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Isolation and Characterization of a Theta-Type Cryptic Plasmid from Bifidobacterium longum FI10564

  • Moon, Gi-Seong;Wegmann, Udo;Gunning, A. Patrick;Gasson, Michael J.;Narbad, Arjan
    • Journal of Microbiology and Biotechnology
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    • v.19 no.4
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    • pp.403-408
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    • 2009
  • A number of bifidobacterial species of human origin were screened for the presence of cryptic plasmids. One strain, Bifidobacterium longum FI10564, harbored plasmids of approximately 2.2 kb, 3.6 kb, and 4.9 kb in size. The smallest plasmid, pFI2576(2,197 bp), was studied in detail and its complete nucleotide sequence was determined. Computer-assisted analysis of this novel plasmid(G+C content 62%) identified 9 putative open reading frames(orfs), 3 of which were shown to be probable genes. These putative genes are arranged in an operon-like structure, in which the overlapping orfs 1 and 2 encode putative Rep proteins and are highly homologous to the rep genes of the B. longum plasmid pMBI(1,847 bp). The mechanism of replication of pFI2576 was investigated using Southern blot analysis of whole cell lysates, with and without S1 nuclease treatment, and atomic force microscopy(AFM). The results indicate that pFI2576 is likely to use the theta mode of replication.

A Newly Isolated Bacteriophage, PBES 02, Infecting Cronobacter sakazakii

  • Lee, Hyung Ju;Kim, Wan Il;Kwon, Young Chan;Cha, Kyung Eun;Kim, Minjin;Myung, Heejoon
    • Journal of Microbiology and Biotechnology
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    • v.26 no.9
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    • pp.1629-1635
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    • 2016
  • A novel bacteriophage, PBES 02, infecting Cronobacter sakazakii was isolated and characterized. It has a spherical head of 90 nm in diameter and a tail of 130 nm in length, and belongs to Myoviridae as observed under a transmission electron microscope. The major virion protein appears to be 38 kilodaltons (kDa) in size. The latent period of PBES 02 is 30 min and the burst size is 250. Infectivity of the phage remained intact after exposure to temperatures ranging from 4℃ to 55℃ for 1 h. It was also stable after exposure to pHs ranging from 6 to 10 for 1 h. The phage effectively removed contaminating Cronobacter sakazakii from broth infant formula. PBES 02 has a double-stranded DNA genome of 149,732 bases. Its GC ratio is 50.7%. Sequence analysis revealed that PBES 02 has 299 open reading frames (ORFs) and 14 tRNA genes. Thirty-nine ORFs were annotated, including 24 related to replication and regulation functions, 10 related to structural proteins, and 5 related to DNA packaging. The genome of PBES 02 is closely related to that of two other C. sakazakii phages, CR3 and CR8. Comparison of DNA sequences of genes encoding the major capsid protein revealed a wide geographical distribution of related phages over Asia, Europe, and America.

Cloning and Characterization of a Gene Cluster for the Production of Polyketide Macrolide Dihydrochalcomycin in Streptomyces sp. KCTC 0041BP

  • Jaishy Bharat Prasad;Lim Si-Kyu;Yoo Ick-Dong;Yoo Jin-Cheol;Sohng Jae-Kyung;Nam Doo-Hyun
    • Journal of Microbiology and Biotechnology
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    • v.16 no.5
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    • pp.764-770
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    • 2006
  • Dihydrochalcomycin (GERI-155), produced by Streptomyces sp. KCTC-0041BP isolated from Korean soil, is a 16-membered macrolide antibiotic consisting of two deoxysugar moieties at C-5 and C-20 positions of a branched lactone ring. The cloning and sequencing of a gene cluster for dihydrochalcomycin biosynthesis revealed a 63-kb nucleotide region containing 25 open reading frames (ORFs). The products of all of these 25 ORFs playa role in dihydrochalcomycin biosynthesis and self-resistance against the compounds synthesized. At the core of this cluster lies a 39.6-kb polyketide synthase (PKS) region encoding eight modules in five giant multifunctional protein-coding genes (gerSI-SV). The genes responsible for the biosynthesis of deoxysugar moieties, D-chalcose and D-mycinose, and their modification and attachment were found on either side of this PKS region. The involvement of this gene cluster in dihydrochalcomycin biosynthesis was confirmed by disruption of the dehydratase (DH) domain in module 3 of the PKS gene and by metabolite analysis.

Characterization of the pcbE Gene Encoding 2-Hydroxypenta-2,4-Dienoate Hydratase in Pseudomonas sp. DJ-12

  • Lim, Jong-Chul;Lee, Jeongrai;Jang, Jeong-Duk;Lim, Jai-Yun;Min, Kyung-Rak;Kim, Chi-Kyung;Kim, Young-Soo
    • Archives of Pharmacal Research
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    • v.23 no.2
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    • pp.187-195
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    • 2000
  • Nucleotide sequence extending 2,3-dihydroxybiphenyl 1,2-dioxygenase gene (pcbC) and 2-hydroxy-6-oxo-6-phenylhexa-2,4-dienoate hydrolase gene (pcbD) of Pseudomonas sp. DJ-12 was previously analyzed and the two genes were present in the order of pcbD-pcbC preceded by a promoter from Pseudomonas sp. DJ-12. In this study, a 3.8-kb nucleotide sequence located downstream of the pcbC gene was analyzed to have three open reading frames (ORFs) that are designated as orf1, pcbE and orf2 genes. All of the ORFs were preceded by each ribosome-binding sequence of 5-GGAXA-3 (X=G or A). However, no promoter-like sequence and transcription terminator sequence were found in the analyzed region, downstream of pcbC gene. Therefore, the gene cluster appeared to be present in the order of pcbD-pcbC-orf1-pcbE-orf2 as an operon, which is unique organization characterized so far in biphenyl- and PCB-degrading bacteria. The orf1 gene was composed of 1,224 base pairs which can encode a polypeptide of molecular weight 44,950 containing 405 amino acid residues. A deduced amino acid sequence of the orf1 gene product exhibited 21-33% identity with those of indole dioxygenase and phenol hydroxylase components. The pcbE gene was composed of 783 base pairs encoding 2-hydroxypenta-2,4-dienoate hydratase involved in the 4-chlorobiphenyl catabolism. The orf2 gene was composed of 1,017 base pairs encoding a polypeptide of molecular weight 37,378 containing 338 amino acid residues. A deduced amino acid sequence of the orf2 gene product exhibited 31% identity with that of a nitrilotriacetate monooxygenase component.

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항생제 다제내성균 Staphylococcus aureus SA2로부터 분리한 테트라사이클린 내성 플라스미드 pKH6의 염기서열

  • Lee, Dae-Woon;Yoon, Sung-Joon;Kim, Woo-Koo;Shin, Chul-Kyo;Im, Sung-Hwan;Lee, Baek-Rak;Moon, Kyung-Ho
    • Microbiology and Biotechnology Letters
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    • v.24 no.4
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    • pp.423-426
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    • 1996
  • The complete nucleotide sequence of pKH6, a tetracycline-resistance (Tc$^{r}$) plasmid isolated from multi-drug resistant Staphylococcus aureus SA2, has been determined and compared with that of the staphylococcal Tc$^{r}$ plasmid pTl8l. The nucleotide sequences of the two plasmids are in agreement except for 7 nucleotides. All differences are caused by base pair substitutions. Among 6 substitutions, 3 occurred in coding regions. However, only two base substitutions in coding regions resulted in changes of amino acid sequences in two different ORFs of repC and Pre proteins.

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Characteristics of Cucumber mosaic virus isolated from Zea mays in Korea

  • Kim, Mi-Kyeong;Kwak, Hae-Ryun;Lee, Su-Heon;Kim, Jeong-Soo;Kim, Kook-Hyung;Cha, Byeong-Jin;Choi, Hong-Soo
    • The Plant Pathology Journal
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    • v.27 no.4
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    • pp.372-377
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    • 2011
  • A virus causing mottle and stunt symptom on Zea mays was observed around Ulleng-do, Korea and identified as Cucumber mosaic virus (CMV-ZM) based upon biological, serological, and molecular characteristics. In host range studies, the CMV-ZM isolate produced local lesions on Datura stramonium, Vigna unguiculata, Cucurbita moschata, Chenopodium amaranticolor, Ch. quinoa, whereas this isolate produced systemic mosaic on Nicotiana tabacum cv. 'Xanthi-nc', Capsicum annuum, Solanum lycopersicum, Solanum melongena, Cucurbita pepo, and Z. mays. In addition, chlorotic local rings on inoculated leaves along with severe mosaic, malformation, and fern leaf symptoms on upper systemic leaves were shown in N. glutinosa plants. Complete nucleotide sequences of each genomic RNA segment was determined and compared to those of the other CMV strains. Comparison of the nucleotide sequence of 1a open reading frame (ORF) revealed approximately 89.2-92.4% sequence identity with each CMV subgroup IA and IB strain, while showing only 78% sequence identity with CMV subgroup II. Nucleotide sequence analysis of RNA2 ORFs revealed 85.3-97.6% sequence identity with subgroup I. In ORFs of RNA3, levels of nucleotide sequence identities were higher than 92-99.2% with CMV subgroup I and lower than 82% with CMV isolates of subgroup II. These results suggest that CMV-ZM isolate is more closely related to subgroup I than subgroup II and therefore, CMV-ZM isolate might be classified into as CMV subgroup I based on biological and molecular analysis.

Genomic Approaches for Understanding the Characteristics of Salmonella enterica subsp. enterica Serovar Typhimurium ST1120, Isolated from Swine Feces in Korea

  • Kim, Seongok;Kim, Eunsuk;Park, Soyeon;Hahn, Tae-Wook;Yoon, Hyunjin
    • Journal of Microbiology and Biotechnology
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    • v.27 no.11
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    • pp.1983-1993
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    • 2017
  • Salmonella enterica subsp. enterica serovar Typhimurium, one of the most common foodborne pathogens, is transmitted mainly through contaminated food derived from infected animals. In this study, S. Typhimurium ST1120, an isolate from pig feces in Korea, was subjected to whole-genome analysis to understand its genomic features associated with virulence. The genome of ST1120 was found to have a circular chromosome of 4,855,001 bp (GC content 52.2%) and a plasmid of 6,863 bp (GC content 46.0%). This chromosome was predicted to have 4,558 open reading frames (ORFs), 17 pseudogenes, 22 rRNA genes, and 86 tRNA genes. Its plasmid was predicted to have three ORFs. Comparative genome analysis revealed that ST1120 was phylogenetically close to S. Typhimurium U288, a critical isolate in piggery farms and food chains in Europe. In silico functional analysis predicted that the ST1120 genome harbored multiple genes associated with virulence and stress resistance, including Salmonella pathogenicity islands (SPIs containing SPI-1 to SPI-5, SPI-13, and SPI-14), C63PI locus, ST104 prophage locus, and various antibiotic resistance genes. In accordance with these analysis results, ST1120 showed competence in invasion and survival abilities when it was added to host cells. It also exhibited robust resistance against antibiotics in comparison with other S. Typhimurium strains. This is the first report of the complete genome sequence of S. Typhimurium isolated from swine in Korea. Comparative genome analysis between ST1120 and other Salmonella strains would provide fruitful information toward understanding Salmonella host specificity and developing control measures against S. Typhimurium infection.

Complete genome and phylogenetic analysis of bovine papillomavirus type 15 in Southern Xinjiang dairy cow

  • Hu, Jianjun;Zhang, Wanqi;Chauhan, Surinder Singh;Shi, Changqing;Song, Yumeng;Zhao, Yubing;Wang, Zhehong;Cheng, Long;Zhang, Yingyu
    • Journal of Veterinary Science
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    • v.21 no.6
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    • pp.73.1-73.10
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    • 2020
  • Background: Bovine papilloma is a neoplastic disease caused by bovine papillomaviruses (BPVs), which were recently divided into 5 genera and at least 24 genotypes. Objectives: The complete genome sequence of BPV type 15 (BPV Aks-02), a novel putative BPV type from skin samples from infected cows in Southern Xinjiang China, was determined by collecting warty lesions, followed by DNA extraction and amplicon sequencing. Methods: DNA was analyzed initially by polymerase chain reaction (PCR) using the degenerate primers FAP59 and FAP64. The complete genome sequences of the BPV Aks-02 were amplified by PCR using the amplification primers and sequencing primers. Sequence analysis and phylogenetic analysis were performed using bio-informatic software. Results: The nucleotide sequence of the L1 open reading frame (ORF) of BPV Aks-02 was 75% identity to the L1 ORF of BPV-9 reference strain from GenBank. The complete genome consisted of 7,189 base pairs (G + C content of 42.50%) that encoded 5 early (E8, E7, E1, E2, and E4) and 2 late (L1 and L2) genes. The E7 protein contained a consensus CX2CX29CX2C zinc-binding domain and a LxCxE motif. Among the different members of this group, the percentages of the complete genome and ORFs (including 5 early and 2 late ORFs) sequence identity of BPV Aks-02 were closer to the genus Xipapillomavirus 1 of the Xipapillomavirus genus. Phylogenetic analysis and sequence similarities based on the L1 ORF of BPV Aks-02 revealed the same cluster. Conclusions: The results suggest that BPV type (BPV Aks-02) clustered with members of the Xipapillomavirus genus as BPV 15 and were closely related to Xipapillomavirus 1.

Interaction between host cell proteins and open reading frames of porcine circovirus type 2

  • Si-Won Park;In-Byung Park;Seok-Jin Kang;Joonbeom Bae;Taehoon Chun
    • Journal of Animal Science and Technology
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    • v.65 no.4
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    • pp.698-719
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    • 2023
  • Postweaning multisystemic wasting syndrome (PMWS) is caused by a systemic inflammation after porcine circovirus type 2 (PCV2) infection. It was one of the most economically important pathogens affecting pig production worldwide before PCV2 vaccine was first introduced in 2006. After the development of a vaccine against PCV2a type, pig farms gradually restored enormous economic losses from PMWS. However, vaccine against PCV2a type could not be fully effective against several different PCV2 genotypes (PCV2b - PCV2h). In addition, PCV2a vaccine itself could generate antigenic drift of PCV2 capsid. Therefore, PCV2 infection still threats pig industry worldwide. PCV2 infection was initially found in local tissues including reproductive, respiratory, and digestive tracks. However, PCV2 infection often leads to a systemic inflammation which can cause severe immunosuppression by depleting peripheral lymphocytes in secondary lymphoid tissues. Subsequently, a secondary infection with other microorganisms can cause PMWS. Eleven putative open reading frames (ORFs) have been predicted to encode PCV2 genome. Among them, gene products of six ORFs from ORF1 to ORF6 have been identified and characterized to estimate its functional role during PCV2 infection. Acquiring knowledge about the specific interaction between each PCV2 ORF protein and host protein might be a key to develop preventive or therapeutic tools to control PCV2 infection. In this article, we reviewed current understanding of how each ORF of PCV2 manipulates host cell signaling related to immune suppression caused by PCV2.

Nucleotide Sequences of Rep and CAT Proteins encoded by Chloramphenicol-Resistance Plasmid pKH7 (클로람페니콜 내성 플라스미드 pKH7의 Rep 단백질과 CAT 단백질의 염기서열 분석)

  • 윤성준;이대운;김우구;신철교;임성환;문경호
    • YAKHAK HOEJI
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    • v.39 no.6
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    • pp.676-680
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    • 1995
  • The nucleotide sequence of Xbal-Mbol fragment of pKH7, a chloramphenicol-resistant($Cm^{r}$) plasmid isolated from multidrug-resistant S. aureus SA2, has been determined. Xbal-Mbol fragment of pKH7 was found to contain two ORFs. One ORF encoded Rap and the other encoded CAT protein. The deduced amino acid sequences of Rep and CAT of pKH7 were compared to those of pUB112 and pC221. Comparisons revealed that there was one amino acid difference in CAT between pKH7 and pUB112. CAT of pKH7 exhibited 98.6% amino acid identity to that of pC221. In case of Rep proteins, a slightly lower homology of 96.4% and 86.7% in amino acid sequences was observed between pKH7 and pUB112 and between pKH7 and pC221, respectively.

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