• 한국식품과학회지
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• 제53권1호
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• pp.1-11
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• 2021

본 논문에서는 최근 10년간(2010-2019년) 바이러스에 의한 식중독 통계 및 바이러스 검출법과 제어법에 대한 자료를 정리하여 나타냈다. 국내에서 지난 10년 동안 바이러스에 의해 발생한 식중독 사고 488건 중 94.9%가 노로바이러스에 의한 것으로 확인되어 노로바이러스가 국내에서 가장 주요한 식중독 바이러스로 생각된다. 노로바이러스를 검출하는 방법으로는 PCR을 이용한 방법이 주로 보고되고 있으며 현재(2020년 12월) 식품 공전에 등록된 방법(고시 제 2010-45호)에 따라 전기 영동을 기반으로 한 one-step RT PCR 및 semi-nested PCR 방법이 널리 이용되고 있다. 또한 최근 DNA sequencing 기술이 발달됨에 따라서 검출된 바이러스의 서열을 분석하여 이미 보고된 바이러스의 서열과 비교한 논문들이 많이 보고되었다. 이 외에도 real-time PCR을 적용한 논문들도 보고되고 있으며 앞으로는 전기 영동을 실시하는 conventional PCR을 대신하여 신속하게 정량 검출이 가능한 realtime PCR의 활용이 늘어날 것으로 생각한다. 기타 바이러스의 검출에 있어서도 역시 PCR을 활용한 방법이 주로 보고되고 있으며 multiplex PCR을 활용하여 여러 종류의 바이러스를 동시에 검출하고자 하는 노력이 이루어지고 있다. 더 나아가서, 자기 면역력 분리와 퀀텀닷 분석 방법 등을 이용한 신속 검출법이 제시되고 있어 앞으로 여러 식품에 오염된 바이러스를 현장에서 신속분리 및 검출하는데 이용할 수 있을 것으로 전망된다. 한편, 노로바이러스는 실험실에서 배양하기가 어렵기 때문에 노로바이러스 제어 연구는 대체재를 이용한 방법들이 주를 이루었다. 옴 가열을 포함한 여러 종류의 열처리와 초고압, 오존, 감마선, 광펄스 등의 비가열 처리를 이용하여 노로바이러스의 저감 정도를 살펴본 연구들이 최근 보고되었다. 일반적으로 물이나 완충용액보다는 식품 샘플에서 바이러스의 저감 정도가 낮게 관찰되었는데 식품 matrix가 이러한 물리적 처리에 간섭 효과를 나타내기 때문으로 생각되며 이를 극복하기 위해서는 여러 물리적, 화학적 처리를 조합하여 처리할 필요가 있다. 기타 바이러스 제어 연구에 있어서는 열, 광펄스, 고압력 등의 물리적 처리와 더불어 살균제(sanitizer)를 적용한 논문들이 보고되고 있으며 식중독 바이러스의 저감 메커니즘에 대한 체계적인 연구가 수행되고 있는 것을 확인하였다. 여러 물리적, 화학적 처리에 대해서 식중독 바이러스가 저항성을 갖는 이유와 사멸되는 메커니즘을 정확하게 이해한다면 추후 여러 식품에서의 바이러스에 대한 안전성을 확보하는데 도움이 될 것으로 사료된다.

• 한국수산과학회지
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• 제40권6호
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• pp.356-361
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• 2007

Recently, the development of various culture-independent identification techniques for environmental microbes has greatly enhanced our knowledge of microbial diversity. In particular, denaturing gradient gel electrophoresis (DGGE) of 16S rDNA fragments, generated using the polymerase chain reaction (PCR) is frequently used to examine the diversity of environmental bacterial populations. This method consists of direct extraction of the environmental DNA, amplification of the 200-600 bp 16S rDNA fragments with universal primers, and separation of the fragments according to their melting point on a denaturing gradient gel. In this study, we investigated the seaside microbial community in coastal areas of Busan, Korea, using culture-independent techniques. First, marine genomic DNA was extracted from seawater samples collected at Songjeong, Gwangahn, and Songdo Beaches. Then, PCR was used to amplify the bacterial 16S rDNA using universal primers, and DGGE was used to separate the amplified 500 bp 16S rDNA fragments. Finally, the tested 16S rDNA genes were further analyzed by sequencing. Based on these experiments, we found that DGGE analysis clearly showed variation among the regional groups. It can be used to monitor rapid changes in the bacterial diversity of various environments. In addition, the sequence analysis indicated the existence of many unculturable bacteria, in addition to Arcobacter, Pseudoaltermonas, and Vibrio species.

• 대한바이러스학회지
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• 제30권1호
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• pp.29-37
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• 2000

The aim of this study was to investigate viral etiology in dilated cardiomyopathy (DCM) by polymerase chain reaction (PCR) or nested reverse transcription PCR (RT-PCR), and characterize the enteroviral RNA presented in the clinical specimens. Twenty-eight paraffin-embedded heart tissue samples were assayed to detect cytomegalovirus, herpes simplex virus type 1, type 2, parvovirus, adenovirus, and enterovirus (EV) with each specific primer. Of these 28 patients (mean age: 27, M: 24, F: 4), 26 were histologically diagnosed as DCM and 2 as myocardial infarction (MI). Nested RT-PCR detected enteroviral RNA in 7 (26.9%) of 26 patients with DCM, and none of patients with MI. And none of DNA viruses tested were detected from the samples. Amplified products were also genotyped by single-strand conformation polymorphism (SSCP). Three subtypes can be differentiated from 7 clinical specimens. Furthermore, direct sequence analysis was performed to determine whether genetic variation of EV is present in the explanted heart tissues from patients with DCM. Although most of the sequences among the wild isolates have the greatest similarity to those of coxsackievirus B3, there are specific regions of variable sequences (no 490 - no 510). The data suggest that enterovirus may be a major viral pathogen for the DCM in Korea and nucleotide sequence data indicate that coxsackievirus B3 may be a leading etiologic agent of DCM.

• 대한임상검사과학회지
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• 제45권2호
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• pp.43-47
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• 2013

This study was designed to evaluate the prevalence of ESBL genes and monitor antimicrobial resistance pattern in Escherichia coli, isolated from a hospital and a community. We tested 200 E. coli strains isolated in the hospitals and community in Chungcheong area from January to March 2012. Antimicrobial susceptibilities were tested by using the disk diffusion method. A search for ESBL genes was conducted by PCR amplification, and the genotypes were determined by direct nucleotide sequence analysis of the amplified products. An Epidemiologic study was performed by repetitive extragenic palindromic sequence-based PCR (REP-PCR). The percentage of ESBL-producing isolates was 17% for hospital associated E. coli and 11% for community associated E. coli. The ESBL gene sequencing results showed that the most common ESBL in E. coli was CTX-M-14 (19/28), followed by CTX-M-15 (9/28). The REP-PCR study also showed the genetic diversity, but there was no difference between the hospital and community associated E. coli. In this study, the most common types of class A ESBLs identified were CTX-M in the hospital and the community in Chungcheong area. ESBL-producing E. coli isolates showed diverse clonality.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• 제26권1호
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• pp.45-52
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• 2000

Nowadays, there are a lot of evidence that mutation of the p53 tumor suppressor gene is one of the most common genetic abnormalities in neoplastic progression. In this study, we analyzed 20 specimens of oral tumors(squamous cell carcinoma 14 cases, ameloblastoma 3 cases, adenoid cystic carcinoma 2 cases, malignant schwannoma 1 case)using polymerase chain reaction and direct sequencing which used an automated DNA sequencer and software for detection of mutations. Polymerase chain reactions were performed with 4 sets of primers encompassing exon 5, 6, 7, 8, and direct sequencing method was employed. The results were as followings. 1. We detected 10 point mutations out of 20 specimens (50%). 2. The genetic alterations included 7 mis-sense mutations resulting in single amino acid subtitutions, 2 silent mutations, 1 non-sense mutations encoding a stop codon. 3. Mutations were mostly in exon 7(7 out of 10 mutations, 70%) and involved codons 225, 234, 235, 236, 238, 247. 4. Therse were 4 cases of $T{\rightarrow}A$ transversion, 2 cases of $C{\rightarrow}A$ transversion, $A{\rightarrow}G$ transition, 1 case of $C{\rightarrow}G$, $T{\rightarrow}G$ transversion respectively. 5. We could find out point mutations more conveniently using PCR - Automated Direct Sequencing method.

• BMB Reports
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• 제29권5호
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• pp.442-447
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• 1996

Mutations in codon 12, 13 and 61 of one of the three ras genes, H-ras, K-ras and N-ras, convert these genes into active oncogenes. The presence of H-ras gene mutations have important prognostic implications in various cancers. In this study, the H-ras gene mutations were investigated by two-step PCRRFLP in patients with bladder and stomach cancer. For the control experiments, T24 and SK2 cell lines were used. In a total of 36 bladder cancer patient cases, five (13.9%) mutations were found by this method. Of these, point 12 mutations were two (5.6%) cases and point 61 mutations were three (8.3%) cases. On the other hand, H-ras mutation was not found in 29 cases of stomach cancer. The results of the mutated H-ras gene confirmed by direct sequencing analysis were correlated well with PCR analysis. From the sensitivity test, the H-ras mutation was found to have about 0.2% of mutated DNA mingled in normal DNA. In conclusion, the H-ras mutation has a higher clinical Significance in bladder cancer than stomach cancer. Moreover the two-step PCR-RFLP method is sensitive, rapid and relatively simple for clinical work in detecting H-ras point mutations.

• 대한의생명과학회지
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• 제8권4호
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• pp.257-268
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• 2002

Chronic infection and inflammation have recently been implicated as important etiologic agents of atherosclerosis. Several agents have been suggested as possible candidates including cytomegalovirus (CMV), herpes simplex vims type 1 (HSV-1), Epstein-Barr virus (EBV), Chlamydia pneumoniae, and Helicobacter pylori. We evaluated the relationship between cytornegalovirus infection and atherosclerosis by in situ hybridization and polymerase chain reaction (PCR). We examined 23 subjects with atherosclerosis and 10 matched control subjects without atherosclerosis. CMV was detected by in situ hybridization in 60.9% (14/23) of aorta and 42.9% (9/21) of coronary arteries in subjects with atherosclerosis. It was also detected by PCR in 65.2% (15/23) of aorta and 52.4% (11/21) of coronary arteries. CMV was detected on areas showing early or advanced atheromatous changes. Cells morphologically identical to smooth muscle cells, endothelial cells, lymphocytes, fibroblasts, and Schwann cells were positively reacted with the CMV probe. However. none of the cells to which the probe hybridized contained inclusion bodies, thus strongly suggesting that the arterial wall may be a site of CMV latency. This result Indicates that CMV may potentially play a direct or indirect role in the pathogenesis of human atherosclerosis.

• Journal of Microbiology and Biotechnology
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• 제20권3호
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• pp.569-573
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• 2010

A quantitative detection method for Salmonella in seafood was developed using a SYBR Green-based real-time PCR assay. The assay was developed using pure Salmonella DNA at different dilution levels [i.e., 1,000 to 2 genome equivalents (GE)]. The sensitivity of the real-time assay for Salmonella in seeded seafood samples was determined, and the minimum detection level was 20 CFU/g, whereas a detection level of 2 CFU/ml was obtained for pure culture in water with an efficiency of ${\geq}85%$. The real-time assay was evaluated in repeated experiments with seeded seafood samples and the regression coefficient ($R^2$) values were calculated. The performance of the real-time assay was further assessed with naturally contaminated seafood samples, where 4 out of 9 seafood samples tested positive for Salmonella and harbored cells <100 GE/g, which were not detected by direct plating on Salmonella Chromagar media. Thus, the method developed here will be useful for the rapid quantification of Salmonella in seafood, as the assay can be completed within 2-3 h. In addition, with the ability to detect a low number of Salmonella cells in seafood, this proposed method can be used to generate quantitative data on Salmonella in seafood, facilitating the implementation of control measures for Salmonella contamination in seafood at harvest and post-harvest levels.

• 대한수의학회지
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• 제38권1호
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• pp.101-106
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• 1998

The present study attempted to apply polymerase chain reaction (PCR) to develop a rapid differential diagnosis among Marek's disease, reticuloendotheliosis and avian leukosis. The primers chosen to detect Marek's disease virus (MDV) flank the 132bp tandem direct repeat of the MDV genome. The primers selected for reticuloendotheliosis virus (REV) and avian leukosis virus (ALV) are based on proviral long terminal repeats of spleen necrosis virus and Rous-associated virus-2 genomes, respectively. The specific PCR products of MDV, REV and ALV were observed with each primer and the reaction was not cross-reacted among the viruses. MDV-specific DNA was also amplified from the MDV-induced lymphoma (MDCC-MSB1) but not from the REV-induced tumor and ALV-induced lymphoma (LSCC-1104B1). In addition, proviral DNA of REV from REV-induced tumor and proviral DNA of ALV from ALV-induced lymphoma were also amplified by REV-specific and ALV-specific PCRs, respectively. Therefore these three PCR methods may be used to rapidly differentiate among MDV, REV and ALV-associated tumors in diagnosis.

• Parasites, Hosts and Diseases
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• 제58권5호
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• pp.589-592
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• 2020

Cyclospora cayetanensis is an apicomplexan protozoan and is one of the most common pathogens causing chronic diarrhea worldwide. Eight stool samples with diarrheal symptom out of 18 Korean residents who traveled to Nepal were obtained, and examined for 25 enteropathogens including 16 bacterial species, 5 viral species, and 4 protozoans in stool samples as causative agents of water-borne and food-borne disease. Only C. cayetanensis was detected by nested PCR, and 3 PCR-positive samples were sequenced to confirm species identification. However, the oocysts of C. cayetanensis in fecal samples could not be detected by direct microscopy of the stained sample. As far as we know, this is the first report of a group infection with C. cayetanensis from a traveler visiting Nepal, and the second report of a traveler's diarrhea by C. cayetanensis imported in Korea.