[Show abstract][Hide abstract] ABSTRACT: ABSTRACT Based on 16S-23S internal transcribed spacer ribosomal DNA sequence data, two padlock probes (PLPs), P-Xoo and P-Xoc, were designed and tested to detect Xanthomonas oryzae pv. oryzae and X. oryzae pv. oryzicola, respectively. These PLPs were combined with dot-blot hybridization to detect X. oryzae pv. oryzae and X. oryzae pv. oryzicola individually in rice seed. Using this technique, a detection sensitivity of 1 pg of X. oryzae pv. oryzae genomic DNA was observed. The technique also facilitated the detection of X. oryzae pv. oryzae in rice seedlots with 2% artificially infested seed. With regards to X. oryzae pv. oryzicola a detection threshold of 1 pg genomic DNA was observed and the pathogen was successful detected in rice seedlots with 0.2% artificially infested seed. The PLP assays detected X. oryzae pv. oryzae and X. oryzae pv. oryzicola in 39.3% (13 of 33) and 21.3% (10 of 47) of naturally infested commercial rice seedlots, respectively. In contrast, conventional polymerase chain reaction using OSF1/OSR1 and XoocF/XoocR primers sets detected X. oryzae pv. oryzae and X. oryzae pv. oryzicola in 9.1% (3 of 33) and 8.5% (4 of 47) of the same rice seedlots, respectively. We also detected both pathogens simultaneously in two seedlots, which successfully proved that PLPs (P-Xoo and P-Xoc) combined with reverse dotblot hybridization can be used to simultaneously detect multiple pathogens in naturally infested commercial rice seedlots. This approach has the potential to be an important tool for detecting multiple pathogens in seed and thereby preventing the spread of important pathogens.
[Show abstract][Hide abstract] ABSTRACT: The type VI protein secretion system (T6SS) is essential for virulence of several Gram-negative bacteria. In this study, we identified a T6SS gene cluster in Acidovorax citrulli, a plant pathogenic bacterium that causes bacterial fruit blotch (BFB) of cucurbits. One T6SS cluster, approximately 25 kb in length, and comprised of 17 genes was found in the A. citrulli AAC00-1 genome. Seventeen A. citrulli mutants were generated, each with a deletion of a single T6SS core gene. There were significant differences in BFB seed-to-seedling transmission between wild type A. citrulli strain, xjl12, and ΔvasD, ΔimpK, ΔimpJ or ΔimpF mutants (71.71%, 9.83%, 8.41%, 7.15%, 5.99% BFB disease index, respectively). In addition, we observed that these four mutants were reduced in melon seed colonization and biofilm formation; however, they were not affected in virulence when infiltrated into melon seedling tissues. There were no significant differences in BFB seed-to-seedling transmission, melon tissue colonization and biofilm formation between xjl12 and the other 13 T6SS mutants. Overall, our results indicate that the T6SS plays a role in seed-to-seedling transmission of BFB on melon.
[Show abstract][Hide abstract] ABSTRACT: Polyketides and nonribosomal peptides represent two large families of natural products (NPs) with diverse structures and important functions. They are synthesized by polyketide synthase (PKS) and nonribosomal peptide synthetase (NRPS), respectively. Lysobacter enzymogenes is emerging as a novel biocontrol agent against pathogens of crop plants and a new source of bioactive NPs, such as antibacterial antibiotic WAP-8294A2 and antifungal antibiotic HSAF. Genome survey of strain OH11, a Chinese L. enzymogenes isolate, detected four novel PKS, NRPS or hybrid gene clusters, designed as cluster A to D. We further individually mutated five genes (PKS or NRPS) located in these four gene clusters, and showed that a PKS gene in cluster A and an NRPS gene in cluster D were involved in the antibacterial activity via a WAP-8294A2 dependent way. The data also showed that none of the five genes was associated with antifungal activity and the regulation of HSAF biosynthesis. Our results reveal the unusual regulatory role of these PKS and NRPS genes that were discovered from genome mining in L. enzymogenes.This article is protected by copyright. All rights reserved.
[Show abstract][Hide abstract] ABSTRACT: Lysobacter enzymogenes is a ubiquitous environmental bacterium emerging as a potentially novel biological control agent and a new source of bioactive secondary metabolites, such as the heat-stable antifungal factor (HSAF) and photo-protective polyene pigments. Thus far, the regulatory mechanism(s) for biosynthesis of these bioactive secondary metabolites remains largely unknown in L. enzymogenes. In the present study, DSF (Diffusible Signal Factor) and DF (Diffusible Factor)-mediated cell-cell signaling systems were identified for the first time from L. enzymogenes. The results show that both Rpf/DSF and DF signaling systems played critical roles in modulating HSAF biosynthesis in L. enzymogenes. Rpf/DSF and DF signaling played negative and positive effects in polyene pigment production, respectively, with DF playing a more important role in regulating this phenotype. Interestingly, only Rpf/DSF, but not DF signaling system, regulated colony morphology of L. enzymgenes. Both Rpf/DSF and DF signaling systems were involved in the modulation of gene expressions with diverse functions in L. enzymogenes, and their own regulons exhibited only a few loci being regulated by both systems. These findings unveil for the first time new roles of Rpf/DSF and DF signaling systems in secondary metabolite biosynthesis of L. enzymogenes.
[Show abstract][Hide abstract] ABSTRACT: A method was developed using a padlock probe (PLP) and dot-blot
hybridization for detecting Acidovorax citrulli in cucurbit seed. The
PLP was designed based on the 16S-23S internal transcribed spacer
ribosomal DNA sequence from A. citrulli. The detection threshold for
the PLP assay was 100 fg of genomic DNA, and A. citrulli was
detected in 100% of artificially infested seedlots with 0.1% infestation
or greater. In addition, using the PLP assay, 4 of 8 melon seedlots
collected from Xinjang province and 15 of 47 watermelon seedlots
collected from Ningxia province were positive for A. citrulli. In contrast,
a conventional polymerase chain reaction (PCR) assay that relied
on primers WFB1 and WFB2 facilitated A. citrulli detection in 1 of 8
and 5 of 47 seedlots from Xinjiang and Ningxia provinces, respectively.
These data indicate that the PLP and dot-blot hybridization technique
was more effective than conventional PCR for seed health testing.
[Show abstract][Hide abstract] ABSTRACT: Quorum sensing (QS) in Xanthomonas oryzae pv. oryzicola (Xoc), the causal agent of bacterial leaf streak, is mediated by the diffusible signal factor (DSF). DSF-mediating QS has been shown to control virulence and a set of virulence-related functions, however, the expression profiles and functions of extracellular proteins controlled by DSF signal remain largely unclear. In the present study, 33 DSF-regulated extracellular proteins, whose functions include small-protein mediating QS, oxidative adaptation, macromolecule metabolism, cell structure, biosynthesis of small molecules, intermediary metabolism, cellular process, protein catabolism and hypothetical function, were identified by proteomics in Xoc. Of these, 15 protein encoding genes were in-frame deleted, and 4 of them, including three genes encoding type II secretion system (T2SS)-dependent proteins and one gene encoding a Ax21 ( a ctivator of X A 21 -mediated immunity)-like protein (a novel small-protein type QS signal) were determined to be required for full virulence in Xoc. The contributions of these four genes to important virulence-associated functions, including bacterial colonization, extracellular polysaccharide, cell motility, biofilm formation and anti-oxidative ability are presented. To our knowledge, our analysis is the first complete list of DSF-regulated extracellular proteins and functions in a Xanthomonas species. Our results show that DSF-type QS played critical roles in regulation of T2SS and Ax21-mediating QS, which sheds light on the role of DSF signaling in Xanthomonas.
Journal of Proteome Research 05/2013; 12(7). DOI:10.1021/pr4001543 · 5.00 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Xanthomonas oryzae pv. oryzicola (Xoc) causes bacterial leaf streak in rice, which is a destructive disease worldwide. Xoc virulence factors are regulated by diffusible signal factor (DSF) and the global regulator Clp. In this study, we have demonstrated that asnB (XOC_3054), encoding an asparagine synthetase, is a novel virulence-related gene regulated by both DSF and Clp in Xoc. A sequence analysis revealed that AsnB is highly conserved in Xanthomonas. An asnB mutation in Xoc dramatically impaired pathogen virulence and growth rate in host rice, but did not affect the ability to trigger the hypersensitive response in nonhost (plant) tobacco. Compared with the wild-type strain, the asnB deletion mutant was unable to grow in basic MMX (-) medium (a minimal medium without ammonium sulphate as the nitrogen source) with or without 10 tested nitrogen sources, except asparagine. The disruption of asnB impaired pathogen resistance to oxidative stress and reduced the transcriptional expression of oxyR, katA and katG, which encode three important proteins responsible for hydrogen peroxide (H(2) O(2) ) sensing and detoxification in Xanthomonas in the presence of H(2) O(2) , and nine important known Xoc virulence-related genes in plant cell-mimicking medium. Furthermore, the asnB mutation did not affect extracellular protease activity, extracellular polysaccharide production, motility or chemotaxis. Taken together, our results demonstrate the role of asnB in Xanthomonas for the first time.
[Show abstract][Hide abstract] ABSTRACT: Xanthomonas oryzae pv. oryzicola causes bacterial leaf streak in rice, a destructive disease worldwide. In this study, six putative hypothetical secreted proteins, which were absent in X. oryzae pv. oryzae, were detected from X. oryzae pv. oryzicola strain BLS256. Disruption-based mutagenesis study revealed that one of them, Xoc_15235, named as extracellular polysaccharide and virulence-related gene (epv), was required for the optimal virulence in host rice but not for the induction of a hypersensitive reaction in nonhost tobacco. Sequence analysis revealed that epv was highly conserved in Xanthomonas spp. (except X. oryzae pv. oryzae). In-frame deletion of epv in X. oryzae pv. oryzicola dramatically impaired pathogen virulence and extracellular polysaccharide (EPS) production, one of the important known virulence-associated functions in Xanthomonas spp. Quantitative real-time reverse-transcription polymerase chain reaction showed that expression of both gumB (a gene encoding exopolysaccharide xanthan biosynthesis export protein) and a known virulence-related gene, pgk (encoding phosphoglycerate kinase), were obviously reduced in the epv-deletion mutant compared with the wild-type strain Rs105. In addition, we observed that epv was positively regulated by both diffusible signal factor and global regulator Clp in X. oryzae pv. oryzicola. Taken together, the novel roles and genetics of epv of X. oryzae pv. oryzicola in the EPS production and virulence were investigated for the first time.
[Show abstract][Hide abstract] ABSTRACT: Here, three different suicide vectors were evaluated for the possibility of performing gene mutagenesis in strain OH11 using the chiA gene (accession number: DQ888611) as a new reporter. Suicide vector pEX18GM was selected, and it was successfully applied for disruption and in-frame deletions in the chiA gene in strain OH11, which was confirmed by PCR amplification and Southern hybridization. The chiA-deletion mutant OH11-3 did not have the ability to produce chitinase on chitine selection medium. Interestingly, the chiA-deletion mutants displayed wild-type antimicrobial activity against Saccharomyces cerevisiae, Magnaporthe grisea, Phytophthora capsici, Rhizoctonia solani, Sclerotinia sclerotiorum and Pythium ultimum. Our data suggest that chitinase might not be a unique lytic enzyme in controlling S. cerevisiae, M. grisea, P. capsici, and P. ultimum. R. solani, S. sclerotiorum. Also, suicide vector pEX18GM might be explored as a potential tool for gene deletions in L. enzymogenes, which will facilitate the molecular study of mechanisms of biological control in L. enzymogenes.
World Journal of Microbiology and Biotechnology (Formerly MIRCEN Journal of Applied Microbiology and Biotechnology) 02/2012; 28(2):549-57. DOI:10.1007/s11274-011-0846-8 · 1.35 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Virulence factors of Xanthomonas oryzae pv. oryzicola, the causal agent of bacterial leaf streak in rice, are regulated by a diffusible signal factor (DSF)-dependent quorum-sensing (QS) system. In this study, a novel pathogenicity-related gene, Xoryp_010100018570 (named hshB), of X. oryzae pv. oryzicola was characterized. hshB encodes a hydrolase with a putative signal peptide, which is a homolog of imidazolonepropionase. Bioinformatic analysis showed that hshB is relatively conserved in the genus Xanthomonas but the homologous gene of hshB was not found in X. oryzae pv. oryzae. Reverse-transcription polymerase chain reaction (PCR) analysis showed that hshB and its upstream gene, Xoryp_010100018565 (named hshA), are co-transcribed in X. oryzae pv. oryzicola. Subsequent experimental results indicated that mutation of hshB remarkably impaired the virulence, extracellular protease activity, extracellular polysaccharide production, growth in minimal medium, and resistance to oxidative stress and bismerthiazol of X. oryzae pv. oryzicola. Mutation of clp, encoding a global regulator, resulted in similar phenotypes. Real-time PCR assays showed that hshB transcription is positively regulated by clp and DSF, and induced by poor nutrition. Our study not only found a novel gene hshB regulated by DSF-dependent QS system and clp but also showed that hshB was required for virulence of X. oryzae pv. oryzicola.
[Show abstract][Hide abstract] ABSTRACT: Acidovorax citrulli (Ac) is an important bacterium that occurs in watermelon, melon and other cucurbits. It mainly damages watermelon and melon, and can cause leaf blight, fruit rot, and even mortality.
To verify the relationship between defects in the synthesis of histidine and the pathogenicity of Ac.
We generated a transposon (Tn5) mutant library on the background of strain xjl12 of Ac. Then we used subclone technology to identify the gene.
The mutant could not elicit the hypersensitive response (HR) in nonhost tobacco, and its virulence was reduced. It is impaired in hisC, which encodes the protein histidinolphosphate aminotransferase. The other three genes (hisA, hisB and hisD) involved in the process of histidine synthesis were also studied. These mutants could not elicit the hypersensitive response (HR) in nonhost tobacco; their virulence was reduced significantly and disease symptoms caused by mutants were delayed for 48 hours when compared to the wild type strain. By adding exogenous histidine, pathogenicity of the mutants was restored.
The change of the characteristics of the mutants was directly related to the synthesis of histidine.
[Show abstract][Hide abstract] ABSTRACT: To investigate functions of flgDxoc and flgExoc genes regulated by diffusible signal factor (DSF) in Xanthomonas oryzae pv. oryzicola(Xoc)Rs105.
TheflgDxoc and flgExoc genes were amplified by PCR. We constructed deltaflgDxoc and deltaflgExoc, the deletion mutants from Rs105 by using double crossover method, and determined cell morphology, motility, pathogenicity in host rice and hypersensitive response (HR) in nonhost tobacco. We tested the differential expression of flgDxoc and flgExoc gene by reverse transcriptional polymerase chain reaction (RT-PCR) between the wide type and deltarpfFxoc (the deletion mutant of rpfFxoc gene, which could not produce DSF).
We cloned flgDxoc and flgExoc from genomic DNA of Rs105. PCR and Southern blot analysis demonstrated that the flgDxoc and flgExoc genes were knocked out successfully. Both mutants were non-flagellated and significantly attenuated motility on the 0.3% semi-solid medium. The pathogenicity on rice were obviously attenuated in deltaflgDxoc and deltaflgExoc compared to the wild type. All the changes in mutant could be restored through the complementation. However, there was no significant difference in bacterial growth in MMX medium and induction of HR between mutant (deltaflgDxoc or deltaflgExoc) and the wild type. In addition, the results of RT-PCR demonstrated that the transcription level of flgDxoc and flgExoc were downregulated in deltarpfFxoc.
This study showed that expressions of flgDxoc and flgExoc were positively regulated by DSF, and necessary for flagellar hook assembly and flagellar structure in Xoc. Meanwhile, FlgD and FlgE contributed to pathogen's virulence, motility and chemotaxis, but no differences at growth rate in MMX medium and HR in nonhost. In addition, our results provided molecular evidences that the contribution of DSF-type quorum sensing to pathogen's virulence might be, at least partially, dependent on bacterial flagellar in Xoc.
[Show abstract][Hide abstract] ABSTRACT: A new specific monoclonal antibody against the organophosphorous pesticide fenthion was produced based on our previous study. A sensitive and specific Enzyme-Linked Immunosorbent Assay (ELISA) for fenthion based on the new immunizing/coating hapten combination was developed. In this study, the H1 which attempts to expose the aromatic ring group was conjugated with bovine serum albumin (BSA) for the immunogen. All of the haptens that have been synthesized were conjugated with ovalbumin (OVA) for the coating antigen. The efficient antibody/coating conjugated combinations were selected, and the optimized ELISA was developed. In the optimized system, the IC50 value was 5.8 ng · mL-1 with the detection limit (IC20) of 0.028 ng · mL-1. The cross reactivity with all of the tested pesticides was lower than the 0.5%. Also, the system can detect the fenthion addition to the real samples such as soil, water, rice, and Chinese cabbages.
[Show abstract][Hide abstract] ABSTRACT: Acidovorax avenae subsp. citrulli is a Gram-negative bacterium and is the causal agent of bacterial fruit blotch (BFB) in cucurbits. In this study, the role
played by the acyl-homoserine lactone (AHL)-type quorum sensing (QS) system in growth, swimming motility and virulence was
characterized in A.
avenae subsp. citrulli strain XJL12. The AHL synthase gene of the QS system from strain XJL12, defined as aacI, was cloned and characterized, and an aacI disruption mutant was generated. The aacI mutant XJL13 abolished the ability to produce AHL molecules, whereas the corresponding complemented strain CPXJL13 produced
wild-type levels of AHL. The aacI mutant exhibited a significant decrease in growth rate relative to the wild type in minimal medium, and was partially impaired
in swimming motility. In plants, the aacI mutant showed a significant reduction of virulence in watermelon fruits and melon seedlings when compared to the wild-type
strain. However, the aacI mutation in strain XJL12 had no effects on biofilm formation, exopolysaccharide production, or induction of hypersentitive
response in Nicotiana tabacum. Our data suggest that the AHL-type QS may play a key role in pathogen virulence and this may provide an opportunity to explore
novel approaches for managing BFB in cucurbits by QS interference.
Acidovorax avenae subsp. citrulli
–Autoinducer–Quorum sensing–Bacterial fruit blotch
World Journal of Microbiology and Biotechnology 01/2011; 27(5):1155-1166. DOI:10.1007/s11274-010-0562-9 · 1.35 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Xanthomonas oryzae pv. oryzicola (Xoc), which caused bacterial leaf streak in rice, is a bacterial pathogen limited to the apoplast of the mesophyll tissue. The rpfF that encodes diffusible signal factor (DSF) synthase, played a key role in the virulence of many plant pathogenic bacteria. In this study, the rpf gene cluster was cloned, and the rpfF was deleted in Xoc. It was observed that the rpfF mutant lost the ability to produce DSF molecular, and exhibited a significant reduction of virulence in rice compared to the wild-type strain. Furthermore, the mutation of rpfF impaired EPS production, and led to Xoc cell aggregation. To analyze the differences of proteome expression between Xoc wild type and rpfF mutant, a comparative proteome analysis was performed by two-dimensional gel electrophoresis (2-DE). The results clearly revealed that 48 protein spots were differentially expressed above the threshold ratio of 1.5. Among them, 18 proteins were identified by MS, which were involved in nitrogen transfer, protein folding, elimination of superoxide radicals and flagellar formation. Our results indicated that DSF might play an important role in virulence and growth of Xoc by mediating expression of proteins.
[Show abstract][Hide abstract] ABSTRACT: A rapid (less than 10 min), qualitative and semi-quantitative immunochromatography using colloidal gold-antibody probe was successfully developed and applied in determination of chlorpyrifos-methyl (a wide-spectrum organophosphorus pesticide) in water samples. The qualitative detection limit of chlorpyrifos-methyl was determined as 0.6 μg ml(-1) by using immunochromatography. In the semi-quantitative experiment, the detection results of chlorpyrifos-methyl were scanned by a membrane strip reader, and a detection curve representing the scanned data average was obtained. After conversion, it was observed that in the range of 50-12,150 ng ml(-1), the graph between logit(B/B(0)) and logarithm of concentration of chlorpyrifos-methyl was linear, from which, the regression equation (y =-2.5229 x +7.5951, R(2)=0.9889) and IC(50) value (1024.39 ng ml(-1)) was obtained, respectively. Meanwhile, the detection limit was calculated as 132.91 ng ml(-1) by the extrapolation of B(0)-2SD. In addition, the cross-reactivities were less than 1% with tested analog compounds and regarded as negligible. The recoveries obtained by standard chlorpyrifos-methyl addition to water samples were 102.5-107.6%. Overall, to our knowledge, this is the first report of qualitative and semi-quantitative detection of chlorpyrifos-methyl by immunochromatography.
[Show abstract][Hide abstract] ABSTRACT: Bacterial soft rot caused by Pectobacterium carotovorum subsp. carotovorum is a serious plant disease in Zantedeschia spp. (also called calla lily). In this study, two independent genes (a N-acyl homoserine lactonase gene attM from Agrobacterium tumefaciens and a hypersensitive response and pathogenicity gene hrf1 from Xanthomonas oryzae pv. oryzae), transcribed by a strong and constitutive Escherichia coli promoter P
, respectively, were cloned into plasmid pUC19, and was transformed into E. coli, creating strain JM109/pPHA. The result of sodium dodecyl sulfate polyacrylamide gel electrophoresis (SDS–PAGE) assay showed
that both genes (hrf1 and attM) were successfully expressed in one plasmid system in strain JM109/pPHA. The expressed HarpinXoo (Hrf1) and AttM protein had the ability of inducing hypersensitive response (HR) in nonhost tobacco and degrading the N-acyl homoserine lactones (AHLs) produced by P. carotovorum subsp. carotovorum, respectively, whereas HarpinXoo and AttM protein did not seem to interfere with the normal growth of this pathogen. In planta, strain JM109/pPHA could significantly
reduce the soft rot disease severity on dormant tubers (control efficiency: 92.8%) or potted plants (control efficiency: 92.4%)
of calla lily. We have first demonstrated the both biocontrol effects of HarpinXoo and AttM proteins (also described as Quorum interference) on the bacterial soft rot disease of calla lily, caused by P. carotovorum subsp. carotovorum. This work provided a potential way to control this serious plant disease.
N-acyl homoserine lactonase AttM–
Pectobacterium carotovorum subsp. carotovorum
World Journal of Microbiology and Biotechnology 02/2010; 27(2):401-410. DOI:10.1007/s11274-010-0471-y · 1.35 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: A generic hapten (H1), 3-(4-Dimethoxyphosphorothioyloxy phenyl)propanoic acid, was synthesized to produce monoclonal antibodies (mAbs) for the determination of organophosphorus pesticides (OPs) in a class-specific manner. Six heterologous haptens were designed to study the effect of hapten heterology on immunoassay sensitivity. Several mice were immunized with this H1-BSA immunogen. Spleen cells of two immunized mice were fused with myeloma cells, and the resulting hybridomas were screened using H1-OVA. In a competitive indirect enzyme-linked immunosorbent assay (ELISA) format, three hybridoma cell lines (B4-C6, D12-B5, E5-H2) that produced mAbs with high selectivity and broad specificity were selected and expanded. The monoclonal antibody D12-B5 with higher titer was chosen for further study. In heterologous assay, the combination of D12-B5 and coating antigen H7-OVA constituted a particularly sensitive assay for competitive indirect ELISA and showed broad specificity for the determination of OPs, including parathion-methyl, chlorpyrifos-methyl, tolclofos-methyl, fenthion, malathion, fenitrothion. This combination resulted in the IC50 of 0.58–10.47 µg/ml.
[Show abstract][Hide abstract] ABSTRACT: Xanthomonas oryzae pv. oryzae causes bacterial leaf blight, one of the most widespread and destructive bacterial diseases in rice. This study identified and characterized the contribution of the twin-arginine translocation (Tat) pathway to motility, chemotaxis, extracellular polysaccharide (EPS) production and virulence in X. oryzae pv. oryzae strain PXO99. The tatC disruption mutant (strain TCM) of strain PXO99 were generated, and confirmed both by PCR and Southern blotting. Strain PXO99 cells were highly motile in NYGB 0.3% soft agar plate. In contrast, the tatC mutation impaired motility. Furthermore, strain TCM cells lacked detectable flagella and exhibited almost no chemotaxis toward glucose under aerobic conditions, indicating that the Tat secretion pathway contributed to flagellar biogenesis and chemotactic responses. It was also observed that strain TCM exhibited a reductive production of extracellular polysaccharide (EPS) and a significant reduction of virulence on rice plants when compared with the wild type PXO99. However, the tatC mutation in strain PXO99 did not affect growth rate and the ability to induce hypersensitive response (HR) in nonhost tobacco (Nicotiana tabacum L. cv. Samsun). Our findings indicated that the Tat system of X. oryzae pv. oryzae played an important role in the pathogen's virulence.
Archives of Microbiology 12/2008; 191(2):163-70. DOI:10.1007/s00203-008-0440-0 · 1.86 Impact Factor
[Show abstract][Hide abstract] ABSTRACT: Harpins constitute one group of effector proteins which elicit a hypersensitive response in nonhost plants, but the subcellular localization and tissue distribution of harpin receptors are still controversial. Antigen mimicry by anti-idiotypic antibodies is employed as a reliable strategy to probe receptors that are present in very low concentrations in the organism. In this study, a monoclonal anti-idiotypic antibody (Ab2), named 6B2, was elicited by F(ab′)2 fragments digested from the purified polyclonal antibody specific for HarpinXoo (Ab1). 6B2 competed with HarpinXoo for binding to Ab1 and the total protein extracted from tobacco leaves indicated its anti-idiotypic character and internal image property. The relevance of antigen mimicry was further confirmed by eliciting a third generation antibody (Ab3), which was shown not only to bind to Ab2 competing with Ab1 but also to react with the original antigen, HarpinXoo. Taken together, these results demonstrate functional and biochemical mimicry of HarpinXoo by Ab2 and suggest that 6B2 can be a useful tool in probing its receptor in nonhost plants and other downstream studies.