• Journal of Food Hygiene and Safety
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• v.16 no.4
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• pp.258-263
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• 2001

Campylobacter spp. isolated and identified from the raw chicken carcasses in food service, were characterized. Total bacterial counts on the skins of raw chicken were 10$^4$~10$^{6}$ CFU/g and a total of 205 strains were primarily isolated after enrichment culture and selective culture of the sample with candle and microaerophilic chamber method. Among them, twenty eight strains of Gram-negative, catalase-positive and oxidase-positive were further isolated by the determination of biochemical characteristics. Only sixteen strains of them were finally identified as Campylobacter with PCR of pA and pB primers. Nine strains, more than half of them, have grown at 42$^{\circ}C$ and $25^{\circ}C$ and seven strains defined as thermophilic Campylobacter grew not at $25^{\circ}C$, but at 42$^{\circ}C$. Therefore, more careful management of food safety for raw chicken is needed in food service. Particularly, we should concern the raw chicken carcasses with high bacteria contamination, more them 10$^{5}$ CFU/g, which possibly includes Campylobacter spp. grown at low temperature.

• Korean Journal of Microbiology
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• v.38 no.3
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• pp.186-191
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• 2002

The usefulness of oligonucleotide DNA chip was evaluated for detection and primary level identification of major waterborne viruses in environmental samples. The enteric waterborne viruses included enterovirus, adenovirus, and rotavirus. Total intracellular RNA of 10 BGM cell plates showing virus-specific cytopathic effects was extracted at the third day after inoculation. The intracellular RNA was then subjected to either enterovirus-specific RT-PCR followed by sequencing analysis, or the DNA chip. Seven out of 10 positive samples in cell culture were positive but the other three sample were turned out to be negative by both RT-PCR and DNA chip analyses. Nucleotide sequencing results and the DNA chip hybridization results of the RT-PCR product were in complete agreement in the identification of the 7 positive samples as enteroviruses. Using the DNA chip, it took only 3∼4 hr to complete detection and primary level identification of target viruses and additional procedures such as gel electrophoresis or nucleotide sequencing were not necessary. We believe that the DNA chip system can be employed as a highly effective and new detection methodology for environmental viruses.

• Korean Journal of Microbiology
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• v.42 no.2
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• pp.89-94
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• 2006

The purpose of this study is to develop the strain-specific PCR primers for the identification of prevotella inter-media ATCC 49046 which is frequently used in the pathogenesis studies of periodontitis. The Hind III-digested genomic DNA of P. intermedia ATCC 49046 were cloned by random cloning method. The specificity of cloned DNA fragments were determined by Southern blot analysis. The nucleotide sequence of cloned DNA probes was determined by chain termination method. The PCR primers were designed based on the nucleotide sequence of cloned DNA fragment. The data showed that Pig6 DNA probe were hybridized with the genomic DNA from P. intermedia strains (ATCC $25611^T$ and 49046) isolated from the Westerns, not the strains isolated from Koreans. The Pig6 DNA probe were consisted of 813 bp. Pig6-F3 and Pig6-R3 primers, designed base on the nucleotide Sequences Of Pig6 DNA Probe, were 3150 specific to the only both P. intermedia ATCC $25611^T$ and P. intermedia ATCC 49046. In the other hand, Pig6-60F and Pig6-770R primers were specific to the only P. intermedia ATCC 49046. The two PCR primer sets could detect as little as 4 pg of chromosomal DNA of P. intermedia. These results indicate that Pig6-60F and Pig6-770R primers have proven useful for the identification of P. intermedia ATCC 49046, especially with regard to the maintenance of the strain.

• Journal of Life Science
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• v.22 no.2
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• pp.232-238
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• 2012

Kumjungsansung-Makgeolli$^{(R)}$ is a traditional Korean rice wine that is fermented from traditional nuruk and rice. In this study, we performed the PCR-denaturing gradient gel electrophoresis (DGGE) analysis targeting the 16S and 28S rRNA genes to characterize bacterial and fungal diversity during Makgeolli fermentation. The predominant bacteria in the PCR-DGGE profile during Makgeolli fermentation were Lactobacillus spp. (Lactobacillus curvatus, L. kisonensis, L. plantarum, L. sakei, and L. gasseri), Pediococcus spp. (P. acidilactici, P. parvulus, P. agglomerans, and P. pentosaceus), Pantoea spp. (P. agglomerans and P. ananatis), and Citrobacter freundii; these were identified on the base of analysis of 16S rRNA gene sequences. The dominant bacterium during Makgeolli fermentation was L. curvatus. The predominant fungi in PCR-DGGE profile during Makgeolli fermentation were Pichia kudriavzevii, Saccharomyces cerevisiae, Asidia idahoensis, Kluyveromyces marxianus, Saccharomycopsis fibuligera, and Torulaspora delbrueckii, and these were identified on the basis of analysis of 28S rRNA gene sequences. The dominant fungal species during Makgeolli fermentation changed from P. kudriavzevii at 0-2 days incubation to S. cerevisiae at 3-6 days incubation. This study suggests that PCR-DGGE analysis could be a suitable tool for the understanding of microbial diversity and structure during Makgeolli fermentation.

• Microbiology and Biotechnology Letters
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• v.32 no.3
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• pp.249-255
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• 2004

Chinese cabbage ('Baechu') Kimchi was fermented at the three different temperatures right after it was prepared. Samples were taken everyday for measuring bacterial populations, pH, and titratable acidity through the whole periods of fermentation up to 50 days. pH values and developed acidity were significantly affected by the fermenting temperatures of 4, 10, and $20^{\circ}C$, suggesting that different bacterial flora has been established by the temperatures exposed. The modified MRS agar containing vancomycin (300 $\mu$g/mL) was used for isolating the vancomycin-resistant LAB strains and 127 isolates were finally obtained. Of the LAB isolates, 13 isolates were subjected to the identification experiments based on the biochemical characteristics and the molecular-typing approach, an ITS-PCR, whether they belong to the genus Leuconostoc or not. The data obtained from API 50 CHL kit resulted that six isolates were identified as the members of Leuconostoc and six as Lactobacillus brevis strains except for a single isolate YKI 30-0401, which was not able to be identified because its biochemical traits were not matched to the database of API 50 CHL kit. It was noted that some isolates were distinct in a couple of some biochemical characteristics compared with those of the reference Leuconostoc species. To overcome the limitations experienced in the commercial identification products above, an ITS-PCR experiment was also conducted for the isolates, resulting that eight isolates belong to Leu. mesenteroides ssp. mesenteroides or dextranicum with a single band of 564 bp, and four to L. brevis strains. The ITS-PCR profiles clearly differentiated the closely-related LAB isolates for which same results were obtained by the biochemical method. This molecular approach, however, failed to produce the amplicons for the YKI 20-1003, leaving the strain unidentified. Judging from the identification data obtained in the Kimchi fermented at $4^{\circ}C$ or $10^{\circ}C$, Leuconostoc spp. including Leu. mesenteroides/dextranicum were likely predominant species in the earlier stage and L. brevis occurred at the high level through the whole period. By contrast, L. brevis, as one of the major flora, possibly lead the fermentation from the beginning in the Kimchi fermented at $20^{\circ}C$.}C$.

• Journal of Korea Association of Health Promotion
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• v.4 no.1
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• pp.68-74
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• 2006

For identification of four sibling species of the Anopheles hyrcanus complex found in Korea, the 5.8 rDNA-ITS2-28S rDNA region of each species was sequenced and the species-specific primers wee designed The amplified PCR products obtained from each species were analyzed by agarose gel electrophoresis. The result showed a single species- specific band, I.e. 559bp, 432bp, 322bp and 192bp for An. sinensis, An. sp., An. lesteri and An. pullus, respectively. In conclusion, the species-specific PCR primers designed from ITS2 variable regions functioned successfully and specifically, and can be applied as a useful tool for identifying species of the Anopheles hyrcanus complex found in Korea.

• Proceedings of the Korean Society for Food Science of Animal Resources Conference
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• 2005.10a
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• pp.346-349
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• 2005

병원성미생물을 분리${\cdot}$동정하는 방법으로 선택배지를 이용하는 검사방법은 4${\sim}$7일의 시간이 소요되는 단점이 있으며, 기존의 PCR방법은 대부분 단일 병원성 미생물만을 검출할 수 있는 반면 본 연구에서 사용된 시스템은 6종류의 세균성 식중독균을 한 번의 분석으로 확인할 수 있었기 때문에 비용과 분석시간을 대폭 절감하는 효과가 있었다. Fig. 1은 6종류의 세균성 식중독균이 혼합된 시료에 각각의 단일 프라이머를 이용하여 특이성을 확인한 결과이며, Fig. 2는 멀티플렉스 프라이머를 이용하여 각각의 세균성 식중독균 열 추출 시료에 대한 특이성을 확인한 결과이다. PCR 산물들이 사다리(ladder) 형태로 증폭됨으로써 동시에 6종류의 세균성 식중독균을 용이하게 확인할 수 있었다(Lane 1${\sim}$Lane 6). 또한 3종류의 세균성 식중독균이 혼합된 열 추출시료에서도 특이적인 PCR 증폭 산물들을 탐지할 수 있었다(Lane 10 & Lane 11).

• Research in Plant Disease
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• v.23 no.3
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• pp.241-248
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• 2017

The purpose of this study was to develop rapid methods for distinguishing between Heterodera schachtii and H. glycines detected from chinese cabbage fields of highland in Gangwon, Korea. To do this, we performed PCR-RFLP and PCR with the primers set developed in this study for GC147, GC408 and PM001 population, H. schachtii, and YS224, DA142 and BC115 population, H. glycines. Eight restriction enzymes generated RFLP profiles of mtDNA COI region for populations of H. schachtii and H. glycines, repectively. As a result, treatment of two restriction enzymes, RsaI and HinfI, were allowed to distinguish H. schachtii from H. glycines based on the differences of DNA band patterns. The primer set, #JBS1, #JBG1 and #JB3R, amplified specific fragments with 277 and 339 bp of H. schachtii, 339 bp of H. glycines, respectively, while it did not amplify fragments from three root-knot nematodes and two root-lesion nematodes. Thus, the primer set developed in this study could be a good method, which is used to distinguish between H. schachtii and H. glycines.

• Journal of Food Hygiene and Safety
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• v.18 no.4
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• pp.229-236
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• 2003

The five different foodborne pathogenic bacterial genera of Escherichi, Salmonella, Shigella, Vibrio and Listeria are important sources of foodpoison. However, the method was not developed to simultaneously differentiate these five bacteria at molecular level. The optimized concentrations of the four major PCR cocktail components of $MgCl_2$, dNTPs, primers and template DNA were determined when ERIC (enterobacterial repetitive intergenic consensus)-PCR reactions were carried out to differentiate the five differnet foodborne pathogenic bacteria. The optimized concentration of $MgCl_2$ was determined to be 2 mM in order to obtain a consistent fingerprinitng pattern. The similar fingerprinting pattern was obtained when ERIC primers and dNTPs were added up to the concentrations of 2 ${\mu}M$ and 200 ${\mu}M$, respectively. As for template DNA, the numbers of PCR fragments were not affected, but their intensities were increased as the concentrations of the DNA were increased.

• The Korean Journal of Mycology
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• v.44 no.2
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• pp.103-107
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• 2016

To establish a rapid and accurate detection of Phytophthora pinifolia, which is a quarantine pathogenic fungus in Korea, a species-specific primer was developed based on the ras-related protein (Ypt1) gene. Species-specific primer based on the DNA sequences of Ypt1 gene amplified 193 bp polymerase chain reaction (PCR) product for P. pinifolia. The primer pair yielded the predicted PCR product size exactly in testing with target pathogen DNAs, but not from the other 10 species of Phytophthora and 14 species of other phytopathogenic fungi. The primer pair also showed only the species-specific amplification curve on realtime PCR on target pathogen DNA. The detection sensitivity of real time PCR using species-specific primer pair was 10 to 100 times higher than conventional PCR, with 1 to $10pg/{\mu}L$.