• Title/Summary/Keyword: C jejuni

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Development of a One-Step PCR Assay with Nine Primer Pairs for the Detection of Five Diarrheagenic Escherichia coli Types

  • Oh, Kyung-Hwan;Kim, Soo-Bok;Park, Mi-Sun;Cho, Seung-Hak
    • Journal of Microbiology and Biotechnology
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    • v.24 no.6
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    • pp.862-868
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    • 2014
  • Certain Escherichia coli (E. coli) strains have the ability to cause diarrheal disease. Five types of diarrheagenic E. coli have been identified, including EHEC, ETEC, EPEC, EAEC, and EIEC. To detect these five diarrheagenic types rapidly, we developed a one-step multiplex PCR (MP-PCR) assay using nine primer pairs to amplify nine virulence genes specific to the different virotypes, with each group being represented (i.e., stx1 and stx2 for EHEC, lt, sth, and stp for ETEC, eaeA and bfpA for EPEC, aggR for EAEC, and ipaH for EIEC). The PCR primers were constructed using MultAlin. The sensitivity and specificity of the constructed multiplex PCR primers were measured using DNA isolated from diarrheagenic E. coli strains representing each group. The limits of detection were as follows: $5{\times}10^1CFU/ml$ for EHEC, $5{\times}10^3CFU/ml$ for ETEC expressing lt and sth, $5{\times}10^4CFU/ml$ for ETEC expressing stp, $5{\times}10^2CFU/ml$ for EPEC, $5{\times}10^4CFU/ml$ for EAEC, and $5{\times}10^2CFU/ml$ for EIEC. To confirm the specificity, C. jejuni, C. perfringens, S. Typhimurium, V. parahaemolyticus, L. monocytogenes, Y. enterocolitica, B. cereus, and S. aureus were used as negative controls, and no amplification was obtained for these. Moreover, this kit was validated using 100 fecal samples from patients with diarrhea and 150 diarrheagenic E. coli strains isolated in Korea. In conclusion, the multiplex PCR assay developed in this study is very useful for the rapid and specific detection of five diarrheagenic E. coli types. This single-step assay will be useful as a rapid and economical method, as it reduces the cost and time required for the identification of diarrheagenic E. coli.

The Analysis for Minimum Infective Dose of Foodborne Disease Pathogens by Meta-analysis (메타분석에 의한 식중독 원인 미생물들의 최소감염량 분석)

  • Park, Myoung Su;Cho, June Ill;Lee, Soon Ho;Bahk, Gyung Jin
    • Journal of Food Hygiene and Safety
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    • v.29 no.4
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    • pp.305-311
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    • 2014
  • Minimum infective dose (MID) data has been recognized as an important and absolutely needed in quantitative microbiological assessment (QMRA). In this study, we performed a comprehensive literature review and meta-analysis to better quantify this association. The meta-analysis applied a final selection of 82 published papers for total 12 species foodborne disease pathogens (bacteria 9, virus 2, and parasite 1 species) which were identified and classified based on the dose-response models related to QMRA studies from PubMed, ScienceDirect database and internet websites during 1980-2012. The main search keywords used the combination "food", "foodborne disease pathogen", "minimum infective dose", and "quantitative microbiological risk assessment". The appropriate minimum infective dose for B. cereus, C. jejuni, Cl. perfringens, Pathogenic E. coli (EHEC, ETEC, EPEC, EIEC), L. monocytogenes, Salmonella spp., Shigella spp., S. aureus, V. parahaemolyticus, Hepatitis A virus, Noro virus, and C. pavum were $10^5cells/g$ (fi = 0.32), 500 cells/g (fi = 0.57), $10^7cells/g$ (fi = 0.56), 10 cells/g (fi = 0.47) / $10^8cells/g$ (fi = 0.71) / $10^6cells/g$ (fi = 0.70) / $10^6cells/g$ (fi = 0.60), $10^2{\sim}10^3cells/g$ (fi = 0.23), 10 cells/g (fi = 0.30), 100 cells/g (fi = 0.32), $10^5cells/g$ (fi = 0.45), $10^6cells/g$ (fi = 0.64), $10{\sim}10^2particles/g$ (fi = 0.33), 10 particles/g (fi = 0.71), and $10{\sim}10^2oocyst/g$ (fi = 0.33), respectively. Therefore, these results provide the preliminary data necessary for the development of foodborne pathogens QMRA.

Analysis of Microbial Contamination in Poultry Slaughtering Operations for the Application of HACCP (HACCP 적용을 위한 도계처리 공정내 미생물 오염의 분석)

  • 홍종해;권혁무;고주언
    • Journal of Environmental Health Sciences
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    • v.22 no.1
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    • pp.45-50
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    • 1996
  • The application of HACCP system, which was adopted by Codex Alimentarius Committee for the safe meat and poultry production, is one of the urgent task for competing in the world trade markets. But there have been no useful analytical studies to identify the causes of contamination in the poultry meat processing plants in Korea. This study was conducted to investigate the potential hazards during the operations by the microbiological examination for the poultry meat processing plant (20,000 birds capacity a day) located in Kangwon province. In spite of air contamination of work places, it may not directly affect the surface contamination of poultry meats. But the risk of Campylobacter jejuni/coli contamination was high. The number of total count was decreased about ten times, but remarkable changes of microbial contamination could not be recognized in each procedure during the operations. The washing water was already contaminated as much as $10^{3-6}CFU/ml$ in SPC before the operations. It means that to keep water tanks hygienic is a primary step to prevent the occurrences of microbial contamination. The overflow and recirculation of water in scalding, washing, and chilling was aslo an important factor for a hygienic control. Based on this study, the followings could be regarded as an important factors for hygenic control in the poultry slaughtering plants on a small scale. The temperature of water used for scalding should be constantly maintained on a required temperature, and the overflow rate of 1~1.5 liter per bird. The carcass surface and the body cavity should be washed thoroughly and the cross-contamination due to facilities, workers, and tools should be prevented. The chilling water sholud be maintained under 5$\circ$C of temperature with ice and overflow, and residual chlorine level of 50 ppm.

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Microbiological Quality of Raw and Cooked Foods in Middle and High School Food Service Establishments (서울시 일부 중.고등학교의 급식용 식재료 및 조리식품의 미생물학적 품질)

  • Kim, Myung-Hee;Shin, Weon-Sun
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.37 no.10
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    • pp.1343-1356
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    • 2008
  • The evaluation of microbiological quality for school food samples collected from 19 selected middle and high schools located in Seoul was undertaken. Eighty-nine food samples consisting of 38 non-pretreated vegetables, 13 pre-washed and cut vegetables, 9 meats and poultry, 3 fish and shellfish, 7 dried fish, and shellfish and 20 processed foods were collected. Aerobic plate count, total coliforms, and Escherichia coli (E. coli ) were detected using $Petrifilm^{TM}$, and the food-borne pathogens were screened by multiplex PCR with species-specific primer sets. Sequentially, the quantitative and confirmative test of the food-borne pathogens were carried out with the selective media and biochemical kits. The contamination of coliform counts was observed on the pre-washed vegetables ($3.4{\sim}4.3\;log\;CFU/g$) and meats ($2.2{\sim}4.3\;log\;CFU/g$). Also, the cooked foods were heavily contaminated with coliform, ranging from 1.0 to $5.5\;log\;CFU/g$. E. coli counts were found in 16 raw and cooked food samples, exceeding the microbiological standards for the guideline of safety management for school foods. Through PCR detection, B acillus cereus was detected in 32 raw and cooked foods, and quantitatively found in pre-washed carrot, radish, and pan-broiled dried shrimp and filefish ranging from $2.3{\sim}3.6\;log\;CFU/g$, respectively. E. coli O157:H7 was detected on frozen pork sample and was confirmed with API kit. Campylobacter jejuni was found in 3 ready-to-eat type vegetables. Vibrio parahaemolyticus were found in 4 pre-washed vegetables and 2 cooked foods, indicating unsatisfactory quality based upon the microbiological standards of ready-to-eat vegetables and cooked foods by Korea Food and Drug Administration. Salmonella spp. was detected in frozen chicken sample and confirmed by API kit and latex antisera agglutination.

Isolation and Identification of Pathogenic Bacteria from Spinach (시금치로부터 병원성세균의 분리 및 동정)

  • Kim, Hye-Jung;Kim, Young-Hoon;Lee, Dong-Sun;Paik, Hyun-Dong
    • Korean Journal of Food Science and Technology
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    • v.35 no.1
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    • pp.97-102
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    • 2003
  • Raw and washed spinaches were tested to evaluate the incidences of Aeromonas hydrophila, Escherichia coli O157:H7, Plesiomonas shigelloides, Pseudomonas aeruginosa, Salmonella spp., Shigella spp., Yersinia enterocolitica, Bacillus cereus, Campylobacter jejuni, Clostridium perfringens, Listeria monocytogenes, and Staphylococcus aureus. Four pathogenic bacteria were isolated from spinach samples, and identified by morphological and biochemical methods, including API and ATB identification systems. Isolates from MacConkey, Cereus Selective, Clostridium Perfringens, and Baird-Parker agar media were in 99.9, 99.8, 99.9, and 97.8% agreements with A. hydrophila, B. cereus, C. perfringens, and S. aureus at the species level, respectively. SET-RPLA revealed, among the five strains of S. aureus isolates, two produced type A enterotoxin. All five strains of B. cereus isolates produced enterotoxin as revealed with CRET-RPLA.

Prevalence of Food Poisoning Bacteria on Hands in Various Age Groups (손 위생에 대한 식중독 원인균 실태조사)

  • Chung, Jae-Keun;Kim, Min-Jee;Kee, Hye-Young;Choi, Mi-Hwa;Seo, Jin-Jong;Kim, Sun-Hee;Park, Jong-Tae;Kim, Myung-Goun;Kim, Eun-Sun
    • Journal of Food Hygiene and Safety
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    • v.23 no.1
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    • pp.40-50
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    • 2008
  • Spread of pathogenic micro-organisms through contaminated hands is a well recognized way of transmitting disease such as food poisoning. We investigated the prevalence of aerobic plate counts, coliform bacteria, and food-poisoning bacteria on hands in various age groups. The average number of aerobic plate counts was 3.3 log CFU/hand in kindergarteners, 3.4 log CFU/hand in elementary students, 3.2 log CFU/hand in middle school students, 3.4 log CFU/hand in high school students, and 3.3 log CFU/hand in adults. Two kindergarteners, 6 elementary students, and 2 adults were positive for the coliform bacteria. Among the food poisoning bacteria we tested, S. aureus was isolated from 47 individuals. Eight isolates of B cereus were all from kindergarteners. C. perfringens was isolated from 7 individuals. Among 47 isolates of S. aureus, 25 isolates produced toxins. Seven of eight isolates of B. cereus produced toxins. None of seven C. peifringens isolates produced toxins. All 47 isolates of S. aureus were sensitive to ciprofloxacin, trimethoprim/sulfamethoxazole, clindamyccin, imipenem, rifampin and vancomycin. Four isolates (8.5%) were resistant to cefepime, chloramphenicol, cefotetan, and gentamycin. Five isolates (10.6%) were resistant to oxacillin and 6 isolates were resistant to tetracycline. This study shows that it is needs to be established policy of school lunch and personal sanitation management.

Microbiological Quality and Potential Pathogen Monitoring for Powdered Infant Formulas from the Local Market (영유아용 분말 조제분유의 미생물 품질분석과 위해세균 모니터링)

  • Hwang, Ji-Yeon;Lee, Ji-Youn;Park, Jong-Hyun
    • Food Science of Animal Resources
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    • v.28 no.5
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    • pp.555-561
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    • 2008
  • Ninety-nine samples of powdered infant formula in a market were collected from the local market and their contaminations for total aerobic bacteria, coliform, FAO/WHO Category A, B, and C pathogens were analyzed. Total aerobic bacteria were detected in 92 of 99 samples (93%) at levels of $1.83{\pm}0.68\;Log\;MPN/g$. These levels were below legal levels specified for infant formulas except for one sample detected by 4.5 Log CFU/g. Coliform was detected in 12 of 99 samples (12%) at levels of $1.26{\pm}1.03\;Log\;MPN/g$ whereas non-detection was required according to the specification of coliform in infant formulas. Escherichia coli was detected in 1 of 99 samples by 0.48 Log MPN/g. Salmonella and Enterobacter sakazakii among Category A weren't detected in all the samples. Enterobacteriaceae, Category B group, were detected in 25 samples of total 99 samples (25%) by $0.83{\pm}1.37\;Log\;MPN/g$. Enterobacteriaceae identified by API 20E were Escherichia vulneris, Es. hermannii, Pantoea spp., Citrobacter koseri, Klebsiella pneumoniae, En. cloaceae. Bacillus cereus among Category C was highly detected in 29 of 99 samples (29%) at levels of $0.69{\pm}0.32\;Log\;MPN/g$ with the most probable number count method, which were below legal levels for the specification of B. cereus in infant formulas. Clostridium perfringens, E. coli O157, Staphyloccus aureus, Listeria monocytogenes, Yersinia enterocolitica, and Campylobacter jejuni/coli were not detected. Contamination level of major pathogens was low and falls within the range of specification of infant formulas. However, Enterobacteriaceae and B.cereus showed the high prevalence and some Enterobacteriaceae causing disease were detected. Therefore, it is necessary to monitor the potential pathogens continually and reduce them to improve the microbial quality of non-sterilized powdered infant formulas.

Microbiological Evaluation of Foods and Kitchen Environments in Childcare Center and Kindergarten Foodservice Operations (보육시설과 유치원 급식의 식품 및 환경 미생물의 오염도 평가)

  • Seol, Hye-Rin;Park, Hyoung-Su;Park, Ki-Hwan;Park, Ae-Kyung;Ryu, Kyung
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.38 no.2
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    • pp.252-260
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    • 2009
  • Whereas the numbers of childcare centers and kindergartens are increasing rapidly, systematic management to control the food safety of foodservice operation is not yet well established. Samples from 12 centers in Seoul and Gyeonggi Province were collected to assess the microbiological quality of 32 raw materials, 24 cooked foods, 76 food-contact surfaces (knives, cutting boards, dish towels and gloves), 17 employees' hands and 12 air-borne bacteria. The microbiological analyses were performed for aerobic plate counts (APC), Enterobacteriaceae, E. coli and 7 pathogens (B. cereus, C. jejuni, C. perfringens, L. monocytogenes, Salmonella spp., S. aureus, and V. parahaemolyticus). Among raw materials, E. coli ($1.39{\sim}2.08\;\log\;CFU/g$) were detected in 4 out of 6 meats and 7.46 log CFU/g of APC in tofu. High enterobacteriaceae levels of 4.23, 5.14 and 4.19 log CFU/g were found in cucumber salad, steamed spinach with seasonings and steamed bean sprout with seasonings, respectively. No pathogens were found in all samples except for C. perfringens detected from raw spinach and raw lotus root. Only APC and enterobacteriaceae were found in food-contact surfaces. Two of the 23 knives and three of the 24 kitchen boards showed over 500 CFU/$100\;cm^2$ of APC; also, APC levels (5.03 to 5.44 log CFU/g) were detected in 4 of the 12 dish towels. Only one glove showed Enterobacteriaceae (2.44 log CFU/glove) contamination. Enterobacteriaceae were found in 2 employees' hands ($2.37{\sim}4.44\;\log\;CFU$/hand) among the 16 employees. The contamination levels of air-borne bacteria were shown unacceptable in two (2.25 and 2.30 log CFU/petri-film/15 min) out of the 12 kitchen areas. These results suggest that the microbiological hazards in some foods and environments are not well controlled and thus a guideline should be provided to ensure the food safety in childcare center and kindergarten foodservice operations.

Change of Harmful Micnoorganisms in Pickling Process of Salted Cabbage According to Salting and Washing Conditions (배추김치의 절임공정 조건에 따른 위해미생물 변화)

  • Kim, Jin-Hee;Lee, Yu-Keun;Yang, Ji-Young
    • Journal of Food Hygiene and Safety
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    • v.26 no.4
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    • pp.417-423
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    • 2011
  • Salted Cabbage products purchased from different companies at 4 different districts in South Korea were detected in this study. Cabbage and salt are the main materials for kimchi manufacture. The results of general bacteria contaminated in the samples were $1.4{\times}10^5$, $6.4{\times}10^5$, $1.7{\times}10^7$, $3.6{\times}10^7$ CFU/g in cabbage and $2.7{\times}10^3$ CFU/g in salt, respectively. The results of coliforms were detected as $2.4{\times}10^4$ CFU/g, and there was no Escherichia coli in any sample. Staphylococcus aureus was detected in cabbage as $9.9{\times}10^2$, $8.0{\times}10^1$, and $3.0{\times}10^3$ CFU/g, Bacillus cereus was also found in cabbage as $4.1{\times}10^3$ and $1.0{\times}10^1$ CFU/g. The results of Campylobacter jejuni and Vibrio paraheamolyticus were $2.4{\times}10^6$ and $1.0{\times}10^4$ CFU/g in cabbage, respectively. $1.0{\times}10^3$ CFU/g for Yersinia enterocolitica was determined in salt. In case of Listeria monocytogenes, the results were $1.5{\times}10^1$, $1.1{\times}10^2$, and $4.5{\times}10^1$ CFU/g in cabbage. Total batcteria ranged from $1.4{\times}10^1$ to $4.4{\times}10^5$ CFU/g were detected in salting solution, from $1.5{\times}10^4$ to $1.2{\times}10^8$ CFU/g in dehydrated salted-cabbage, from $9.4{\times}10^4{\sim}1.3{\times}10^8$ CFU/g in minced salted-cabbage. The results of E. coli in samples from different companies were different from one to anther. The results of the contamination of S. aureus and B. cereus showed positive in salting solution and dehydrated salted-cabbage at a portion of companies. V. paraheamolyticus was detected in salting solution. The contamination of Y. enterocolitica ranged from $9.5{\times}10^2$ to $1.8{\times}10^3$ CFU/g in salting solution, from $1.7{\times}10^1$ to $2.7{\times}10^2$ CFU/g in dehydrated salted-cabbage, from $1.2{\times}10^2$ to $1.3{\times}10^8$ CFU/g in minced salted-cabbage. The contamination of L. monocytogenes ranged from $8.0{\times}10^2$ to $1.7{\times}10^4$ CFU/g in salting solution, from $2.8{\times}10^2$ to $1.2{\times}10^4$ CFU/g in dehydrated salted-cabbage. During the manufacture processing of Kim chi, microorganisms were detected in cabbages salted in different concentrations of salt solution at 8%, 10%, 12% and 15% for 5-20 hours. As the results, $3.5{\times}10^5-1.7{\times}10^6$, $3.4{\times}10^5-2.5{\times}10^6$, $5.4{\times}10^5-2.3{\times}10^6$, $4.0{\times}10^5-2.3{\times}10^6$ CFU/g were detected for E. coli in samples at different treatment conditions. $1.9{\times}10^4-4.1{\times}10^4$, $4.1{\times}10^3-2.8{\times}10^4$, $1.5{\times}10^3-7.8{\times}10^3$, $2.2{\times}10^4-6.6{\times}10^4$ CFU/g were detected for S. aureus in samples at different treatment conditions. Salmonella typhimurium was detected in salted cabbage with various salt concentration after salting for 5 hrs, the result ranged from $2.5{\times}10^5$ to $3.8{\times}10^6$ CFU/g, and change of microorganism was the smallest in salted cabbage under the concentration of salting solution at 10% for 15 hours. The cabbage salted in 10% salting solution for 15 hours were washed with water for 2 and 3 times, with chlorine for 3 times, and with acetic acid for 3 times. E. coli was detected in the samples washed with water for 2 and 3 times, washed with chlorine for 3 times. The contamination of S. aureus was $3.0{\times}10^5$ CFU/g in the samples washed with water for 2 times, $5.6{\times}10^3$ CFU/g in the samples washed with acetic acid for 3 times, $3.6{\times}10^5$ CFU/g in the samples washed with water for 3 times and same amount in the samples washed with chlorine for 3 times. According to the results, the contamination of S. aureus was $5.6{\times}10^3$ CFU/g lower in samples washed with chlorine and acetic acid than that in samples washed with water. In case of S. typhimurium, it has been detected in samples washed with water and chlorine, $3.0{\times}10^1$ CFU/g as the lowest concentration among all the samples was measured in the samples washed with acetic acid for 3 times.

Microbial and Pathogenic Contamination of Ready-to-eat Fresh Vegetables in Korea (한국에 유통중인 신선편이 채소류의 미생물 품질 및 병원성 세균의 오염도 조사)

  • Bae, Young-Min;Hong, Yu-Jin;Kang, Dong-Hyun;Heu, Sung-Gi;Lee, Sun-Young
    • Korean Journal of Food Science and Technology
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    • v.43 no.2
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    • pp.161-168
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    • 2011
  • The purpose of this study was to evaluate microbiological contamination of fresh vegetables in Korea. Twenty types of vegetables were tested for total aerobic bacteria, coliforms, Escherichia coli, yeast and mold, and pathogenic bacteria such as Staphylococcus aureus, Listeria monocytogenes, Bacillus cereus, Salmonella, E. coli O157:H7, Cronobacter sakazakii, Shigella, and Campylobacter. Levels of total aerobic bacteria and coliform on 20 vegetables were between 3.74 and 8.04 log CFU/g, and 0.16 and 5.02 log CFU/g, respectively. The highest contamination levels of total aerobic bacteria were observed on water dropwort, sprouts, mungbean sprout, and ballflower root. There was no significant difference (p>0.05) in microbial contamination levels of total aerobic count, coliform, E. coli, yeast and mold between organic and nonorganic vegetables. When isolation methods using selective agars were applied, L. monocytogenes, B. cereus, Salmonella and Campylobacter were isolated from some fresh vegetable samples. Results of API kit tests showed that L. monocytogenes was identified on Chinese cabbage, cucumber, soybean sprouts, and iceberg lettuce while Salmonella was identified on Korean leek. Furthermore, Campylobacter jejuni was also identified in more than 50 of the 100 samples. However, when positive samples from API kit were tested for real-time PCR or 16S rRNA sequencing method, only B. cereus from perilla leaf, carrot, water dropwort, and sprouts showed positive results. These results indicate that selective agar and API kit detection methods might result in false positive results for some pathogens. Therefore, studies need to improve isolation or confirmation methods for such pathogens.