless mice. The results showed that L. plantarum HY7714 treatment effectively rescued UVB-reduced procollagen expression through the inhibition of UVB-induced matrix metalloproteinase (MMP)-1 expression in human dermal fibroblasts. Data from a western blot showed that L. plantarum HY7714 inhibited the phosphorylation of Jun N-terminal kinase, thereby suppressing the UVB-induced phosphorylation and expression of c-Jun. Oral administration of L. plantarum HY7714 clearly inhibited the number, depth, and area of wrinkles in hairless mouse skin. Histological data showed that L. plantarum HY7714 significantly inhibited UVB-induced epidermal thickness in mice. Western blot and zymography data also revealed that L. plantarum HY7714 effectively inhibited MMP-13 expression as well as MMP-2 and -9 activities in dermal tissue. Collectively, these results provide further insight regarding the skin biological actions of L. plantarum HY7714, a potential skin anti-photoaging agent.