In vitro OECD test methods applied to screen the estrogenic effect of chemicals, used in Korea.

In this study, 27 chemicals found in household products, which became an issue in Korea were screened for the agonistoc and antagonistic effects against human estrogen receptor using official Organization for Economic Cooperation and Development (OECD) in vitro assays, STTA assay using ERα-HeLa-9903 cell line and BG1Luc ER TA assay. In the case of human ER agonist screening by two assays, all tested chemicals did not show agonist effect against ER. In ER antagonist test by BG1Luc ER TA assay, five surfactants α-dodecyl-ω-hydroxypoly(oxyethylene), alcohols C16-18 ethoxylated, nonylphenol, ethoxylated, 3,6,9,12,15,18,21-heptaoxatritriacontan-1-ol, and α-dodecyl-ω-hydroxypoly(oxy-1,2-ethanediyl)) were found to exhibit weak antagonistic activities. The agonist/antagonist effects against human estrogen receptor of various chemicals, used in Korea by OECD test guideline are reported in this study. These results indicated that two OECD in vitro assays will can be applied in Korea by screening of agonistic/antagonistic effects against human ER of various chemicals.

Risk assessment based on urinary bisphenol A levels in the general Korean population.

Bisphenol A (BPA) is a high-volume industrial chemical used in the global production of polycarbonate plastics and epoxy resins, which are used in food and drink containers, such as tableware (plates and mugs). Due to its broad applications, BPA has been detected in human blood, urine and breast milk as well as environmental substances, including water, indoor and outdoor air, and dust. Indeed, exposure to high concentrations of BPA can result in a variety of harmful effects, including reproductive toxicity, through a mechanism of endocrine disruption. Our comparison of reported BPA urinary concentrations among different countries revealed that exposures in Korea may be higher than those in other Asian countries and North America, but lower than or similar to those in European countries. The current study included a total of 2044 eligible subjects of all ages. The subjects were evenly divided between males and females (48.58% and 51.42%, respectively). The geometric mean (GM) of pre-adjusted (adjusted) urinary BPA concentrations was 1.83µg/L (2.01µg/g creatinine) for subjects of all ages, and there was no statistically difference in BPA concentrations between males (1.90µg/L, 1.87µg/g creatinine) and females (1.76µg/L, 2.16µg/g creatinine). Multiple regression analysis revealed only one positive association between creatinine pre-adjusted urinary BPA concentration and age (ß=-0.0868, p<0.001). The 95th percentile levels of 24-hour recall (HR), food frequency questionnaires (FFQ) and estimated daily intake (EDI) through urinary BPA concentrations were 0.14, 0.13, and 0.22µg/kg bw/day, respectively. According to the Ministry of Food and Drug Safety (MFDS), a tolerable daily intake (tDI) of 20µg/kg bw/day was established for BPA from the available toxicological data. Recently, the European Food Safety Authority (EFSA) established a temporary TDI of 4µg/kg bw/day based on current toxicological data. By comparing these TDIs with subjects' exposure, we conclude that there are no health concerns for any age group as a result of current levels of dietary exposure to BPA.

Optimization of the elution buffer and concentration method for detecting hepatitis E virus in swine liver using a nested reverse transcription-polymerase chain reaction and real-time reverse transcription-polymerase chain reaction.

The aim of this study was to develop an optimal technique for detecting hepatitis E virus (HEV) in swine livers. Here, three elution buffers and two concentration methods were compared with respect to enhancing recovery of HEV from swine liver samples. Real-time reverse transcription-polymerase chain reaction (RT-PCR) and nested RT-PCR were performed to detect HEV RNA. When phosphate-buffered saline (PBS, pH 7.4) was used to concentrate HEV in swine liver samples using ultrafiltration, real-time RT-PCR detected HEV in 6 of the 26 samples. When threonine buffer was used to concentrate HEV using polyethylene glycol (PEG) precipitation and ultrafiltration, real-time RT-PCR detected HEV in 1 and 3 of the 26 samples, respectively. When glycine buffer was used to concentrate HEV using ultrafiltration and PEG precipitation, real-time RT-PCR detected HEV in 1 and 3 samples of the 26 samples, respectively. When nested RT-PCR was used to detect HEV, all samples tested negative regardless of the type of elution buffer or concentration method used. Therefore, the combination of real-time RT-PCR and ultrafiltration with PBS buffer was the most sensitive and reliable method for detecting HEV in swine livers.

Expression of intestinal transporter genes in beagle dogs.

This study was performed to produce a transcriptional database of the intestinal transporters of beagle dogs. Total RNA was isolated from the duodenum and the expression of various mRNAs was measured using GeneChip(®) oligonucleotide arrays. A total of 124 transporter genes were detected. Genes for fatty acid, peptide, amino acid and glucose and multidrug resistance/multidrug resistance-associated protein (MDR/MRP) transport were expressed at relatively higher levels than the other transporter types. The dogs exhibited abundant mRNA expression of the fatty acid transporters (fatty acid binding proteins, FABPs) FABP1 and FABP2, the ATP-binding cassettes (ABCs) ABCB1A and ABCC2, the amino acid/peptide transporters SLC3A1 and SLC15A1, the glucose transporters SLC5A1, SLC2A2 and SLC2A5, the organic anion transporter SLC22A9 and the phosphate transporters SLC20A1 and SLC37A4. In mice, a similar profile was observed with high expression of the glucose transporters SLC5A1 and SLC2As, the fatty acid transporters FABP1 and FABP2, the MDR/MRP transporters ABCB1A and ABCC2 and the phosphate transporter SLC37A4. However, the overall data reveal diverse transcriptomic profiles of the intestinal transporters of dogs and mice. Therefore, the current database may be useful for comparing the intestinal transport systems of dogs with those of mice to better evaluate xenobiotics.

Detection and identification of Vibrio species using whole-cell protein pattern analysis.

Outbreaks of foodborne diseases associated with Vibrio species such as V. parahaemolyticus, V. vulnificus, and V. cholerae frequently occur in countries having a dietary habit of raw seafood consumption. For rapid identification of different Vibrio species involved in foodborne diseases, whole-cell protein pattern analysis for 13 type strains of 12 Vibrio species was performed using SDS-PAGE analysis. Pathogenic Vibrio species such as V. parahaemolyticus, V. vulnificus, V. cholerae, V. alginolyticus, V. fluvialis, and V. mimicus were included in the 12 Vibrio species used in this study. Each of the 12 Vibrio species showed clearly specific band patterns of its own. Two different strains of V. parahaemolyticus showed two different SDS-PAGE wholecell protein patterns, giving the possibility of categorizing isolated strains in the same V. parahaemolyticus species into two subgroups. The 36 Vibrio isolates collected from sushi restaurants in Busan were all identified as V. parahaemolyticus by comparing their protein patterns with those of Vibrio type strains. The identified isolates were categorized into two different subgroups of V. parahaemolyticus. The whole-cell protein pattern analysis by SDS-PAGE can be used as a specific, rapid, and simple identification method for Vibrio spp. involved in foodborne diseases at the subspecies level.

Development of real-time PCR for the detection of Clostridium perfringens in meats and vegetables.

A real-time PCR assay was developed and validated inhouse specifically for the detection of Clostridium perfringens (Cl. perfringens) in meats and vegetables by comparing with the culture method. The detection limit of the real-time PCR assay in phosphate-buffered saline was 10² CFU/ml. When the two methods were compared in food samples inoculated with Cl. perfringens, the culture method detected 52 positives, whereas real-time PCR detected 51 positives out of 160 samples. The difference was without statistical significance (p>0.05). Real-time PCR assay is an option for quality assurance laboratories to perform standard diagnostic tests, considering its detection ability and time-saving efficiency.

Development of predictive models for the growth of Escherichia coli O157:H7 on cabbage in Korea.

Cabbage is the main material of coleslaw, a popular side dish in Korea as well as many other countries. In the present study, the combined effect of temperature (15, 25, and 35 °C) and relative humidity (60%, 70%, and 80%) on the growth of Escherichia coli O157:H7 on cabbage was investigated. The polynomial models for growth rate (GR), lag time (LT), and maximum population density (MPD) estimated from the Baranyi model were conducted with high coefficients of determination (R(2)> 0.98). Subsequently, performance and reliability of the models were assessed through external validation, employing three indices as bias factor (B(f)), accuracy factor (A(f)), and the standard error of prediction expressed in percentage (%SEP). The B(f), A(f), and %SEP values of the predictive models for GR were 1.008, 1.127 and 18.70%, while 1.033, 1.187 and 20.79% for LT and 0.960, 1.044 and 5.22% for MPD, respectively. The results demonstrated that the developed secondary models showed a good agreement between the observed and predicted values. Therefore, the established models can be suitable to estimate and control E. coli O157:H7 growth risk on cabbage at some steps from farm to table in Korea as a valuable tool.

Effect of wash treatments on reducing human norovirus on iceberg lettuce and perilla leaf.

Human noroviruses (NoVs) are major causes of nonbacterial gastroenteritis; they are transmitted by food and water, as well as person-to-person. The consumption of contaminated raw or uncooked food such as vegetables and fruits has been identified as a common source of human NoV outbreaks. In an effort to understand the survival and persistence of human NoVs on fresh produce, the efficacy of washing treatments in the removal of human NoVs from vegetables was evaluated. This study used artificially contaminated vegetables (iceberg lettuce and perilla leaf), and washing was done with tap water for convenience. Wash treatments included immersion in water, rinsing with running water, and a combination of immersion and rinsing (treatments I to III, respectively). The effect of a class I detergent, a commercial product used for washing fruits and vegetables, was also evaluated (treatment IV). After the wash treatments, the remnants of human NoVs on samples were measured via real-time reverse transcriptase PCR. The results varied among treatments and by vegetable. For iceberg lettuce, a reduction of 0.9 log was noted in the treatment III group. The wash treatment was more effective in the perilla leaf samples: each treatment significantly reduced the numbers of human NoVs (0.69- to 1.29-log reduction). These data demonstrated that wash treatments reduced numbers of virus from the surfaces of the vegetables. Therefore, washing would seem to be a basic step in reducing numbers of virus in food preparation and in viral transmission routes.

Prevalence, antibiotic resistance, and molecular characterization of Salmonella serovars in retail meat products.

The prevalence of Salmonella was determined in chicken meat (n = 26), beef (n = 49), and pork (n = 56) collected from wholesale markets, retail stores, and traditional markets in Seoul, South Korea, in 2009. Antibiotic resistance was assessed, and the molecular subtypes of Salmonella isolates were ascertained using an automated repetitive sequence-based PCR (rep-PCR) system (DiversiLab). A total of 18 Salmonella strains were isolated from 17 of 131 samples: 16 strains from each of 16 samples and 2 strains from the same pork sample. The prevalence of Salmonella from the retail meats was 2.0% in beef, 8.9% in pork, and 42.3% in chicken meat. Among 10 different serotypes, Salmonella enterica Panama was recovered from a beef sample, and Salmonella London and Salmonella Montevideo were the predominant serotypes from pork and chicken meat, respectively. The highest antibiotic resistance observed was to erythromycin (100%) followed by streptomycin (22.2%) and tetracycline and chloramphenicol (16.7%). Of the 18 isolates, 5 (27.8%) were resistant to two or more antibiotics, and 1 isolate from chicken meat was resistant to eight antibiotics, including cephalosporins. Differentiation between all of the Salmonella isolates except between Salmonella Montevideo and Salmonella London was successfully performed with the automated rep-PCR system, indicating that it can be added to the toolbox for source tracking of foodborne pathogens associated with outbreaks.

Evaluation of various PCR assays for the detection of emetic toxin producing Bacillus cereus.

Because conventional methods for detecting emetic-toxin-producing B. cereus are laborious and costly, various PCR assays, which are easy and cheap, have recently been reported. Therefore, this study estimated and compared the ability of various PCR assays to detect emetic-toxin-producing B. cereus strains isolated in Korea. The PCR assays were performed on 160 B. cereus strains, including 40 emetic-toxin-producing strains. Although the species-specific PCR assays were all shown to be highly specific, the sensitivities varied greatly. The accuracies of the primers were 97.5% (CER), 95.6% (EM1), 96.3% (RE234), 89.4% (CES), and 83.1% (Ces3R/CESR2). Moreover, the CER primer had a higher sensitivity (100%) than all the other primers tested, and a specificity of 96.7%. Thus, the CER primer was shown to be the most effective for screening the emetic-toxin-producing B. cereus strains tested in this study. However, the ability of these PCR assays to identify emetic-toxin-producing B. cereus should also be confirmed using other methods.

Disinfection of iceberg lettuce by titanium dioxide-UV photocatalytic reaction.

Securing the physical quality and microbial safety of fresh foods has been a major focus in the food industry. To improve quality and increase the shelf life of fresh produce, disinfection methods have been developed. Titanium dioxide (TiO2) photocatalytic reactions under UV radiation produce hydroxyl radicals that can be used for disinfection of foodborne pathogenic bacteria. We investigated the effects of TiO2-UV photocatalytic disinfection on the shelf life of iceberg lettuce. Counts of natural microflora (total aerobic bacteria, coliforms, psychrotrophic bacteria, and yeasts and molds) and inoculated pathogenic bacteria (Escherichia coli, Listeria monocytogenes, Staphylococcus aureus, and Salmonella Typhimurium) on iceberg lettuce were determined after 20-min treatments with TiO2-UV, UV radiation, a sodium hypochlorite (NaOCl) solution, and tap water. TiO2-UV treatment reduced the number of microorganisms by 1.8 to 2.8 log CFU/g compared with reductions of 0.9 to 1.4 and 0.7 to 1.1 log CFU/g obtained with UV radiation and NaOCl treatments, respectively. Treatment with tap water was used as a control and resulted in no reductions. Counts of microflora for iceberg lettuce at 4 and 25 degrees C were determined during a 9-day period. TiO2-UV treatment resulted in 1.2- and 4.3-log increases in the counts of total aerobic bacteria at 4 and 25 degrees C, respectively, compared with 1.3- to 1.6-log and 4.4- to 4.8-log increases due to UV radiation and NaOCl treatments.

Optimization of methods for detecting norovirus on various fruit.

Methods for detecting norovirus (NoV) in food are crucial for investigation and prevention of outbreaks caused by NoV-contaminated food. However, current NoV detection methods have not been well examined or optimized. In this study, the effectiveness of various methods for eluting NoV from various fruit, concentrating the virus using polyethylene glycol (PEG), and extracting the viral RNA for subsequent assay by RT-PCR was optimized. First, six different buffers previously described for eluting NoV from fruit surfaces were evaluated. A known amount of NoV was spiked onto the surface of grapes, strawberries, and raspberries, and the virus was recovered with distilled water, 0.05 M glycine-0.14 M NaCl (pH 7.5), 2.9% tryptose phosphate broth-6% glycine, 100 mM Tris-HCl (pH 9.5), 50 mM glycine-50 mM MgCl(2) (pH 9.5), or 3% beef extract. Quantitation of the recovered virus using RT-PCR revealed that the most effective elution buffer was 3% beef extract. Secondly, to optimize a method for concentrating the recovered NoV, the key parameters of PEG precipitation, a typical method for concentrating enteric virus, were investigated. The influence of PEG molecular weight and the duration and temperature of the precipitation procedure were examined. NoV was concentrated most efficiently by precipitation when PEG (10,000) was used for 4h at room temperature. Finally, five different methods for nucleic acid extraction were evaluated. Among RNA extraction methods examined, QIAamp Viral RNA Mini kit showed the best recovery efficiency. Using the optimized method, approximately 6-80% of the seeded NoV was recovered from the various fruit.