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Specific Detection of Xanthomonas oryzae pv. oryzicola in Infected Rice Plant by Use of PCR Assay Targeting a Membrane Fusion Protein Gene

  • Kang, Man-Jung;Shim, Jae-Kyung;Cho, Min-Seok;Seol, Young-Joo;Hahn, Jang-Ho;Hwang, Duk-Ju;Park, Dong-Suk
    • Journal of Microbiology and Biotechnology
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    • v.18 no.9
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    • pp.1492-1495
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    • 2008
  • Successful control of Xanthomonas oryzae pv. oryzicola, the causal agent of bacterial leaf streak, requires a specific and reliable diagnostic tool. A pathovar-specific PCR assay was developed for the rapid and accurate detection ofthe plant pathogenic bacterium Xanthomonas oryzae pv. oryzicola in diseased plant. Based on differences in a membrane fusion protein gene of Xanthomonas oryzae pv. oryzicola and other microorganisms, which was generated from NCBI (http://www.ncbi.nlm.nih.gov/) and CMR (http://cmr.tigr.org/) BLAST searches, one pair of pathovar-specific primers, XOCMF/XOCMR, was synthesized. Primers XOCMF and XOCMR from a membrane fusion protein gene were used to amplity a 488-bp DNA fragment. The PCR product was only produced from 4 isolates of Xanthomonas oryzae pv. oryzicola among 37 isolates of other pathovars and species of Xanthomonas, Pectobacterium, Pseudomonas, Burkholderia, Escherichia coli, and Fusarium oxysporum f.sp. dianthi. The results suggested that the assay detected the pathogen more rapidly and accurately than standard isolation methods.

Current Status on Molecular Genetic Study and Comparative Genomic Analysis of Virulence Related Genes in Xanthomonas oryzae pv. oryzae (벼 흰잎마름병균(Xanthomonas oryzae pv. oryzae)의 병원성 유전자의 분자유전학적 연구현황 및 비교유전체 분석)

  • Kang, Hee-Wan;Park, Young-Jin;Lee, Byeong-Moo
    • Korean Journal of Microbiology
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    • v.44 no.1
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    • pp.1-9
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    • 2008
  • Xanthomonas oryzae pv. oryzae (Xoo) is the causal agent of bacterial blight on rice. In this paper, current status on molecular genetic study of major virulence genes, hypersensitive response and pathogenicity (hrp), productions of extracellular polysaccharide (EPS), extracellular enzymes and lipopolysaccharides (LPS), avr genes were reviewed. The IS elements with 611 copies including 133 ORF IS were inserted in various regions of the Xoo genome and in expecially regions franking virulence genes. Whole genome sequence of X. oryzae pv. oryzae KACC10331 were used for defining genetic organization of the virulence genes. Futhermore, the virulence genes in Xoo genome were compared to those of other Xanthomonas species in Blast GenBank data base.

PCR-Based Sensitive Detection and Identification of Xanthomonas oryzae pv. oryzae (중합효소연쇄 반응에 의한 벼 흰잎마름병균의 특이적 검출)

  • Lee, Byoung-Moo;Park, Young-Jin;Park, Dong-Suk;Kim, Jeong-Gu;Kang, Hee-Wan;Noh, Tae-Hwan;Lee, Gil-Bok;Ahn, Joung-Kuk
    • Microbiology and Biotechnology Letters
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    • v.32 no.3
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    • pp.256-264
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    • 2004
  • A new primer set was developed for the detection and identification of Xanthomonas oryzae pv. oryzae, the bacterial leaf blight (BLB) pathogen in rice plant. The nucleotide sequence of hpaA gene was determined from X. o. pv. oryzae str. KACC10331, and the sequence information was used to design primers for the application of the polymerase chain reaction (PCR). The nucleotide sequence of hpaA from X. o. pv. oryzae str. KACC 10331 was aligned with those of X. campestris pv. vesicatoria, X. campestris pv. campestris, X. axonopodis pv. citri, and X. axonopodis pv. glycines. Based on these results, a primer set(XOF and XOR) was designed for the specific detection of hpaA in X. o. pv. oryzae. The length of PCR products amplified using the primer set was 534-bp. The PCR product was detected from only X. o. pv. oryzae among other Xanthomonas strains and reference bacteria. This product was used to confirm the conservation of hpaA among Xanthomonas strains by Southern-blotting. Furthermore, PCR amplification with XOF and XOR was used to detect the pathogen in an artificially infected leaf. The sensitivity of PCR detection in the pure culture suspension was also determined. This PCR-based detection methods will be a useful method for the detection and identification of X. o. pv. oryzae as well as disease forecasting.

A Semiselective Medium for the Isolation of Xanthomonas campestris pv. oryzae from Rice Seed (벼 종자에서 Xanthomonas campestris pv. oryzae의 분리를 위한 선택배지)

  • 김형무;송완엽;소인영;이두구
    • Korean Journal Plant Pathology
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    • v.10 no.1
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    • pp.13-17
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    • 1994
  • A semiselective agar medium (XCO) was developed for the isolation of bacterial blight pathogen, Xanthomonas campestris pv. oryzae, from rice seed. The medium contained yeast extract 1.0 g, peptone 2.0 g, sucrose 5.0 g, sodium glutamate 1.0 g, FeSO4.7H2O 0.05 g, Fe.EDTA 1 mg, cephalexin 20 mg, Evan blue (0.1%) 1.5 ml, bromcresol purple (0.1%) 2.5ml, cycloheximide 100 mg and agar 15.0 g per liter. Colonies of X. c. pv. oryzae were 4~5 mm in diameter, smooth, round, blue (darker center) and convex after 6 days incubation at 28$^{\circ}C$. The recovery of 6 isolates of X. c. pv. oryzae on the XOC medium ranged from 81% to 120% (mean 98.2%) in comparison to Wakimoto's medium. The number of saprophytic bacteria from 10 rice seed lots on XCO medium was reduced to 70.4% of that on Wakimoto's medium. The recovery of X. c. pv. oryzae added to rice seed on XOC medium ranged from 67% to 87% (mean 75.6%) of that on Wakimoto's medium.

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Recombinant Expression and Purification of Functional XorII, a Restriction Endonuclease from Xanthomonas oryzae pv. oryzae

  • Hwang, Dong-Kyu;Cho, Jae-Yong;Chae, Young-Kee
    • Journal of Microbiology
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    • v.45 no.2
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    • pp.175-178
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    • 2007
  • An endonuclease from Xanthomonas oryzae pathovar oryzae KACC 10331, XorII, was recombinantly produced in Escherichia coli using a T7 system. XorII was purified using a combination of ion exchange and immobilized metal affinity chromatography (IMAC). An optimized washing protocol was carried out on an IMAC in order to obtain a high purity product. The final amount of purified XorII was approximately 2.5 mg/L of LB medium. The purified recombinant XorII was functional and showed the same cleavage pattern as PvuI. The enzyme activity tested the highest at $25^{\circ}C$ in 50 mM NaCl, 10 mM Tris-HCl, 10 mM $MgCl_{2}$, and 1 mM dithiothreitol at a pH of 7.9.

Biosynthesis of Silver Nanoparticles by Phytopathogen Xanthomonas oryzae pv. oryzae Strain BXO8

  • Narayanan, Kannan Badri;Sakthivel, Natarajan
    • Journal of Microbiology and Biotechnology
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    • v.23 no.9
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    • pp.1287-1292
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    • 2013
  • Extracellular biogenic synthesis of silver nanoparticles with various shapes using the rice bacterial blight bacterium Xanthomonas oryzae pv. oryzae BXO8 is reported. The synthesized silver nanoparticles were characterized by UV-Vis spectroscopy, powder X-ray diffractometry (XRD), scanning electron microscopy, energy dispersive X-ray spectrometry, and high-resolution transmission electron microscopy (HR-TEM). Based on the evidence of HR-TEM, the synthesized particles were found to be spherical, with anisotropic structures such as triangles and rods, with an average size of 14.86 nm. The crystalline nature of silver nanoparticles was evident from the bright circular spots in the SAED pattern, clear lattice fringes in the high-resolution TEM images, and peaks in the XRD pattern. The FTIR spectrum showed that biomolecules containing amide and carboxylate groups are involved in the reduction and stabilization of the silver nanoparticles. Using such a biological method for the synthesis of silver nanoparticles is a simple, viable, cost-effective, and environmentally friendly process, which can be used in antimicrobial therapy.

Comparative AFLP Profiles among Strains of Korean Races of Xanthomonas oryzae pv. oryzae.

  • Kang, Mi-Hyung;Lee, Du-Ku;Noh, Tae-Hwan;Shim, Hyeong-Kwon;Na, Seung-Yong;Kim, Jae-Duk
    • Plant Resources
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    • v.7 no.1
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    • pp.65-68
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    • 2004
  • We used an amplified fragment length polymorphism (AFLP) analysis, a novel PCR-based technique, to differentiate Xanthomonas oryzae pv. oryzae (Xoo) of Korean races. The 6 strains of Xoo K1, K2, K3 races were tested with 81 AFLP primer combinations to identify the best selective primers. The primer combinations were selected according to their reproducibility, number of polymorphic bands and polymorphism detected among Xoo strains. 18 strains of Xoo K1, K2 and K3 races were analyzed with the selected combinations of primer set. Some primer combinations (Eco R I +1 / Mse I+1) could differentiate Xoo of Korean races that were not distinguished by other fingerprinting analysis. Thus AFLP fingerprinting permitted very fine discrimination among different races.

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Nitrogen Sources Inhibit Biofilm Formation by Xanthomonas oryzae pv. oryzae

  • Ham, Youngseok;Kim, Tae-Jong
    • Journal of Microbiology and Biotechnology
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    • v.28 no.12
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    • pp.2071-2078
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    • 2018
  • Xanthomonas oryzae pv. oryzae (Xoo) causes bacterial blight, which results in severe economic damage to rice farms. Xoo produces biofilms for pathogenesis and survival both inside and outside the host. Biofilms, which are important virulence factors, play a key role in causing the symptoms of Xoo infection. In the present study, we investigated the nutritional conditions for biofilm formation by Xoo. Although Xoo biofilm formation may be initiated by interactions with the host, Xoo biofilm cannot mature without the support of favorable nutritional conditions. Nitrogen sources inhibited Xoo biofilm formation by overwhelming the positive effect that cell growth has on it. However, limited nutrients with low amino acid concentration supported biofilm formation by Xoo in the xylem sap rather than in the phloem sap of rice.

Genetic Diversity of avrBs-like Genes in Three Different Xanthomonas Species Isolated in Korea

  • Oh, Chang-Sik;Lee, Seung-Don;Heu, Sung-Gi
    • The Plant Pathology Journal
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    • v.27 no.1
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    • pp.26-32
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    • 2011
  • Plant-pathogenic bacteria including Xanthomonas spp. carry genetic diversity in composition of avirulence genes for interaction with their host plants. Previously, we reported genetic diversity of avirulence genes in X. axonopodis pv. glycines. In this study, we determined genetic diversity of five avirulence genes, avrBs1, avrBs2, avrBs3, avrBs4, and avrRxv, in three other Xanthomonas species isolated in Korea by genomic southern hybridization. Although Korean races of X. campestris pv. vesicatoria that were isolated from year 1995 to 2002 had the same avirulence gene patterns as those that already reported, there was race shift from race 3 to race 1 by acquisition of avrBs3 genes. X. campestris pv. campestris isolated from Chinese cabbage, but not from cabbage or radish, carried two avrBs3 genes, and one of them affected HR-eliciting ability of this bacterium in broccoli. X. oryzae pv. oryzae carried eight to thirteen avrBs3 gene homologs, and this bacterium showed dynamic changes of resistance patterns in rice probably by losing or obtaining avrBs3 genes. These results indicate that avrBs3 gene is more diverse in Xanthomonas spp. than other four avirulence genes and also host ranges of these bacteria can be easily changed by loss or acquisition of avrBs3 genes.

Multiplication and Movement of Xanthomonas oryzae pv. oryzae in Rice Leaves with Resistance Genes Derived from Different Origins (기원이 다른 저항성 유전자를 갖는 근동질 계통에서 Xanthomonas oryzae pv. oryzae의 증식과 이동)

  • Kang, Sun-Joo;Lee, Sung-Eun;Kim, Min-Jeong;Han, Jin-Soo;Choi, Jae-Eul
    • KOREAN JOURNAL OF CROP SCIENCE
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    • v.55 no.4
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    • pp.306-311
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    • 2010
  • The multiplication and movement of Xanthomonas oryzae pv. oryzae in rice leaves of seven near-isogenic lines(NILs) derived from different genetic sources and from the susceptible cultivar Toyonishiki were examined. The bacterium populations increased rapidly in susceptible cultivar leaves of the inoculation sites but increased gradually in NIL leaves. X oryzae pv. oryzae were detected at 20cm above the leaves of the inoculated sites in IRBB 103 and Toyonishiki but were not detected in the other NILs at 25 days after inoculation. These results support that resistant genes restrict bacterial movement not multiplication.