• 디지털융복합연구
• /
• 제14권4호
• /
• pp.371-378
• /
• 2016

본 연구에서는 한국의 닭에서 분리된 E. coli 균주들로부터 퀴놀론계 항생제 내성을 나타내는 균주를 분리 동정하고 그 내성 기전과 유병률에 관하여 조사하였다. 또한 multilocus sequence typing (MLST)을 이용하여 E. coli 균주들의 분자생물학적 성상을 분석하였다. 항생제 감수성 테스트에서 63.5% (54/85) 의 E. coli 균주들에서 퀴놀론계 항생제 내성률을 보였다. 또한 퀴놀론계 항생제 내성을 보이는 54개 모두에서 gyrA 유전자의 sense mutations과 parC 유전자의 $57^{th}$, $80^{th}$, or $84^{th}$residues에서 점돌연변이를 관찰할 수 있었다. MLST를 통한 분석에서 E. coli ST는 parE 유전자의 염기치환과 깊은 상관관계를 보이는 것으로 관찰되었다. 이 결과들을 바탕으로 우리가 먹는 가축 및 가금류에 대한 무분별한 항생제 사용은 항생제 내성균의 증가와 유전변이를 초래함을 알 수 있었다. 따라서 식용 동물에 대한 지속적인 감시와 모니터링을 통하여 항생제 내성균의 확산방지를 통제하는 것이 필요할 것으로 사료된다.

• Journal of Microbiology and Biotechnology
• /
• 제20권10호
• /
• pp.1457-1462
• /
• 2010

Three types of serotypically atypical Shigella flexneri isolates were collected between 2007 and 2008 from Korean patients at the Korea National Institute of Health (NIH). These atypical isolates were characterized and compared with serologically typical S. flexneri. The first grouping of 11 atypical isolates displayed agglutination only with polyB antiserum and exhibited no reaction with any typing or grouping sera (PolyB:un). The second group of 3 isolates displayed reactions with typing sera IV, but also did not bind with any grouping sera (IV:un). The third group of 14 isolates exhibited a plural agglutination pattern, reacting with typing sera II, and two grouping sera (II:(3)4,7(8)). Amongst these atypical isolates, isolates belonging to IV:un and II:(3)4,7(8) exhibited greater antibiotic resistance, in particular to ampicillin, streptomycin, and trimethoprim-sulfamethoxazole, than typical S. flexneri strains. Furthermore, all II:(3)4,7(8) strains harbored integrons. This study suggests that these multiple antibiotic-resistant atypical S. flexneri are new subserotypes of S. flexneri that await further serological classification.

• BMB Reports
• /
• 제32권6호
• /
• pp.529-534
• /
• 1999

Asymmetric PCR and single-strand conformation polymorphism (SSCP) methods were combined to analyze human leukocyte antigen (HLA)-DRB allele polymorphism. Asymmetric PCR amplification was applied to generate single-stranded DNA (ssDNA) using the nonradioactive oligonucleotide primers desinged for the polymorphic exon 2 region. The conformational differences of ssDNAs, depending on the allele type, were analyzed by nondenaturing polyacrylamide gel electrophoresis and visualized by ethidium bromide staining. The ssDNAs were clearly separated from double-stranded DNA without interference and obviously migrated depending on their allele type. This method was applied to the genomic DNA either from homozygous or from heterozygous cell lines containing the DR4 allele as template DNA using DR4-specific primers, and satisfying results were obtained. Compared to the standard PCR-SSCP method, this asymmetric PCR-SSCP method has advantages of increased speed, reproducibility, and convenience. Along with PCR-SSP or sequence-based typing, this method will be useful in routine typing of HLA-DRB allele.

• 한국동물위생학회지
• /
• 제32권3호
• /
• pp.257-264
• /
• 2009

Subtyping of Brucella abortus isolates is epidemiologically important for monitoring of bovine brucellosis outbreaks. Pulsed-field gel electrophoresis (PFGE) is considered as a gold standard of molecular typing methods to study the DNA polymorphisms of bacteria. In this study, we analyzed using PFGE the DNA fragment profiles of B. abortus isolated in Gyeongbuk province from 1998 to 2006. The genomic DNA was digested with the restriction endonuclease Xba I, Xho I and Smi I followed gel electrophoresis. No distinguishable patterns of the genomic DNA digested with Xba I and Xho I were observed among the field isolates of B. abortus tested in this study. But Smi I restriction enzyme resulted in two PFGE patterns consisting of 13-15 bands that ranged in size from 33 to 668bp by standard marker. The cluster analysis by DNA fingerprinting software showed 93.75% similarity between two PFGE patterns. No different PFGE patterns were recognized among the isolates originated from various years, regions and cow breeds.

• Journal of Microbiology and Biotechnology
• /
• 제26권9호
• /
• pp.1643-1649
• /
• 2016

The aims of this study were to characterize the molecular epidemiological profiles of CTX-M-producing uropathogenic Escherichia coli isolates from a tertiary hospital in Daejeon, Korea, and to investigate the genetic diversity and compare the prevalence of sequence types (STs) in different areas. Extended spectrum β-lactamase-producing E. coli strains isolated from urine were analyzed for CTX-M, integrons, and insertion sequence common regions (ISCRs) by PCR and sequencing. Multilocus sequence typing (MLST), pulsed-field gel electrophoresis (PFGE), phylogenetic analysis, and rep-PCR were also used for molecular typing of the isolates. Of 80 CTX-M producers, 31 and 46 expressed CTX-M-15 and CTX-M-14, respectively. MLST analysis indicated that the most prevalent ST was ST131 (n = 34, 42.5%), followed by ST38 (n = 22, 27.5%), ST405 (n = 8, 10.0%), and ST69 (n = 6, 7.5%). Most CTX-M producers harbored class 1 integrons. ST131 strains belonged to phylogenetic group B2 and showed identical rep-PCR patterns, whereas ST69, ST38, and ST405 strains belonged to phylogenetic group D; the ST38 and ST405 strains displayed the same rep-PCR pattern, respectively. ST131 and ST38 isolates showed 21 and 19 distinct types, respectively, by PFGE. In Daejeon, D-ST38 CTX-M-14 producers were relatively more prevalent than in other countries and Korean cities. Our results indicate that CTX-M-producing E. coli isolates belonged mostly to ST131 or ST38 and were more related to hospital-onset than to community-onset infections and that the bla_CTX-M gene may vary according to the ST.

• Fisheries and Aquatic Sciences
• /
• 제9권2호
• /
• pp.70-74
• /
• 2006

DNA sequence-based typing is considered a robust tool for the discrimination of dinoflagellate species because of the availability of extensive rDNA sequences. Here, we present a rapid, cost-effective DNA-sequencing technique for various PCR products. This sequencing strategy relies on 'nested' or 'tailed' primer labeled with near-infrared dye, and uses a minimal volume of unpurified PCR product (ca. $5{\mu}L$) as the DNA template for sequencing reactions. Reliable and accurate base identification was obtained for several hundred PCR fragments of rRNA genes. This quick, inexpensive technique is widely applicable to sequence-based typing in clinical applications, as well as to large-scale DNA sequencing of the same genomic regions from related species for studies of molecular evolution.

• Journal of Microbiology
• /
• 제42권2호
• /
• pp.80-86
• /
• 2004

Pulsed-field gel electrophoresis (PFGE) typing was applied to the epidemiological investigation of 21 Candida tropicalis isolates collected from urine specimens of 11 patients and one healthcare worker, in an intensive care unit (ICU) over a 4-month period. Seventeen epidemiologically unrelated strains from 14 patients were also tested to determine the discriminatory power of PFGE. PFGE typing consisted of electrophoretic karyotyping (EK) and restriction endonuclease analysis of genomic DNA (REAG), using two restriction enzymes (BssHII and SfiI). The EK pattern was the same in all 38 isolates, while REAG using SfiI separated the isolates into nine types. However, 16 different PFGE types were iden-tified by REAG with BssHII, and the same results were obtained when the results of both REAG tests were combined. In serial urinary isolates from 10 patients, all strains from each patient had the same PFGE pattern. While the epidemiologically unrelated strains from 14 patients consisted of 13 different PFGE types, the 20 isolates from the 11 ICU patients fell into only two PFGE types (types Cl and C2), and these apparently originated from the two different outbreaks. All strains of type Cl (n = 12) were isolated from six patients, between November 1999 and January 2000, and all of the type C2 strains (n=8) were isolated from five patients, during January and February 2000. This study shows two con-secutive clusters of C. tropicalis candiduria in an ICU, defined by PFGE typing, and also demonstrates that a PFGE typing method using BssHII is perhaps the most useful method for investigating C. tropi-calis candiduria.

• 생명과학회지
• /
• 제15권2호
• /
• pp.161-168
• /
• 2005

2001년부터 2003년까지 부산지역의 냉각탑수에서 분리한 L. pneumophila (serogroup 1) 39균주에 대해 제한효소 SfiI 처리한 경우 PFGE 양상은 dice coefficient $<65\%$의 유사성을 가진 band를 Awl로 10개의 pulsotype으로 나누었고, 가장 많았던 유형은 E pulsotype으로 39주중 18주로 $46.2\%$를 나타내었으며, 그 외 A pulsotype $17.9\%$, C pulsotype $15.4\%$, F pulsotype $7.7\%$ 및 B, D, G, H, I, J pulsotype이 각각 $2.6\%$로 유전적 양상이 다양하였다. 제한효소 NotI 처리한 경우 PFGE 양상은 dice coefficient $<60\%$의 유사성을 가진 band를 $a\~h$로 8개의 pulsotype으로 나누었고, 가장 많았던 유형은 f pulsotype이 $38.5\%$였으며 그 외 d pulsotype $20.5\%$, e pulsotype $17.9\%$, a pulsotype $10.3\%$, h pulsotype $7.7\%$ 및 b, c, g pulsotype이 각각 $2.6\%$를 차지하였다. 본 연구를 통하여 부산지역에서 분리되는 레지오넬라균에 대한 유전자 유형분석 결과를 데이터베이스화하여, 레지오넬라증의 집단 발생 또는 산발적인 질병 발생이 있을 경우 분자학적 측면에서 사람과 환경에서 분리된 균으로부터 감염원과 감염경로를 규명하는 역학적인 도구로써 이용 가능할 것으로 사료되었다.

• Journal of Microbiology and Biotechnology
• /
• 제16권1호
• /
• pp.68-76
• /
• 2006

Despites many attempts to explore the microbial diversity in kimchi fermentation, the predominant flora remains controversial to date. In the present study, major lactic acid bacteria (LAB) were investigated in Chinese cabbage kimchi in the early phase of fermention. For the samples over pH 4.0, viable cell counts of Leuconostoc and Pediococcus were $10^6\;cfu/ml$ and below $10^2\;cfu/ml$, respectively, and 20 isolates out of 172 were subjected to a biochemical identification (API 50 CH kit) as well as molecular-typing methods including ITSPCR with a RsaI digestion and 16s rRNA gene sequence analysis for species confirmation. Seven isolates were nicely assigned to Lb. brevis, 6 to Leuconostoc spp. (2 mesenteroides, 2 citreum, I carnosum, I gasicomitatum), 4 to Weissella (3 kimchii/cibaria, 1 hanii) and 2 to other Lactobacillus spp. (1 farciminis, 1 plantarum). On the other hand, the biochemical identification data revealed 9 strains of Lb. brevis, 6 strains of Leuconostocs,2 strains of Lb. plantarum and 1 strain each of Lb. coprophilus and Lactococcus lactis. However, a single isolates, YSM 16, was not matched to the ITS-PCR database constructed in the present study. Two Lb. brevis strains by API 50 CH kit were reassigned to W kimchii/cibaria, Lb. coprophilus or W hanii, respectively, judging from the results by the above molecular typing approaches. As a whole, the identification data obtained by the biochemical test were different from those of ITS-PCR molecular method by about $63\%$ at genus-level and $42\%$ at species-level. The data by the ITS-PCR method conclusively suggest that predominant LAB species is probably heterolactic Lb. brevis, followed by W kimchii/cibaria, Leuc. mesenteroides, and Leuc. citreum, in contrast to the previous reports [3] that Leuc. mesenteroides is the only a predominant species in the early phase kimchi fermentation.

• Animal cells and systems
• /
• 제11권2호
• /
• pp.165-168
• /
• 2007

The nucleotide sequences of a male-specific marker sex determining region Y (SRY) gene and a mitochondrial cytochrome B (CYTB) gene were characterized and analyzed to establish a molecular method for identification of species and sex of Korean roe deer (Capreolus pygargus tianschanicus). Similarity search result of SRY sequences showed very similar result to those reported in Moose (Alces alces) and Reindeer (Rangifer tarandus), both of which had 95.9% similarity in identity. CYTB genes were very similar to those reported in Siberian roe deer (C. pygargus pygargus) which had 98.6% similarity and not to European roe deer (C. capreolus), suggesting that the DNA samples tested were of Siberian roe deer lineage. Polymerase chain reaction (PCR)-based sex typing successfully discriminated between carcasses of male and female roe deer. Males had SRY band on agarose gels and females did not. The result of this molecular sex typing provided similar information with that obtained by genital organ observation. Therefore, this molecular method using male specific marker SRY and mitochondrial CYTB genes would be very useful for identification of the species and sex of the carcass remains of roe deer.