• Journal of Food Hygiene and Safety
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• v.26 no.2
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• pp.114-119
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• 2011

In this study, two duplex real-time PCR approach with melting curve analysis is presented for the detection of Escherichia coli O157:H7, Listeria monocytogenes, Salmonella spp. and Staphylococcus aureus, which are important food-borne bacterial pathogens usually present in fresh and/or minimally processed vegetables. Reaction conditions were adjusted for the simultaneous amplification and detection of specific fragments in the ${\beta}$-glucuronidase (uidA, E. coli), thermonuclease (nuc, S. aureus), hemolycin (hly, L. monocytogenes) and tetrathionate reductase (ttr, Salmonella spp.) genes. Melting curve analysis using a SYBR Green I real-time PCR approach showed characteristic $T_m$ values demonstrating the specific and efficient amplification of the four pathogens; $80.6{\pm}0.9^{\circ}C$, $86.9{\pm}0.5^{\circ}C$, $80.4{\pm}0.6^{\circ}C$ and $88.1{\pm}0.11^{\circ}C$ for S. aureus, E. coli O157:H7, L. monocytogenes and Salmonella spp., respectively. For all the pathogens, the two duplex, real-time PCR was equally sensitive to uniplex real-time PCR, using same amounts of purified DNA, and allowed detection of 10 genome equivalents. When our established duplex real-time PCR assay was applied to artificially inoculated fresh lettuce, the detection limit was $10^3$ CFU/g for each of these pathogens without enrichment. The results from this study showed that the developed duplex real-time PCR with melting curve analysis is promising as a rapid and cost-effective test method for improving food safety.

• Journal of Food Hygiene and Safety
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• v.32 no.2
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• pp.129-134
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• 2017

Diarrheagenic Escherichia coli is now recognized as an important cause of diarrhea, hemorrhagic colitis and hemolytic-uremic syndrome (HUS) worldwide. E. coli were isolated from 80 of 356 (22.5%) chicken and chilled chicken products in Korea. Fifteen virulence genes specific for pathogenic E. coli, including Shiga toxin-producing E. coli (STEC), enteropathogenic E. coli (EPEC), enterotoxigenic E. coli (ETEC), enteroinvasive E. coli (EIEC) and enteroaggregative E. coli (EAEC), were examined by multiplex PCR. STEC virulence markers were detected for eaeA (20.0%), escV (21.3%), stx1 (3.8%), ent (2.5%), EHEC-hly (1.3%), stx2 (1.3%), EAEC virulence marker (astA) was detected in 32.5%. ETEC and EIEC were not detected. STEC serotypes O152, O1, O116, O26, O25, O119 and O153 were found in chicken samples. This suggests the importance of diarrheagenic Escherichia coli control in raw chicken and chilled chicken food for food safety.

• Korean Journal of Veterinary Service
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• v.26 no.2
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• pp.105-111
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• 2003

From February 2000 to December 2001, A total of 1,785 samples was taken from beef and pork carcasses in Seoul. Seven(0.69%) Listeria spp. were isolated from the 1,014 of beef carcasses, and five(0.65%) were isolated from the 771 of pork carcasses. The isolates were identified L monocytogenes by API listeria, and VIDAS LMO kit, serological test and PCR assay were preformed. A total 12 strains of L monocytogenes were isolated form samples tested and all of the strains were classified into serotype 1. PCR primers are selected to amplify a 520-base pair(bp) DNA fragment from the listeolysin O gene(hlyA) of Listeria monocytogenes. A 520-bp product was detected in PCR with DNA from L monocytogenes, but not from the other Listeria species tested.

• Microbiology and Biotechnology Letters
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• v.48 no.2
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• pp.121-128
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• 2020

Diarrhea is a major public health concern associated with pathogenic Escherichia coli infections. Food-borne pathogenic E. coli can lead to large diarrheal outbreaks and hence, there is a need to estimate the frequency of pathogenic E. coli load in the various types of meat available in markets. In the present study, we classified and characterized diarrheagenic E. coli isolates collected from 399 raw meat samples from retail sources in Korea. Shiga toxin-producing E. coli (STEC) were detected in 11 (9.7%) samples, including nine strains (8.0%) in beef and two strains (1.8%) in chicken. The frequency of the detected virulence markers were as follows: astA, 28.3%; escV,18.6%; eaeA,17.7%; ent, 7.0%; EHEC-hly, 4.4%; stx1, 3.5%; and stx2, 3.5%. We did not observe any typical EPEC, EIEC, or ETEC virulence determinants in any of the samples. The STEC serotype O26 was detected in one sample, but no other serogroups (O91, O103, O128, O157, O145, O111, and O121) were found. Further research is needed to better understand the virulence mechanism of STEC serotypes, their ecology, and prevalence in animals, food, and the environment. These results will help improve risk assessment and predict the sources of food poisoning outbreaks.

• Journal of Microbiology
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• v.44 no.1
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• pp.92-97
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• 2006

We developed an mPCR assay for the simultaneous detection, in one tube, of Escherichia coli O157:H7, Salmonella spp., Staphylococcus aureus and Listeria monocytogenes using species-specific primers. The mPCR employed the E. coli O157:H7 specific primer Stx2A, Salmonella spp. specific primer Its, S. aureus specific primer Cap8A-B and L. monocytogenes specific primer Hly. Amplification with these primers produced products of 553, 312, 405 and 210 bp, respectively. All PCR products were easily detected by agarose gel electrophoresis, and the sequences of the specific amplicons assessed. Potential pathogenic bacteria, in laboratory-prepared and four commercially available kimchi products, were using this mPCR assay, and the amplicons cloned and sequenced. The results correlated exactly with sequences derived for amplicons obtained during preliminry tests with known organisms. The sensitivity of the assay was determined for the purified pathogen DNAs from four strains. The mPCR detected pathogen DNA at concentrations ranging from approximately 0.45 to $0.05\;pM/{\mu}l$. Thus, this mPCR assay may allow for the rapid, reliable and cost-effective identification of four potentially pathogens present in the mixed bacterial communities of commercially available kimchi.

• Korean Journal of Fisheries and Aquatic Sciences
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• v.53 no.6
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• pp.870-877
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• 2020

Twenty-three Bacillus cereus strain isolated from commercial jeotgal were investigated for 11 toxin genes and susceptibility to 25 different antimicrobials. The hemolytic enterotoxins hblA, hblC, and hblD were detected in 13.0%, and non-hemolytic enterotoxins nheA, nheB, and nheC were detected in 26.1%, 100%, and 100% of the isolates, respectively. The positive rates of cytK, entFM, becT, hlyII, and ces were 73.9%, 60.9%, 26.1%, 8.7%, and 0.0%, respectively. According to the disk diffusion susceptibility test, all of the strains studied were resistant to cefuroxime, followed by cefoxitin (78.3%), oxacillin (78.3%), ampicillin (69.6%), penicillin G (69.6%), and amoxicillin (65.2%). However, all the strains were susceptible to 11 other antimicrobials, including amikacin, chloramphenicol, and ciprofloxacin. The average minimum inhibitory concentrations of amoxicillin, ampicillin, and cefuroxime against B. cereus were 462.9, 235.0, and 135.0 ㎍/mL, respectively. These results highlight the need for sanitizing commercial jeotgal, and provide evidence to help reduce the risk of jeotgal contamination by antimicrobial-resistant bacteria.

• Ocean and Polar Research
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• v.30 no.3
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• pp.215-224
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• 2008

A piston core (MR06-04 PC23A) collected from the northern continental slope in the central Bering Sea has recorded the high-resolution millennial-scale variation of calcium carbonate ($CaCO3$) content during the last 65 kyr. An estimation of the age of the core sediments was carried out by using the lithologic correlation of the deglacial laminated layers with a neighboring core (HLY02023JPC), complementing the last appearance datum of both Lychnocanoma nipponica sakaii (54 kyr) and Amphimelissa setosa (85 kyr). The probable age of core MR06-04 PC23A was approximately younger than 65 kyr. Two distinct events of a significant increase of $CaCO3$ in the deglacial laminated sediments clearly correspond to MWP1A and MWP1B in the Bering Sea (Gorbarenko et al. 2005) and to T1ANP and T1BNP in the North Pacific (Gorbarenko 1996). These pronounced peaks of $CaCO3$ contents result from the elevated carbonate production in the surface water and the subsequent weakened dilution due to terrestrial input, along with an enhanced oxygen minimum zone. The $CaCO3$ contents are low (${\sim}2%$) during the last glacial period mainly because of a low carbonate production caused by an expanded sea-ice cover and an increased dilution by terrigenous particles due to their closer distance to the continent during the sea-level low stand. The occurrence of seven distinct $CaCO3$ peaks in core MR06-04 PC23A is remarkable during MIS 3 and MIS 4, and they most likely correlate to the short-term millennial Dansgaard-Oeschger events.

• Korean Journal of Veterinary Service
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• v.30 no.3
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• pp.353-362
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• 2007

A total of 1,354 samples was collected from bovine and porcine carcass from January 2005 to December 2006 in a slaughter house. Twenty five strains(1.8%) of Listeria monocytogenes were isolated from 1,354 samples using selective media. Ten(1.4%) L monocytogenes were isolated from the 677 of bovine carcasses, and 15(2.2%) were isolated from the 677 of porcine carcasses. Among 15 L mono-cytogenes from porcine, 11 siolates were serovars 1/2c, followed by 1/2b (3 strains, 20.0%) and 1/2a(1 strain) Out of 10 bovine samples, positive cases in 1/2a were 9 strains (90.0%), 1/2b were 1 strains(10.0%). PCR primers were selected to amplify a 520-base pair(bp) DNA fragment from the listeolysin O gene (hlyA) of L mono-cytogenes. A 520-bp product was detected in PCR with DNA from L monocytogenes, but not from the other Listeria species tested. A total of 25 L monocytogenes strains were analysed by PFGE after digestion with Apa I. PFGE analysis of genomic DNA showed the $14{\sim}18$ fragments ranging in size from 30 to 550 kb, resulting in 14 patterns.

• Clinical and Experimental Pediatrics
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• v.55 no.11
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• pp.420-423
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• 2012

Purpose: We phylogenetically analyzed the Escherichia coli strains isolated from children with urinary tract infection (UTI) in 2 regions of Korea. Virulence factors (VFs) and antibiotic resistance of the strains were also determined to compare the possible differences. Methods: A total of 138 E. coli strains were collected from the 2 regions; Gyeongin (78 strains) and Gyeongnam (60 strains). The phylogenetic groups were determined using the triplex polymerase chain reaction (PCR) method and multiplex PCRs were used to detect 7 VFs genes (fimH, papC, iutA, hlyA, sfa/focDE, afa/draBC, and kpsMT II). We also tested for antibiotic resistance. Results: Phylogenetic groups, B2 (61.6%) and D (26.8%), comprised the majority of all isolated strains. Regional comparisons revealed that more B2 strains and fewer non-B2 (A+B1+D) strains were found in Gyeongnam, than in the Gyeongin region (P=0.033), and certain VFs were predominantly detected in Gyeongnam (P<0.05). Neither regional nor phylogenetic differences, in antibiotic resistance of the strains, were significant. Conclusion: We were able to confirm that the geographic location is an important determinant of the distribution of the phylogenetic groups and VFs among the E. coli strains that cause UTI in children.

• Korean Journal of Veterinary Service
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• v.28 no.1
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• pp.71-79
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• 2005

In this study, a total of 2,119 samples was taken from bovine feces and carcass from March 2002 to December 2003. And those were examined for the presence of enterohemorrhagic E coli O157:H7. The properties of the isolates were characterized for biochemical features, serotypes, virulence genes and antimicrobial susceptibility. Forty five strains($3.7\%$) of E coli O157:H7 were isolated from 1,208 fecal samples and were not detected in carcass using immunomagnetic separation technique and selective media. In multiplex PCR using stx1, stx2, eaeA and hlyA primers, the amplified bands at 180 bp, 255bp, 384bp and 534bp were observed, respectively. In antimicrobial susceptibility test, all isolates were susceptible to amoxicillin/clavulanic acid and cefazolin. The isolates were most resistant to sulfisoxazole($24.4\%$), followed by streptomycin($22.2\%$), tetracycline($20.0\%$). Eight strains($17.8\%$) of 45 isolates showed the multi-resistant patterns with over 3 drugs.