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August 6, 2007
[Identification of a hrp-associated gene hpaB in the role of pathogenicity of Xanthomonas oryzae pv. oryzae]
Wei Sheng Wu Xue Bao. 2007 Jun 4;47(3):402-6
Authors: Zeng SY, Hu J, Huang GX, He CZ
Xanthomonas oryzae pv. oryzae (Xoo), a Gram-negative bacterium, is the causal agent of rice bacterial blight disease, which can cause severe yield loss of rice worldwide. To identify genes contributing to virulence and explore the possible mechanism of pathogenicity, transposon mutagenesis was used to isolate nonpathogenic mutants. By screening of a high-quality Tn5-like transposon (EZ: :TN) insertional mutant library of Xoo PXO99 against a host plant (rice cultivar IR24), one virulence-deficient mutant, XOG11, was identified. Genomic fragment flanking the insertion site of the mutant was amplified by thermal asymmetric interlaced polymerase chian reaction ( TAIL-PCR) and sequenced. The result of NCBI blast homologue searching of the fragment shows that the transposon was inserted into a hrp associated gene, hpaB. Xoo hpaB gene is one of the hrp gene cluster members that encode a type [I secretion system (TTSS) and locates at the downstream of hrpE. The product of hpaB in Xoo is a small (Molecular Weight, 17.6kDa), acidic (PI, 4.28) and Leucine-rich (14.4%) protein and shares high homology with corresponding proteins in other Xanthomonas. It suggests that HpaB may play as a TTSS chaperone. Mutant XOGl1 was confirmed both by PCR and Southern blotting: The PCR result by using primers upstream and downstream of hpaB respectively verified Tn5 insertion in hpaB and excluded the rare case of second transfer of the transposon associated with flanking sequence; Southern blot of digested genomic DNA with the probe of Km resistance gene aph proved that XOG11 was inserted by a single-copy transposon, indicating that the loss of pathogenicity in XOG11 was due to the Tn5 insertion in hpaB gene. Genetic complementation by cloning hpaB in the wide host range plasmid pHMI and transferring the recombinant plasmid into XOG11 restored its pathogenicity in IR24. These results suggest that the pathogenicity deficiency of XOG11 is due to the mutation of hpaB gene.
PMID: 17672295 [PubMed - in process]
[Establishment of the hrp-inducing systems for the expression of the hrp genes of Xanthomonas oryzae pv. oryzicola]
Wei Sheng Wu Xue Bao. 2007 Jun 4;47(3):396-401
Authors: Xiao YL, Li YR, Liu ZY, Xiang Y, Chen GY
The hrp genes of Xanthomonas oryzae pv. oryzicola (Xooc), which is the causal agent of bacterial leaf streak in rice, possesses the ability to elicit hypersensitive response on nonhost plants and the pathogenicity in host rice. In order to analyze the function of the hrp genes, we developed hrp-inducing systems using transcriptional hrp: :gfp fusions with the promoters of hrpX and hpa 1 of Xooc. The levels of GFP protein expression indicated that the hrp gene expression in Xooc was not efficiently induced in NB medium, but efficiently in XOM3 medium. Using the hrpG and hrpX mutants of Xooc as the controls, the results by RT-PCR demonstrated that in wild type strain the expression of the hpa1 gene was suppressed in NB medium, but was increased in XOM3 medium. When incubated in XOM3, the expression of the hpa1 gene was abolished in hrpX mutant, while the level of the hpa1 gene expression was lower in the hrpG mutant than that in wild-type strain. More importantly, it was found that the induction of the hrp gene expression was strongly increased in response to rice suspension cells and callus in this study. This suggests that the hrp-inducing systems, XOM3 or rice suspension cells or rice callus, for the induction of the hrp genes expression be useful for functionally analyzing the hrp genes, mining effectors secreted by the type III secretion apparatus and understanding pathogenicity determinats of Xooc.
PMID: 17672294 [PubMed - in process]
August 4, 2007
Crystallization and preliminary crystallographic studies of LipA, a secretory lipase/esterase from Xanthomonas oryzae pv. oryzae.
Acta Crystallograph Sect F Struct Biol Cryst Commun. 2007 Aug 1;63(Pt 8):708-10
Authors: Aparna G, Chatterjee A, Jha G, Sonti RV, Sankaranarayanan R
Xanthomonas oryzae pv. oryzae is the causal agent of bacterial leaf blight, a serious disease of rice. Several enzymes that are secreted through the type II secretion system of this bacterium play an important role in the plant-microbe interaction, being important for virulence and also being able to induce potent host defence responses. One of these enzymes is a secretory lipase/esterase, LipA, which shows a very weak homology to other bacterial lipases and gives a positive tributyrin plate assay. In this study, LipA was purified from the culture supernatant of an overexpressing clone of X. oryzae pv. oryzae and two types of crystals belonging to space group C2 but with two different unit-cell parameters were obtained using the hanging-drop vapour-diffusion method. Type I crystals diffract to a maximum resolution of 1.89 A and have unit-cell parameters a = 93.1, b = 62.3, c = 66.1 A, beta = 90.8 degrees . Type II crystals have unit-cell parameters a = 103.6, b = 54.6, c = 66.3 A, beta = 92.6 degrees and diffract to 1.86 A. Solvent-content analysis shows one monomer in the asymmetric unit in both the crystal forms.
PMID: 17671374 [PubMed - in process]
July 31, 2007
Functional analysis of Xa3/Xa26 family members in rice resistance to Xanthomonas oryzae pv. oryzae.
Theor Appl Genet. 2007 Jul 27;
Authors: Cao Y, Duan L, Li H, Sun X, Zhao Y, Xu C, Li X, Wang S
Plant disease resistant (R) genes are frequently clustered in the genome. The diversity of members in a complex R-gene family may provide variation in resistance specificity. Rice Xa3/Xa26, conferring resistance to Xanthomonas oryzae pv. oryzae (Xoo) encodes a leucine-rich repeat (LRR) receptor kinase-type protein and belongs to a multigene family, consisting of Xa3/Xa26, MRKa, MRKc and MRKd in rice cultivar Minghui 63. MRKa and MRKc are intact genes, while MRKd is a pseudogene. Complementary analyses showed that MRKa and MRKc could not mediate resistance to Xoo when regulated by their native promoters, but MRKa not MRKc conferred partial resistance to Xoo when regulated by a strong constitutive promoter. Plants carrying truncated XA3/XA26, which lacked the kinase domain, were compromised in their resistance to Xoo. However, the kinase domain of MRKa could partially restore the function of the truncated XA3/XA26 in resistance. MRKa and MRKc showed similar expression pattern as Xa3/Xa26, which expressed only in the vascular systems of different tissues. The expressional characteristic of MRKa and MRKc perfectly fits the function of genes conferring resistance to Xoo, a vascular pathogen. These results suggest that although MRKa and MRKc cannot mediate bacterial blight resistance nowadays, they may be once effective genes for Xoo resistance. Their expressional characteristic and sequence similarity to Xa3/Xa26 will provide templates for generating novel recognition specificity to face the evolution of Xoo. In addition, both LRR and kinase domains encoded by Xa3/Xa26 and MRKa are the functional determinants and MRKa-mediated resistance is dosage-dependent.
PMID: 17657469 [PubMed - as supplied by publisher]
Xanthan production by Xanthomonas albilineans infecting sugarcane stalks.
J Plant Physiol. 2007 Jul 21;
Authors: Blanch M, Legaz ME, Vicente C
Xanthomonas albilineans is the causal organism of leaf scald, a bacterial vascular disease of sugarcane. Xanthomonas may invade the parenchyma between the bundles and cause reddened pockets of gum, identified as a xanthan-like polysaccharide. Since xanthan contains glucuronic acid, the ability of Xanthomonas to produce an active UDP glucose dehydrogenase is often seen as a virulence factor. X. albilineans axenically cultured did not secrete xanthans to Willbrink liquid media, but the use of inoculated sugarcane tissues for producing and characterizing xanthans has been required. A hypothesis about the role of sugarcane polysaccharides to assure the production of bacterial xanthan is discussed.
PMID: 17646030 [PubMed - as supplied by publisher]
The crystal structure of XC1258 from Xanthomonas campestris: A putative procaryotic Nit protein with an arsenic adduct in the active site.
Proteins. 2007 Jul 19;
Authors: Chin KH, Tsai YD, Chan NL, Huang KF, Wang AH, Chou SH
PMID: 17640068 [PubMed - as supplied by publisher]
Controlled synthesis of the DSF cell-cell signal is required for biofilm formation and virulence in Xanthomonas campestris.
Environ Microbiol. 2007 Aug;9(8):2101-9
Authors: Torres PS, Malamud F, Rigano LA, Russo DM, Marano MR, Castagnaro AP, Zorreguieta A, Bouarab K, Dow JM, Vojnov AA
Virulence of the black rot pathogen Xanthomonas campestris pv. campestris (Xcc) is regulated by cell-cell signalling involving the diffusible signal factor DSF. Synthesis and perception of DSF require products of genes within the rpf cluster (for regulation of pathogenicity factors). RpfF directs DSF synthesis whereas RpfC and RpfG are involved in DSF perception. Here we have examined the role of the rpf/DSF system in biofilm formation in minimal medium using confocal laser-scanning microscopy of GFP-labelled bacteria. Wild-type Xcc formed microcolonies that developed into a structured biofilm. In contrast, an rpfF mutant (DSF-minus) and an rpfC mutant (DSF overproducer) formed only unstructured arrangements of bacteria. A gumB mutant, defective in xanthan biosynthesis, was also unable to develop the typical wild-type biofilm. Mixed cultures of gumB and rpfF mutants formed a typical biofilm in vitro. In contrast, in mixed cultures the rpfC mutant prevented the formation of the structured biofilm by the wild-type and did not restore wild-type biofilm phenotypes to gumB or rpfF mutants. These effects on structured biofilm formation were correlated with growth and disease development by Xcc strains in Nicotiana benthamiana leaves. These findings suggest that DSF signalling is finely balanced during both biofilm formation and virulence.
PMID: 17635553 [PubMed - in process]
Mutation of a key residue in the type II secretion system ATPase uncouples ATP hydrolysis from protein translocation.
Mol Microbiol. 2007 Jul;65(2):401-12
Authors: Shiue SJ, Chien IL, Chan NL, Leu WM, Hu NT
Membrane-associated ATPase constitutes an essential element common to all secretion machineries in Gram-negative bacteria. How ATP hydrolysis by these ATPases is coupled to secretion process remains unclear. Here we identified R286 as a key residue in the type II secretion system (T2SS) ATPase XpsE of Xanthomonas campestris that plays a pivotal role in coupling ATP hydrolysis to protein translocation. Mutation of R286 to alanine made XpsE hydrolyse ATP at a rate five times that of the wild-type XpsE. Yet the mutant XpsE(R286A) is non-functional in protein secretion via T2SS. Detailed analyses indicated that the mutant XpsE(R286A) lost the ability co-ordinating the N- and C-domain of XpsE. Without significantly influencing XpsE binding affinity with ATP or its oligomerization, R286A mutation however, caused XpsE lose the ability to associate with the cytoplasmic membrane via XpsL(N). As a consequence, ATP hydrolysis by XpsE was uncoupled from protein secretion. Because R286 is highly conserved among members of the secretion NTPase superfamily, we speculate that its equivalent in other homologues may also play a critical energy coupling role for T2SS, type IV pilus assembly and type IV secretion system.
PMID: 17630971 [PubMed - in process]
Substrate Specificity Regions of the NRPS Adenylation Domains Involved in Albicidin Pathotoxin Biosynthesis are Highly Conserved within the Species Xanthomonas albilineans.
Appl Environ Microbiol. 2007 Jul 13;
Authors: Renier A, Vivien E, Cociancich S, Letourmy P, Perrier X, Rott PC, Royer M
Albicidin is a pathotoxin produced by Xanthomonas albilineans, a xylem-invading pathogen that causes leaf scald disease of sugarcane. Albicidin is synthesized by a non-ribosomal pathway via modular PKS (polyketide synthase) and NRPS (non-ribosomal peptide synthetase) megasynthases, and NRPS adenylation (NRPS-A) domains are responsible for the recognition and activation of specific amino-acid substrates. 0.5 kb DNA fragments encoding the region responsible for substrate specificity of six albicidin NRPS-A domains were amplified and sequenced from 16 strains of X. albilineans representing the known diversity of this pathogen. Polymorphism analysis of these DNA fragments at different levels (DNA, protein, NRPS signature) showed that these pathogenicity loci were highly conserved. Conservation of these loci is most likely under purifying selection, as revealed by a comparison with the variability of nucleotide and amino-acid sequences of two housekeeping genes (atpD and efp) of X. albilineans. Nevertheless, the 16 strains of X. albilineans were differentiated into several groups by a phylogenetic analysis of the nucleotide sequences of the NRPS-A domains. One of these groups was representative of the genetic diversity previously found within the pathogen by RFLP (Random Fragment Length Polymorphism) and AFLP (Amplified Fragment Length Polymorphism). This group, which differed by three single synonymous nucleotide mutations, contained only four strains of X. albilineans that were all involved in outbreaks of sugarcane leaf scald. The amount of albicidin produced in vitro in agar and liquid media varied according to the 16 strains of X. albilineans. However, no relationship was found between the amount of albicidin produced in vitro, the pathotypes and the genetic diversity of the pathogen. The NRPS loci contributing to the synthesis of the primary structure of albicidin apparently are not involved in the pathogenicity differences observed between strains of X. albilineans.
PMID: 17630307 [PubMed - as supplied by publisher]
Impact of the extrusion process on xanthan gum behaviour.
Carbohydr Res. 2007 Jul 23;342(10):1333-42
Authors: Sereno NM, Hill SE, Mitchell JR
Processing xanthan gum by extrusion and subsequent drying produces a biopolymer showing particulate, rather than molecular behaviour in aqueous solution. This form of xanthan disperses very readily to give a viscosity that is strongly dependent on salt concentration. On heating above the temperature of the order-disorder transition as determined by calorimetry, there is a viscosity transition that is indicative of the irreversible loss of the particulate structure. It is suggested that the extrusion process melts and aligns xanthan macromolecules. On cooling reordering will occur but in the highly concentrated environment in the extruder ( approximately 45% water w/w), inter-molecular association between neighbouring macromolecules cannot proceed to completion due to kinetic trapping. As a consequence a network structure is created maintained by associations involving ordered regions. A xanthan solution can be prepared from this particulate material by dispersing and subsequent heating far more readily than can be achieved with non-processed xanthan.
PMID: 17466288 [PubMed - indexed for MEDLINE]
Isolation and molecular characterization of a Spotted leaf 18 mutant by modified activation-tagging in rice.
Plant Mol Biol. 2007 Apr;63(6):847-60
Authors: Mori M, Tomita C, Sugimoto K, Hasegawa M, Hayashi N, Dubouzet JG, Ochiai H, Sekimoto H, Hirochika H, Kikuchi S
A lesion mimic mutant that we designated Spotted leaf 18 (Spl18) was isolated from 13,000 activation-tagging lines of rice produced by our modified activation-tagging vector and further characterized. Spl18 was dominant and its phenotype was linked to the T-DNA insertion. An ORF was located about 500 bp downstream of the inserted T-DNA, and the deduced protein, designated OsAT1, showed sequence similarity to an acyltransferase whose expression is induced by hypersensitive reaction in tobacco. The transcriptional level of OsAT1 was very low in the WT leaf blade but high in Spl18 leaf blade. In wild-type rice, OsAT1 was transcribed mainly in the young panicle, in the panicle just after heading, and in the leaf sheath. In addition, transcription of the genes for PR protein was upregulated in Spl18, accumulation of phytoalexins (both momilactone A and sakuranetin) was increased, and resistance to blast disease was improved. We then combined OsAT1 genomic DNA downstream of the modified 35S promoter and re-transformed it into rice. Lesion mimic and blast resistance phenotypes were detected in the transgenic lines produced, clearly indicating that overexpression of OsAT1 caused the Spl18 phenotypes. In addition, plants overexpressing OsAT1 showed resistance to bacterial blight.
PMID: 17273822 [PubMed - indexed for MEDLINE]
July 15, 2007
Biodegradability of chlorinated anilines in waters.
Biomed Environ Sci. 2007 Apr;20(2):141-5
Authors: Wang C, Lu GH, Zhou YJ
OBJECTIVE: To identify the bacteria tolerating chlorinated anilines and to study the biodegradability of o-chloroaniline and its coexistent compounds. METHODS: Microbial community of complex bacteria was identified by plate culture observation techniques and Gram stain method. Bacterial growth inhibition test was used to determine t he tolerance of complex bacteria to toxicant. Biodegradability of chlorinated anilines was determined using domesticated complex bacteria as an inoculum by shaking-flask test. RESULTS: The complex bacteria were identified, consisting of Xanthomonas, Bacillus alcaligenes, Acinetobacter Pseudomonas, and Actinomycetaceae nocardia. The obtained complex bacteria were more tolerant to o-chloroaniline than mixture bacteria in natural river waters. The effects of exposure concentration and inoculum size on the biodegradability of o-chloroaniline were analyzed, and the biodegradation characteristics of single o-chloroaniline and 2, 4-dichloroaniline were compared with the coexistent compounds. CONCLUSION: The biodegradation rates can be improved by decreasing concentration of compounds and increasing inoculum size of complex bacteria. When o-chloroaniline coexists with aniline, the latter is biodegraded prior to the former, and as a consequence the metabolic efficiency of o-chloroaniline is improved with the increase of aniline concentration. Meanwhile, when o-chloroaniline coexists with 2,4-dichloroaniline, the metabolic efficiency of 2,4-dichloroaniline is markedly improved.
PMID: 17624189 [PubMed - in process]
July 13, 2007
Structure of Xanthomonas axonopodis pv. citri YaeQ reveals a new compact protein fold built around a variation of the PD-(D/E)XK nuclease motif.
Proteins. 2007 Jul 10;
Authors: Guzzo CR, Nagem RA, Barbosa JA, Farah CS
The YaeQ family of proteins are found in many Gram-negative and a few Gram-positive bacteria. We have determined the first structure of a member of the YaeQ family by X-ray crystallography. Comparisons with other structures indicate that YaeQ represents a new compact protein fold built around a variation of the PD-(D/E)XK nuclease motif found in type II endonucleases and enzymes involved in DNA replication, repair, and recombination. We show that catalytically important residues in the PD-(D/E)XK nuclease superfamily are spatially conserved in YaeQ and other highly conserved YaeQ residues may be poised to interact with nucleic acid structures. Proteins 2007. (c) 2007 Wiley-Liss, Inc.
PMID: 17623842 [PubMed - as supplied by publisher]
Investigation of the chemical structure and biological activity of oligosaccharides isolated from rough-type Xanthomonas campestris pv. campestris B100 lipopolysaccharide.
J Endotoxin Res. 2007;13(2):101-8
Authors: Kaczyński Z, Braun S, Lindner B, Niehaus K, Holst O
The rough-type lipopolysaccharide (LPS) of the phytopathogenic bacterium Xanthomonas campestris pv. campestris B 100 was isolated utilizing the hot phenol-water method and successively de-acylated by treatment with hydrazine and hot potassium hydroxide. Four compounds were separated by preparative high-performance anion-exchange chromatography and studied by sugar analysis and by 1D and 2D homonuclear and heteronuclear (1)H-, (13)C- and (31)P-NMR spectroscopy as well as ESI FT-MS. The two main products were a heptasaccharide and a pentasaccharide of the structures alpha-D-Manp-(1-->3)-alpha-D-Man p-(1-->4)-beta-D-Glcp-(1-->4)-alpha-D-Manp-3P -(1-->5)-alpha-Kdo-(2-->6)-beta-D-GlcpN-4P-(1-->6)-alpha-D-Glc pN-1P (1) and beta-D-Glcp-(1-->4)-alpha-D-Man p-3P-(1-->5)-alpha-Kdo-(2-->6)-beta-D-GlcpN-4 P-(1-->6)-alpha-D-GlcpN-1P (2), respectively. The products in smaller amounts were a heptasaccharide and pentasaccharide possessing the above structures plus a phosphate group at C-4 of the Kdo residue (compounds 3 and 4). Both, heptasaccharide 1 and pentasaccharide 2 were able to induce an oxidative burst in cell cultures of the non-host plant tobacco.
PMID: 17621551 [PubMed - in process]
July 11, 2007
Three New Loci of Insertion Element IS1112 in Chinese Strains of Xanthomonas oryzae pv. oryzae.
J Microbiol. 2007 Jun;45(3):219-26
Authors: Xie J, Wang X, Li F, Peng Y, Zhou G
Insertion sequence IS1112 is a repetitive element with a relatively high number of copies in Xanthomonas oryzae pv. oryzae (Xoo), the causal agent of bacterial blight of rice (Oryza sativa L.). Three new loci of IS1112 were identified in seven Chinese strains of Xoo using a single oligonucleotide primer J3; 5'-GCTCAGGTCAGGTGGCCTGG-3' by insertion-sequence-based polymerase chain reaction (IS-PCR). Among the three new loci of IS1112, two were located in the open-reading frame region of genes fhuA and cirA, which encode TonB-dependent receptors, and the third in ISXo2, another type of insertion sequence in Xoo genome. Three variants of IS1112 were identified in those three loci based on their sequence similarities: two were identical to IS1112a and IS1112b, reported in strain PXO86 from the Philippines, while the third was a new member of IS1112, defined as IS1112d. Inserting IS1112 in gene fhuA caused three bases, GGT, to be duplicated at the target site, but inserting it in gene cirA did not cause any duplication in the target site. The diversity of IS1112 sequence and insertion loci in Xoo genome and their potential effects are discussed.
PMID: 17618227 [PubMed - in process]
July 10, 2007
Science Daily (press release)New Dry Edible Bean Resists Bacterial Disease
Science Daily (press release) - Jul 10, 2007
(Credit: Photo by Stephen Ausmus) Caused by the pathogen Xanthomonas campestris pv. phaseoli, bacterial blight is an endemic disease affecting bean crops ...
Science Daily (press release)New Dry Edible Bean Resists Bacterial Disease
Science Daily (press release) - Jul 10, 2007
(Credit: Photo by Stephen Ausmus) Caused by the pathogen Xanthomonas campestris pv. phaseoli, bacterial blight is an endemic disease affecting bean crops ...
Science Daily (press release)New Dry Edible Bean Resists Bacterial Disease
Science Daily (press release) - Jul 10, 2007
(Credit: Photo by Stephen Ausmus) Caused by the pathogen Xanthomonas campestris pv. phaseoli, bacterial blight is an endemic disease affecting bean crops ...
Science Daily (press release)New Dry Edible Bean Resists Bacterial Disease
Science Daily (press release) - Jul 10, 2007
(Credit: Photo by Stephen Ausmus) Caused by the pathogen Xanthomonas campestris pv. phaseoli, bacterial blight is an endemic disease affecting bean crops ...
Science Daily (press release)New Dry Edible Bean Resists Bacterial Disease
Science Daily (press release) - Jul 10, 2007
(Credit: Photo by Stephen Ausmus) Caused by the pathogen Xanthomonas campestris pv. phaseoli, bacterial blight is an endemic disease affecting bean crops ...