• 한국동물위생학회지
• /
• 제45권3호
• /
• pp.171-180
• /
• 2022

The aim of this study was to investigate the prevalence of antimicrobial resistance and virulence factors in staphylococci isolated from canine otitis externa. A total 295 causative microorganisms were isolated. The most common isolated species were Staphylococcus (S) pseudintermedius (94 isolates) followed by Pseudomonas aeruginosa (60 isolates), S. schleiferi (25 isolates), Escherichia coli (23 isolates) and Proteus mirabilis (20 isolates). Staphylococci isolates were showed high resistance to penicillin (78.6%), erythromycin (55.9%), tetracycline (52.4%), clindamycin (51.7%) and ciprofloxacin (42.8%). Of the 145 staphylococci isolates, 49 (33.8%) methicillin-resistant staphylococci (MRS) were observed, distributed among S. pseudintermedius (n=34), S. schleiferi (n=6), S. epidermis (n=4), S. hominis (n=2), S. aureus, S. caprae and S. saprophyticus (n=1, respectively). Forty-three (87.8%) of 49 MRS and 10 (10.4%) of 96 methicillin-susceptibility staphylococci harbored mecA gene. About 80% of MRS were multidrug-resistant with resistance to at least one antibiotic in three or more antibiotic classes. Resistance genes blaZ (93/114, 81.5%), ermB (35/81, 43.2%), ermC (3/81, 3.7%), aacA-aphD (50/54, 92.5%), tetM (69/76, 90.7%) and tetK (6/76, 7.8%) were detected among resistant isolates. Virulence factors genes lukF and lukS were found in 100%(145/145) and 43.4%(63/145), respectively. Genes encoding ermA, eta, etb and tsst were not detected. To the best of our knowledge, this is the first study which investigated for the presence of genes encoding antimicrobial resistance and staphylococcal toxins in staphylococci isolated from canine otitis externa. A continuous monitoring and surveillance program to prevent antimicrobial resistance in companion animals is demanded.

• BMB Reports
• /
• 제57권3호
• /
• pp.135-142
• /
• 2024

DNA methylation is one of the most extensively studied epigenetic regulatory mechanisms, known to play crucial roles in various organisms. It has been implicated in the regulation of gene expression and chromatin changes, ranging from global alterations during cell state transitions to locus-specific modifications. 5-hydroxymethylcytosine (5hmC) is produced by a major oxidation, from 5-methylcytosine (5mC), catalyzed by the ten-eleven translocation (TET) enzymes, and is gradually being recognized for its significant role in genome regulation. With the development of state-of-the-art experimental techniques, it has become possible to detect and distinguish 5mC and 5hmC at base resolution. Various techniques have evolved, encompassing chemical and enzymatic approaches, as well as third-generation sequencing techniques. These advancements have paved the way for a thorough exploration of the role of 5hmC across a diverse array of cell types, from embryonic stem cells (ESCs) to various differentiated cells. This review aims to comprehensively report on recent techniques and discuss the emerging roles of 5hmC.

• Journal of Microbiology
• /
• 제40권3호
• /
• pp.211-218
• /
• 2002

A 4.8-kb DNA fragment encoding the P-450 type hydroxylase and ferredoxin genes was cloned from Pseudonocardia autotrophica IFO 12743 that can convert vitamin D$\_$3/ into its hydroxylated active forms. In order to isolate the P-450 gene cluster in this organism, we designed PCR primers on the basis of the regions of an oxygen binding site and a heme ligand pocket that are general characteristics of the P-450 hydroxylase. Sequencing analysis of the BamHI fragment revealed the presence of four complete and one incomplete ORFs, named PauA, PauB, PauC, and PauD, respectively. As a result of computer-based analyses, PauA and PauB have homology with enoyl-CoA hydratase from several organisms and the positive regulators belonging to the tetR family, respectively. PauC and PauD show similarity with SuaB/C proteins and ferredoxins, respectively, which are composed of P-450 monooxygenase systems for metabolizing two sulfonylurea herbicides in Streptomyces griseolus PauC shows the highest similarity with another CytP-450$\_$Sca2/ protein that is responsible for production of a specific HMG-CoA reductase inhibitor, pravastatin, in S. carbophilus. Cultures of Steptomyces lividans transformant, containing the P-450 gene cluster on the pWHM3 plasmid, was unable to convert vitamin D$\_$3/ to its hydroxylated forms.

• Reproductive and Developmental Biology
• /
• 제29권1호
• /
• pp.57-62
• /
• 2005

본 연구에서는 retrovirus를 이용한 유전자 전이에 있어서 대두되는 큰 문제점의 하나인 외래 유전자의 지속적인 발현으로 인한 개체의 생리적인 손상을 최소화하기 위하여 tetracycline계 물질의 공급 여부에 따라서 발현을 유도적으로 조절할 수 있는 one vector 형태의 Tet-On system을 구축하고자 하였다. 또한 WPRE 서열을 이 vector 상에 도입하여 유도적 조건에서 외래 유전자의 발현이 보다 강하게 일어날 수 있는 효율적인 retrovirus vector system을 확립하고자 하였다. 구축한 각각의 vector system에서 fluorometry와 western blotting을 이용하여 GFP 유전자의 발현 정도를 비교 측정한 결과, RevTRE-EGFP-WPRE-RSVp-rtTA2SM2 virus를 이용하여 유전자를 전이시킨 표적세포에서 GFP의 절대적인 발현량이 가장 큰 것으로 나타났고, 유전자 발현의 turn on/off에 의한 유도율은 RevTRE-EGFP-RSVp-rtTA2SM2-WPRE virus의 경우에서 8∼21배로 가장 높은 것으로 나타났다. 이상의 결과에서 외래 유전자의 발현을 효율적으로 조절할 수 있는 vector system은 WPRE가 rtTA2SM2 서열의 3에 위치한 형태로, 이 system을 이용하여 생산한 고감염가의 virus는 유전자 치료나 형질전환 동물의 생산에 있어서 요구되는 외래 유전자의 발현을 효율적으로 조절할 수 있는 수단이 될 것이다.

• Journal of Microbiology
• /
• 제44권2호
• /
• pp.192-199
• /
• 2006

Pseudomonas putida KL47 is a natural isolate that assimilates benzene, 1-alkylbenzene $(C_1-C_4)$, biphenyl, p-cumate, and p-cymene. The genetic background of strain KL47 underlying the broad range of growth substrates was examined. It was found that the cym and cmt operons are constitutively expressed due to a lack of the cymR gene, and the tod operon is still inducible by toluene and biphenyl. The entire array of gene clusters responsible for the catabolism of toluene and p-cymene/p-cumate has been cloned in a cosmid vector, pLAFR3, and were named pEK6 and pEK27, respectively. The two inserts overlap one another and the nucleotide sequence (42,505 bp) comprising the cym, cmt, and tod operons and its flanking genes in KL47 are almost identical (>99 %) to those of P. putida F1. In the cloned DNA fragment, two genes with unknown functions, labeled cymZ and cmtR, were newly identified and show high sequence homology to dienelactone hydrolase and CymR proteins, respectively. The cmtR gene was identified in the place of the cmtI gene of previous annotation. Western blot analysis showed that, in strains F1 and KL47, the todT gene is not expressed during growth on Luria Bertani medium. In minimal basal salt medium, expression of the todT gene is inducible by toluene, but not by biphenyl in strain F1; however, it is constantly expressed in strain KL47, indicating that high levels of expression of the todST genes with one amino acid substitution in TodS might provide strain KL47 with a means of adaptation of the tod catabolic operon to various aromatic hydrocarbons.

• Journal of Microbiology
• /
• 제45권5호
• /
• pp.379-387
• /
• 2007

A hundred and seventeen antibiotic-resistant Escherichia coli strains were isolated from public tap and spring waters which were polluted by fecal coliforms. There were no significant differences between two water sources as to the coliform pollution level (p> 0.05). All E. coli isolates were detected to be resistant to one or more antibiotics tested. Nearly 42% of the isolates showed multiresistant phenotype. Three (2.5%) of these isolates contained class 1 integron. Sequencing analysis of variable regions of the class 1 integrons showed two gene cassette arrays, dfr1-aadA1 and dhfrA17-aadA5. Resistance to ampicillin, tetracycline or trimethoprim-sulfamethoxazole was transferable according to the results of conjugation experiments. The rate of tetracycline resistance was 15%. tet(A)-mediated tetracycline resistance was widespread among tetracycline-resistant E. coli isolates. Genotyping by BOX-polymerase chain reaction (BOX-PCR) showed that some of the strains were epidemiologically related. This is the first report on the prevalence and characterization of class 1 integron-containing E. coli isolates of environmental origin in Turkey.

• 한국발생생물학회:학술대회논문집
• /
• 한국발생생물학회 2003년도 제3회 국제심포지움 및 학술대회
• /
• pp.107-107
• /
• 2003

The activation of protooncogenes or the inactivation of their gene products may be a specific and effective functional study for human neoplasia. To examine this possibility, we have used the tetracycline regulatory system to generate transgenic mice that conditionally express the HccR-2 protooncogene in vivo. The new human cervical cancer protooncogene (HccR-2) was detected from cervical cancer cell line. To elucidate its biological functions, we generated transgenic mice that expressed the HccR-2 gene. The sustained expression of the HccR-2 transgene culminated chronic neutrophilic leukemia (CNL). CNL is a rare chronic myeloproliferative disorder that presents as a sustained, mature neutrophilic leukocytosis with few or no circulating immature granulocytes, the absence of peripheral blood monocytosis, basophilia, or eosinophilia, and infiltration of neutrophils at the liver, spleen and kidney. Mice expressing the HccR-2 and tetracycline-transactivating protein (tTa) transgene were found to have altered myeloid development that was characterized by increased percentages of mature neutrophil and band form neutrophil in the peripheral blood, liver and spleen. Activation of the transgene causes CNL. In our model, expression of HccR-2 transgene mice was similar in many respects to the human CNL. This model will be valuable not only for investigating the biological properties of the HccR-2 and other protooncogenes in vivo but also for analyzing the mechanism involved in the progression of CNL.

• Biomolecules & Therapeutics
• /
• 제6권4호
• /
• pp.351-357
• /
• 1998

In order to understand the mechanism of the regulation of CYPIAI gene expression and ethoxy-resorufin deethylase (EROD) activity in ex vivo system, we have studied the action of TCDD and 3MC in theisolated perfused male rat liver. CYPIAI myNA level and EROD activity were measured in rat liver that wasisolated and perfused with va.ious chemicals such as 2,3,7,8-tet.achlorodibenzo-p-dioxin (TCDD), 3-methyl-cholanthrene (3MC), $17{\beta}$-est.adios ($E_2$), morin. TCDD or 3MC alone perfusion into male rat liver resulted in increase of CYPIAI mRNA level and the magnitude of stimulation was one and half times higher with TCDD treatment than 3MC treatment. However $E_2$ perfusion into male rat liver showed slight stimulation of CYPIAI mRNA level. When $10_{-8}$ M $E_2$ was perfused concomitantly with either $10_{-9}$ M TCDD or $10_{-9}$ M 3MC, stimulated CYPIAI mRNA by either TCDD or 3MC was inhibited. Morin was examined for its effects on CYPIAI mRNA level and result was similar to that was observed with estrogen except that morin alone did not change the level of CYPIAI mRNA. EROD activity was also stimulated with either TCDD or 3MC perfusion, and the magnitude of EROD stiumlation was similar to that of CYPIAI mRNA stimulation in response to TCDD or 3MC perfusion. This data is different from the data that we have obtained with female rat liver. Concomitant perfusion either $E_2$ or morin with TCDD or 3MC inhibited 3MC perFusion or TCDD perfusion stimulated EROD activity. These data confirm the hypothesis that TCDD and 3MC might act through the same mechanism of action on the regulation of CYPIAI gene expression in male rat liver.

• Reproductive and Developmental Biology
• /
• 제30권3호
• /
• pp.157-162
• /
• 2006

본 연구에서는 인간 부자상선 호르몬의 발현을 유도적으로 조절할 수 있는 retrocvirus vector system을 확립하고자 하였다. 이에 tetracycline계 물질로 발현을 유도적으로 조절할 수 있는 one vector 형태의 Tet-On system을 이용하였으며 WPRE 서열을 도입하여 유도적 조건에서 외래 유전자의 발현을 증가시켰다. 구축한 각각의 표적세포에서 RT-PCR과 ELISA를 이용하여 hPTH유전자의 발현 정도를 비교 측정한 결과, WPRE가 hPTH의 3' 위치에 도입된 $RevTRE-PTH-WPRE-CMVp-rtTA2^sM2$ virus를 이용하여 유전자를 전이시킨 경우에 hPTH의 발현량이 가장 높은 것으로 나타났고, 또한 유도율도 가장 큰 것으로 확인되었다. 이 system을 이용하여 생산한 고감염가의 virus는 인간의 부갑상선 호르몬을 생산하기 위한 동물세포주의 구축이나 형질전환 동물의 생산에 있어서 매우 효율적인 유전자 전이 수단이 될 것으로 사료된다.

• 한국미생물·생명공학회지
• /
• 제34권1호
• /
• pp.15-22
• /
• 2006

Ochrobactrum anthropi JW-2의 염색체 DNA로부터 paraquat 내성 유전자 pqrA를 포함하는 4,971 bp의 DNA 염기서열을 결정하였다. 염기서열 분석 결과 2개의 불완전한 ORF(orf1, orf5)와 4개의 완전한 ORF(orf2, pqrA, orf3, orf4)가 존재하는 것으로 나타났는데 orf1, pqrA, orf4, orf5는 direct strand에 orf2와 orf3은 reverse complementary strand 존재하였다. Orf1은 개시코돈이 결손된 불완전한 서열로서, response regulator receiver의 ATP binding region과 상동성을 나타내었다. Orf2는 tetR family에 속하는 transcription repressor와 높은 상동성을 나타내었고 H-T-H motif가 존재하는 것으로 나타났다. 따라서 orf2가 pqrA 유전자의 전사조절에 관여하는 repressor로 추정되어 pqrR2로 명명하였다. Orf3은 lysR type의 transcription activator와 높은 상동성을 나타내었고 N-terminal 부위에 H-T-H motif와 C-terminal 부위에 substrate binding domain이 존재하는 것으로 나타났다. 따라서 orf3은 pqrA의 전사조절에 관여하는 transcription activator로 추정되어 pqrR1로 명명하였다. Orf4는 amino acid ABC transporter의 periplasmic amino acid-binding protein과 상동성을 나타내었으며, orf5는 종결 코돈이 없는 불완전한 ORF로서 amino acid ABC transporter의 permease protein과 상동성을 나타내었다. 이와 같은 결과로 미루어 pqrA 유전자 주위에 존재하는 전사조절 유전자들이 paraquat 내성유전자인 pqrA의 발현조절을 통하여 paraquat에 대한 내성획득에 관여하는 것으로 판단되었다.