Design, Synthesis, and Antibacterial Evaluation of Novel Isoindolin-1-ones Derivatives Containing Piperidine Fragments.

A series of novel isoindoline-1-one derivatives containing piperidine moiety were designed and synthesized using natural compounds as raw materials, and their biological activities were tested for three bacterial and three fungal pathogens. These derivatives exhibited good against phytopathogenic bacteria activities against Pseudomonas syringae pv actinidiae (Psa) and Xanthomonas axonopodis pv.citri (Xac). Some compounds exhibited excellent antibacterial activities against Xanthomonas oryzae pv oryzae (Xoo). The dose of Y8 against Xoo (the maximum half lethal effective concentration (EC50) = 21.3 µg/mL) was better than that of the thiediazole copper dose (EC50 = 53.3 µg/mL). Excitingly, further studies have shown that the molecular docking of Y8 with 2FBW indicates that it can fully locate the interior of the binding pocket through hydrogen bonding and hydrophobic interactions, thereby enhancing its anti-Xoo activity. Scanning electron microscopy (SEM) studies revealed that Y8 induced the Xoo cell membrane collapse. Moreover, the proteomic results also indicate that Y8 may be a multifunctional candidate as it affects the formation of bacterial Xoo biofilms, thereby exerting antibacterial effects.

The PilB-PilZ-FimX regulatory complex of the Type IV pilus from Xanthomonas citri.

Type IV pili (T4P) are thin and flexible filaments found on the surface of a wide range of Gram-negative bacteria that undergo cycles of extension and retraction and participate in a variety of important functions related to lifestyle, defense and pathogenesis. During pilus extensions, the PilB ATPase energizes the polymerization of pilin monomers from the inner membrane. In Xanthomonas citri, two cytosolic proteins, PilZ and the c-di-GMP receptor FimX, are involved in the regulation of T4P biogenesis through interactions with PilB. In vivo fluorescence microscopy studies show that PilB, PilZ and FimX all colocalize to the leading poles of X. citri cells during twitching motility and that this colocalization is dependent on the presence of all three proteins. We demonstrate that full-length PilB, PilZ and FimX can interact to form a stable complex as can PilB N-terminal, PilZ and FimX C-terminal fragments. We present the crystal structures of two binary complexes: i) that of the PilB N-terminal domain, encompassing sub-domains ND0 and ND1, bound to PilZ and ii) PilZ bound to the FimX EAL domain within a larger fragment containing both GGDEF and EAL domains. Evaluation of PilZ interactions with PilB and the FimX EAL domain in these and previously published structures, in conjunction with mutagenesis studies and functional assays, allow us to propose an internally consistent model for the PilB-PilZ-FimX complex and its interactions with the PilM-PilN complex in the context of the inner membrane platform of the X. citri Type IV pilus.

Photodynamic inactivation of the phytopathogenic bacterium Xanthomonas citri subsp. citri.

The present work intended to evaluate the applicability of photodynamic inactivation (PDI) of Xanthomonas citri subsp. citri with toluidine blue O (TBO), a commercial photosensitizer, as a strategy to control citrus canker. Assays were conducted with cell suspensions and biofilms, constructed either on polypropylene microtubes (in vitro assays) or on the surface of orange leaves (ex vivo assays), in the presence of TBO and under irradiation with artificial white light or natural sunlight. PDI assays using TBO alone caused a maximum 5·8 log10 reduction of X. citri viable cells in suspensions, and a much smaller inactivation (1·5 log10) in biofilms. However, concomitant use of KI potentiated the TBO photosensitization. Biofilms were inactivated down to the detection limit (>6 log10 reduction) with 5·0 µmol l-1 TBO + 10 mmol l-1 KI (in vitro) or 5·0 µmol l-1 TBO + 100 mmol l-1 KI (ex vivo) after artificial white light irradiation. Under natural sunlight, a reduction down to the detection limit of the Miles-Misra method was achieved with 50 µmol l-1 TBO and 100 mmol l-1 KI. PDI has potential to be applied in the control of citrus canker in field conditions although further studies are needed to show that there are no risks to plant physiology or fruit quality. SIGNIFICANCE AND IMPACT OF THE STUDY: Xanthomonas citri subsp. citri is a major cause of disease in citrus orchards. Because of the low efficacy and high environmental toxicity of copper-based treatments, there is growing interest on more sustainable phytosanitary approaches. Photodynamic inactivation (PDI) is being successfully used to control infectious agents and literature reports indicate that it is effective against some fungi and bacteria attacking fruit crops. The results of the present work open the perspective of using a low-cost photosensitizer and sunlight, as energy source, to control of the causative agent of citrus canker.

Rice OsAAA-ATPase1 is Induced during Blast Infection in a Salicylic Acid-Dependent Manner, and Promotes Blast Fungus Resistance.

Fatty acids (FAs) have been implicated in signaling roles in plant defense responses. We previously reported that mutation or RNAi-knockdown (OsSSI2-kd) of the rice OsSSI2 gene, encoding a stearoyl acyl carrier protein FA desaturase (SACPD), remarkably enhanced resistance to blast fungus Magnaporthe oryzae and the leaf-blight bacterium Xanthomonas oryzae pv. oryzae (Xoo). Transcriptomic analysis identified six AAA-ATPase family genes (hereafter OsAAA-ATPase1-6) upregulated in the OsSSI2-kd plants, in addition to other well-known defense-related genes. Here, we report the functional analysis of OsAAA-ATPase1 in rice's defense response to M. oryzae. Recombinant OsAAA-ATPase1 synthesized in Escherichia coli showed ATPase activity. OsAAA-ATPase1 transcription was induced by exogenous treatment with a functional analogue of salicylic acid (SA), benzothiadiazole (BTH), but not by other plant hormones tested. The transcription of OsAAA-ATPase1 was also highly induced in response to M. oryzae infection in an SA-dependent manner, as gene induction was significantly attenuated in a transgenic rice line expressing a bacterial gene (nahG) encoding salicylate hydroxylase. Overexpression of OsAAA-ATPase1 significantly enhanced pathogenesis-related gene expression and the resistance to M. oryzae; conversely, RNAi-mediated suppression of this gene compromised this resistance. These results suggest that OsAAA-APTase1 plays an important role in SA-mediated defense responses against blast fungus M. oryzae.

Gene expression analysis identifies hypothetical genes that may be critical during the infection process of Xanthomonas citri subsp. citri

Background: Gene expression analysis via microarray is widely used in phytobacteria to validate differential gene expression associated with virulence or to compare biological profiles of wild type and mutant strains. Here, we employed DNA microarrays to study the early stages of the infection process (24, 72 and 120 h post-inoculation) of Xanthomonas citri subsp. citri (Xac) infecting Citrus sinensis to interrogate the expression profiles of hypothetical genes. Results: Under infective conditions, 446 genes were up- and 306 downregulated. Outstanding among genes upregulated during infection were those involved in synthesizing the Type 3 Secretion System and effectors, xanthan gum and quorum-sensing induction, and flagellum synthesis and regulation. Additionally, 161 hypothetical genes were up- and 100 were downregulated, 49 of which are known to have a significant biological role. To understand hypothetical gene co-regulation or -expression, nine expression profiles including 158 genes were identified during the three infection phases. Of these, 47 hypothetical genes were identified as having expression profiles associated with at least one connected to a gene associated with adaptation and virulence. Conclusions: Expression patterns of six differentially expressed genes were validated by quantitative reverse transcription polymerase chain reaction, thus demonstrating the effectiveness of this tool in global gene expression analysis in Xac.

Small Molecule Inhibitors Specifically Targeting the Type III Secretion System of Xanthomonas oryzae on Rice.

The initiative strategy for the development of novel anti-microbial agents usually uses the virulence factors of bacteria as a target, without affecting their growth and survival. The type III secretion system (T3SS), one of the essential virulence factors in most Gram-negative pathogenic bacteria because of its highly conserved construct, has been regarded as an effective target that developed new anti-microbial drugs. Xanthomonas oryzae pv. oryzae (Xoo) causes leaf blight diseases and is one of the most important pathogens on rice. To find potential anti-virulence agents against this pathogen, a number of natural compounds were screened for their effects on the T3SS of Xoo. Three of 34 compounds significantly inhibited the promoter activity of the harpin gene, hpa1, and were further checked for their impact on bacterial growth and on the hypersensitive response (HR) caused by Xoo on non-host tobacco plants. The results indicated that treatment of Xoo with CZ-1, CZ-4 and CZ-9 resulted in an obviously attenuated HR without affecting bacterial growth and survival. Moreover, quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis showed that the expression of the Xoo T3SS was suppressed by treatment with the three inhibitors. The mRNA levels of representative genes in the hypersensitive response and pathogenicity (hrp) cluster, as well as the regulatory genes hrpG and hrpX, were reduced. Finally, the in vivo test demonstrated that the compounds could reduce the disease symptoms of Xoo on the rice cultivar (Oryza sativa) IR24.

Antimicrobial activity of linear lipopeptides derived from BP100 towards plant pathogens.

A collection of 36 lipopeptides were designed from the cecropin A-melittin hybrid peptide BP100 (H-Lys-Lys-Leu-Phe-Lys-Lys-Ile-Leu-Lys-Tyr-Leu-NH2) previously described with activity against phytopathogenic bacteria. These lipopeptides were synthesized on solid-phase and screened for their antimicrobial activity, toxicity and proteolytic stability. They incorporated a butanoyl, a hexanoyl or a lauroyl group at the N-terminus or at the side chain of a lysine residue placed at each position of the sequence. Their antimicrobial activity and hemolysis depended on the fatty acid length and its position. In particular, lipopeptides containing a butanoyl or a hexanoyl chain exhibited the best biological activity profile. In addition, we observed that the incorporation of the acyl group did not induce the overexpression of defense-related genes in tomato. Best lipopeptides were BP370, BP378, BP381, BP387 and BP389, which were highly active against all the pathogens tested (minimum inhibitory concentration of 0.8 to 12.5 µM), low hemolytic, low phytotoxic and significantly stable to protease degradation. This family of lipopeptides might be promising functional peptides useful for plant protection.

Synthesis and bioactivity of thiazolidin-2-cyanamide derivatives against type III secretion system of Xanthomonas oryzae on rice.

Targeting virulence factors of bacterial without affecting their growth and survival, has been an initiative strategy for the development of novel anti-microbial agents. The type III secretion system (T3SS), one of essential and highly conserved virulence factors in most Gram-negative pathogenic bacteria, has been regarded as an effective target that developed new anti-microbial drugs. Xanthomonas oryzae pv. oryzae (Xoo) is one of the most Important bacterial pathogens on rice, which causes leaf blight disease. To discover potential anti-virulence agents against the pathogens, a new series of thiazolidin-2-cyanamide derivatives containing 5-phenyl-2-furan were designed and synthesized. Their structures were characterized by 1H NMR, 13C NMR, MS, and elemental analysis. All the title compounds inhibited the promoter activity of a harpin gene hpa1, significantly, that were further checked for the impact on bacterial growth and on the hypersensitive response (HR) caused by Xoo on non-host tobacco plants. The results indicated that treatment of Xoo with the title compounds II-2, II-3 and II-4 resulted in significantly attenuated HR without affecting bacterial growth or survival. Moreover, quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis showed that the expression of the Xoo T3SS was suppressed by treatment with the three inhibitors. The mRNA levels of representative genes in the hrp (hypersensitive response and pathogenicity) cluster, as well as the regulatory genes hrpG and hrpX, were reduced. Finally, the in vivo test demonstrated that the compounds could reduce the disease symptoms of Xoo on the rice cultivar (Oryza sativa) IR24.

Glycolate oxidase gene family in Nicotiana benthamiana: genome-wide identification and functional analyses in disease resistance.

Glycolate oxidase (GOX)-dependent production of H2O2 in response to pathogens and its function in disease resistance are still poorly understood. In this study, we performed genome-wide identification of GOX gene family in Nicotiana benthamiana and analyzed their function in various types of disease resistance. Sixteen GOX genes were identified in N. benthamiana genome. They consisted of GOX and HAOX groups. All but two NbGOX proteins contained an alpha_hydroxyacid_oxid_FMN domain with extra 43-52 amino acids compared to that of FMN-dependent alpha-hydroxyacid oxidizing enzymes (NCBI-CDD cd02809). Silencing of three NbGOX family genes NbHAOX8, NbGOX1 and NbGOX4 differently affected resistance to various pathogens including Tobacco rattle virus, Xanthomonas oryzae pv. oryzae (Xoo) and Sclerotinia sclerotiorum. Effect of these genes on resistance to Xoo is well correlated with that on Xoo-responsive H2O2 accumulation. Additionally, silencing of these genes enhanced PAMP-triggered immunity as shown by increased flg22-elicited H2O2 accumulation in NbGOX-silenced plants. These NbGOX family genes were distinguishable in altering expression of defense genes. Analysis of mutual effect on gene expression indicated that NbGOX4 might function through repressing NbHAOX8 and NbGOX1. Collectively, our results reveal the important roles and functional complexity of GOX genes in disease resistance in N. benthamiana.

Low-iron conditions induces the hypersensitive reaction and pathogenicity hrp genes expression in Xanthomonas and is involved in modulation of hypersensitive response and virulence.

Expression of hrp (hypersensitive reaction and pathogenicity) genes inside the host is crucial for virulence of phytopathogenic bacteria. The hrp genes encode components of type3 secretion system (T3SS), HR elicitors and several regulators, which are involved in the co-ordinated expression of hrp genes in the host environment and in hrp inducing chemically defined medium. However, little is known about specific host or environmental factors which may play a role in the induction of hrp gene expression. In this study, we show that iron-limiting condition elicits induced expression of hrp genes, including type3 secretion system (T3SS) and effectors (T3E). Expression analysis using qRT-PCR and promoter probe strains suggest significant induction in the expression of Hrp and T3S-associated genes of Xanthomonas campestris pv. campestris (Xcc) under low-iron condition, and is suppressed by exogenous supplementation of iron. Furthermore, we show that with exogenous iron supplementation, wild type Xcc exhibited reduced disease symptoms in host-plant, and exhibited significant reduction in HR and callose deposition in the non-host plants. Xanthomonas oryzae and oryzicola pathovars also exhibited the iron affect, albeit to a lesser extend compared with the Xcc. Overall, our results suggest that low-iron condition inside the host may play a crucial role in pathogenicity.

Artificial Agrobacterium tumefaciens strains exhibit diverse mechanisms to repress Xanthomonas oryzae pv. oryzae-induced hypersensitive response and non-host resistance in Nicotiana benthamiana.

Xanthomonas oryzae pv. oryzae (Xoo) rapidly triggers a hypersensitive response (HR) and non-host resistance in its non-host plant Nicotiana benthamiana. Here, we report that Agrobacterium tumefaciens strain GV3101 blocks Xoo-induced HR in N. benthamiana when pre-infiltrated or co-infiltrated, but not when post-infiltrated at 4 h after Xoo inoculation. This suppression by A. tumefaciens is local and highly efficient to Xoo. The HR-inhibiting efficiency of A. tumefaciens is strain dependent. Strain C58C1 has almost no effect on Xoo-induced HR, whereas strains GV3101, EHA105 and LBA4404 nearly completely block HR formation. Intriguingly, these three HR-inhibiting strains employ different strategies to repress HR. Strain GV3101 displays strong antibiotic activity and thus suppresses Xoo growth. Comparison of the genotype and Xoo antibiosis activity of wild-type A. tumefaciens strain C58 and a set of C58-derived strains reveals that this Xoo antibiosis activity of A. tumefaciens is negatively, but not solely, regulated by the transferred-DNA (T-DNA) of the Ti plasmid pTiC58. Unlike GV3101, strains LBA4404 and EHA105 exhibit no significant antibiotic effect on Xoo, but rather abolish hydrogen peroxide accumulation. In addition, expression assays indicate that strains LBA4404 and EHA105 may inhibit Xoo-induced HR by suppression of the expression of Xoo type III secretion system (T3SS) effector genes hpa1 and hrpD6. Collectively, our results unveil the multiple levels of effects of A. tumefaciens on Xoo in N. benthamiana and provide insights into the molecular mechanisms underlying the bacterial antibiosis of A. tumefaciens and the non-host resistance induced by Xoo.

Identification of phenolic compounds that suppress the virulence of Xanthomonas oryzae on rice via the type III secretion system.

The targeting of bacterial type III secretion systems (T3SSs), which are critical virulence factors in most Gram-negative pathogens, is regarded as an alternative strategy for the development of novel anti-microbial drugs. Xanthomonas oryzae pv. oryzae (Xoo) and X. oryzae pv. oryzicola (Xoc) are two of the most important bacterial pathogens on rice, which cause leaf blight and leaf streak diseases, respectively. To identify potential anti-virulence drugs against these two pathogens, we screened a library of plant phenolic compounds and derivatives for their effects on the Xoo T3SS. Ten of 56 compounds significantly inhibited the promoter activity of a harpin gene, hpa1. These inhibitors were further tested for their impact on the hypersensitive response (HR) caused by Xoo on non-host tobacco plants. The results showed that pretreatment of Xoo with TS006 (o-coumaric acid, OCA), TS010, TS015 and TS018 resulted in significantly attenuated HR without affecting bacterial growth or survival. In addition, Cya translocation assays demonstrated that the translocation of two T3 effectors was suppressed by the four inhibitors. Quantitative reverse transcription-polymerase chain reaction (qRT-PCR) analysis showed that mRNA levels of representative genes in the hrp (hypersensitive response and pathogenicity) cluster, as well as the regulatory genes hrpG and hrpX, were reduced by treatment with the four inhibitors, suggesting that expression of the Xoo T3SS was suppressed. The expression of other virulence factors was not suppressed, which indicated possible T3SS-specific inhibition. Finally, we demonstrated that these inhibitors reduced the disease symptoms of Xoo and Xoc on the rice cultivar (Oryza sativa) IR24 to varying extents.

Rapid, Bioassay-Guided Process for the Detection and Identification of Antibacterial Neem Oil Compounds.

Bioassay guidance was used along the whole process including method development, isolation and identification of antibacterial neem (Azadirachta indica) oil compounds. The biomonitoring was performed by direct bioautography (DB), a combination of thin-layer chromatography (TLC) and antimicrobial detection. DB of neem oil showed one antibacterial zone that was not UV-active; therefore, the TLC separation was improved under DB control. The chromatographic zone that exhibited activity against Bacillus subtilis, Xanthomonas euvesicatoria, Aliivibrio fischeri, Staphylococcus aureus and methicillin-resistant Staphylococcus aureus was characterized by TLC reagents, indicating a lipophilic, fatty acid-like chemical feature. Two compounds were found and identified in the active zone by high-performance liquid chromatography-electrospray ionization mass spectrometry as linoleic and oleic acids. Both fatty acids inhibited B. subtilis, but A. fischeri was sensitive only against linoleic acid.

Niclosamide inhibits leaf blight caused by Xanthomonas oryzae in rice.

Rice leaf blight, which is caused by the bacterial pathogen Xanthomonas oryzae pv. oryzae (Xoo), results in huge losses in grain yield. Here, we show that Xoo-induced rice leaf blight is effectively controlled by niclosamide, an oral antihelminthic drug and molluscicide, which also functions as an anti-tumor agent. Niclosamide directly inhibited the growth of the three Xoo strains PXO99, 10208 and K3a. Niclosamide moved long distances from the site of local application to distant rice tissues. Niclosamide also increased the levels of salicylate and induced the expression of defense-related genes such as OsPR1 and OsWRKY45, which suppressed Xoo-induced leaf wilting. Niclosamide had no detrimental effects on vegetative/reproductive growth and yield. These combined results indicate that niclosamide can be used to block bacterial leaf blight in rice with no negative side effects.

Chemical products induce resistance to Xanthomonas perforans in tomato.

The bacterial spot of tomato, caused by Xanthomonas spp., is a very important disease, especially in the hot and humid periods of the year. The chemical control of the disease has not been very effective for a number of reasons. This study aimed to evaluate, under greenhouse conditions, the efficacy of leaf-spraying chemicals (acibenzolar-S-methyl (ASM) (0.025 g.L(-1)), fluazinam (0.25 g.L(-1)), pyraclostrobin (0.08 g.L(-1)), pyraclostrobin + methiran (0.02 g.L(-1) + 2.2 g.L(-1)), copper oxychloride (1.50 g.L(-1)), mancozeb + copper oxychloride (0.88 g.L(-1) + 0.60 g.L(-1)), and oxytetracycline (0.40 g.L(-1))) on control of bacterial spot. Tomatoes Santa Clara and Gisele cultivars were pulverized 3 days before inoculation with Xanthomonas perforans. The production of enzymes associated with resistance induction (peroxidase, polyphenol oxidase, phenylalanine ammonia-lyase, ß-1,3-glucanase, and protease) was quantified from leaf samples collected 24 hours before and 24 hours after chemical spraying and at 1, 2, 4, 6, and 8 days after bacterial inoculation. All products tested controlled bacterial spot, but only ASM, pyraclostrobin, and pyraclostrobin + metiram increased the production of peroxidase in the leaves of the two tomato cultivars, and increased the production of polyphenol oxidase and ß-1,3-glucanase in the Santa Clara cultivar.

Chemical products induce resistance to Xanthomonas perforans in tomato

The bacterial spot of tomato, caused by Xanthomonas spp., is a very important disease, especially in the hot and humid periods of the year. The chemical control of the disease has not been very effective for a number of reasons. This study aimed to evaluate, under greenhouse conditions, the efficacy of leaf-spraying chemicals (acibenzolar-S-methyl (ASM) (0.025 g.L⁻¹), fluazinam (0.25 g.L⁻¹), pyraclostrobin (0.08 g.L⁻¹), pyraclostrobin + methiran (0.02 g.L⁻¹ + 2.2 g.L⁻¹), copper oxychloride (1.50 g.L⁻¹), mancozeb + copper oxychloride (0.88 g.L⁻¹ + 0.60 g.L⁻¹), and oxytetracycline (0.40 g.L⁻¹)) on control of bacterial spot. Tomatoes Santa Clara and Gisele cultivars were pulverized 3 days before inoculation with Xanthomonas perforans. The production of enzymes associated with resistance induction (peroxidase, polyphenol oxidase, phenylalanine ammonia-lyase, β-1,3-glucanase, and protease) was quantified from leaf samples collected 24 hours before and 24 hours after chemical spraying and at 1, 2, 4, 6, and 8 days after bacterial inoculation. All products tested controlled bacterial spot, but only ASM, pyraclostrobin, and pyraclostrobin + metiram increased the production of peroxidase in the leaves of the two tomato cultivars, and increased the production of polyphenol oxidase and β-1,3-glucanase in the Santa Clara cultivar.

XocR, a LuxR solo required for virulence in Xanthomonas oryzae pv. oryzicola.

Xanthomonas oryzae pv. oryzicola (Xoc) causes bacterial leaf streak (BLS) in rice, a serious bacterial disease of rice in Asia and parts of Africa. The virulence mechanisms of Xoc are not entirely clear and control measures for BLS are poorly developed. The solo LuxR proteins are widespread and shown to be involved in virulence in some plant associated bacteria (PAB). Here, we have cloned and characterized a PAB LuxR solo from Xoc, named as XocR. Mutation of xocR almost completely impaired the virulence ability of Xoc on host rice, but did not alter the ability to trigger HR (hypersensitive response, a programmed cell death) on non-host (plant) tobacco, suggesting the diversity of function of xocR in host and non-host plants. We also provide evidence to show that xocR is involved in the regulation of growth-independent cell motility in response to a yet-to-be-identified rice signal, as mutation of xocR impaired cell swimming motility of wild-type Rs105 in the presence but not absence of rice macerate. We further found that xocR regulated the transcription of two characterized virulence-associated genes (recN and trpE) in the presence of rice macerate. The promoter regions of recN and trpE possessed a potential binding motif (an imperfect pip box-like element) of XocR, raising the possibility that XocR might directly bind the promoter regions of these two genes to regulate their transcriptional activity. Our studies add a new member of PAB LuxR solos and also provide new insights into the role of PAB LuxR solo in the virulence of Xanthomonas species.

Cell-cell signalling promotes ferric iron uptake in Xanthomonas oryzae pv. oryzicola that contribute to its virulence and growth inside rice.

Cell-cell communication mediated by diffusible signal factor (DSF) plays an important role in virulence of several Xanthomonas group of plant pathogens. In the bacterial pathogen of rice, Xanthomonas oryzae pv. oryzicola, DSF is required for virulence and in planta growth. In order to understand the role of DSF in promoting in planta growth and virulence, we have characterized the DSF deficient mutant of X. oryzae pv. oryzicola. Mutant analysis by expression analysis, radiolabelled iron uptake studies and growth under low-iron conditions indicated that DSF positively regulates ferric iron uptake. Further, the DSF deficient mutant of X. oryzae pv. oryzicola exhibited a reduced capacity to use ferric form of iron for growth under low-iron conditions. Exogenous iron supplementation in the rice leaves rescued the in planta growth deficiency of the DSF deficient mutant. These data suggest that DSF promotes in planta growth of X. oryzae pv. oryzicola by positively regulating functions involved in ferric iron uptake which is important for its virulence. Our results also indicate that requirement of iron uptake strategies to utilize either Fe(3+) or Fe(2+) form of iron for colonization may vary substantially among closely related members of the Xanthomonas group of plant pathogens.

[THE SENSITIVITY OF PHYTOPATHOGENIC BACTERIA TO STREPTOMYCIN UNDER THE INFLUENCE OF PESTICIDES].

The results of the streptomycin sensitivity changes of phytopathogenic Pseudomonas syringae and Xanthomonas translucens bacteria under the action of pesticides are pre- sented. It is demonstrated that phytopathogenic strains show greater changes of strepto- mycin sensitivity compared to epiphytic Pantoea agglomerans strain under the pesticides influence. Granstar herbicide, Tviks and Alpha Super insecticides increase the number of streptomycin resistant cells of Xanthomonas translucens 3164, P syringae pv. syringae YKM B-1027 and P syringae pv. atrofaciens YKM B-1011. This fact indicates mutagenic action of these pesticides against researched phytopathogenic bacteria.

Extracellular matrix-associated proteome changes during non-host resistance in citrus-Xanthomonas interactions.

Non-host resistance (NHR) is a most durable broad-spectrum resistance employed by the plants to restrict majority of pathogens. Plant extracellular matrix (ECM) is a critical defense barrier. Understanding ECM responses during interaction with non-host pathogen will provide insights into molecular events of NHR. In this study, the ECM-associated proteome was compared during interaction of citrus with pathogen Xanthomonas axonopodis pv. citri (Xac) and non-host pathogen Xanthomonas oryzae pv. oryzae (Xoo) at 8, 16, 24 and 48 h post inoculation. Comprehensive analysis of ECM-associated proteins was performed by extracting wall-bound and soluble ECM components using both destructive and non-destructive procedures. A total of 53 proteins was differentially expressed in citrus-Xanthomonas host and non-host interaction, out of which 44 were identified by mass spectrometry. The differentially expressed proteins were related to (1) defense-response (5 pathogenesis-related proteins, 3 miraculin-like proteins (MIR, MIR1 and MIR2) and 2 proteases); (2) enzymes of reactive oxygen species (ROS) metabolism [Cu/Zn superoxide dismutase (SOD), Fe-SOD, ascorbate peroxidase and 2-cysteine-peroxiredoxin]; (3) signaling (lectin, curculin-like lectin and concanavalin A-like lectin kinase); and (4) cell-wall modification (α-xylosidase, glucan 1, 3 ß-glucosidase, xyloglucan endotransglucosylase/hydrolase). The decrease in ascorbate peroxidase and cysteine-peroxiredoxin could be involved in maintenance of ROS levels. Increase in defense, cell-wall remodeling and signaling proteins in citrus-Xoo interaction suggests an active involvement of ECM in execution of NHR. Partially compromised NHR in citrus against Xoo, upon Brefeldin A pre-treatment supported the role of non-classical secretory proteins in this phenomenon.