RESUMO
Fully sequenced genomes of Xanthomonas campestris pv. campestris (Xcc) strains are reported. However, intra-pathovar differences are still intriguing and far from clear. In this work, the contrasting virulence between two isolates of Xcc - Xcc51 (more virulent) and XccY21 (less virulent) is evaluated by determining their pan proteome profiles. The bacteria are grown in NYG and XVM1 (optimal for induction of hrp regulon) broths and collected at the max-exponential growth phase. Shotgun proteomics reveals a total of 329 proteins when Xcc isolates are grown in XVM1. A comparison of both profiles reveals 47 proteins with significant abundance fluctuations, out of which, 39 show an increased abundance in Xcc51 and are mainly involved in virulence/adaptation mechanisms, genetic information processing, and membrane receptor/iron transport systems, such as BfeA, BtuB, Cap, Clp, Dcp, FyuA, GroEs, HpaG, Tig, and OmpP6. Several differential proteins are further analyzed by qRT-PCR, which reveals a similar expression pattern to the protein abundance. The data shed light on the complex Xcc pathogenicity mechanisms and point out a set of proteins related to the higher virulence of Xcc51. This information is essential for the development of more efficient strategies aiming at the control of black rot disease.
Assuntos
Proteínas de Bactérias/análise , Proteoma/análise , Fatores de Virulência/análise , Xanthomonas campestris/patogenicidade , Proteínas de Bactérias/genética , Meios de Cultura/química , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica/genética , Proteoma/genética , Virulência/genética , Fatores de Virulência/genética , Xanthomonas campestris/genética , Xanthomonas campestris/isolamento & purificaçãoRESUMO
Bacterial diseases caused by members of the genus Xanthomonas affect agricultural crops of great importance in the world. At least 350 different plant diseases are caused by species of Xanthomonas. Important crops, such as: rice, citrus, cassava, tomato, sugar cane, passionfruit and brassicas are severely affected by bacteria of this genus. Due to its rapid propagation, handling difficulties, problems with chemical control and severity of the losses of the affected plantations Xanthomonas is a difficult obstacle for agriculture around the world. In addition, chemical control of some of these diseases is carried out using copper-based chemicals, which causes a negative impact on health and the environment. A more sustainable alternative to combat these diseases is the control of Xanthomonas by microorganisms directly or indirectly through the use of its secondary metabolites involved in biocontrol. This review is a report concerning the recent advances in the search for microorganisms for the biocontrol of several Xanthomonas that are important for the world economy.
Assuntos
Doenças das Plantas/microbiologia , Doenças das Plantas/prevenção & controle , Xanthomonas/patogenicidade , Bactérias/metabolismo , Agentes de Controle Biológico/metabolismo , Agentes de Controle Biológico/farmacologia , Produtos Agrícolas/microbiologia , Fungos/metabolismo , Controle Biológico de Vetores/métodos , Metabolismo Secundário , Xanthomonas/classificação , Xanthomonas/efeitos dos fármacos , Xanthomonas campestris/patogenicidadeRESUMO
Citrus is an economically important fruit crop that is severely afflicted by citrus canker, a disease caused by Xanthomonas citri ssp. citri (X. citri); thus, new sustainable strategies to manage this disease are needed. Although all Citrus spp. are susceptible to this pathogen, they are resistant to other Xanthomonas species, exhibiting non-host resistance (NHR), for example, to the brassica pathogen X. campestris pv. campestris (Xcc) and a gene-for-gene host defence response (HDR) to the canker-causing X. fuscans ssp. aurantifolii (Xfa) strain C. Here, we examine the plant factors associated with the NHR of C. limon to Xcc. We show that Xcc induced asymptomatic type I NHR, allowing the bacterium to survive in a stationary phase in the non-host tissue. In C. limon, this NHR shared some similarities with HDR; both defence responses interfered with biofilm formation, and were associated with callose deposition, induction of the salicylic acid (SA) signalling pathway and the repression of abscisic acid (ABA) signalling. However, greater stomatal closure was seen during NHR than during HDR, together with different patterns of accumulation of reactive oxygen species and phenolic compounds and the expression of secondary metabolites. Overall, these differences, independent of Xcc type III effector proteins, could contribute to the higher protection elicited against canker development. We propose that Xcc may have the potential to steadily activate inducible defence responses. An understanding of these plant responses (and their triggers) may allow the development of a sustained and sustainable resistance to citrus canker.
Assuntos
Citrus/microbiologia , Doenças das Plantas/genética , Doenças das Plantas/microbiologia , Xanthomonas campestris/patogenicidade , Ácido Abscísico/metabolismo , Citrus/metabolismo , Regulação da Expressão Gênica de Plantas , Espécies Reativas de Oxigênio/metabolismo , Ácido Salicílico/metabolismoRESUMO
Black rot of crucifers, (Xanthomonas campestris pv. campestris) is the principal yield-limiting and destructive pathogen of cruciferous crop worldwide. In order to validate a bio-based control alternative for this disease, whey, lime sulfur, biofertilizer, Bordeaux mixture or raw milk were applied to kale (Brassica oleracea var. acephala) plants. The disease control was achieved by most of the tested products. Milk-based products (raw milk and whey) and biofertilizer reduced the severity by 44 and 56% in the field. Antioxidants, crude fibber, crude protein and lipid contents and kale yield were verified in the five treatments on the leaves with and without X. campestris pv. campestris inoculation. In the absence of the pathogen (non-inoculated), lime sulfur and Bordeaux mixture improved plant nutritional value compared to organic treatments, nevertheless milk-based products and biofertilizer improved the evaluated variables more than the control. However, on leaves inoculated with X. campestris pv. campestris raw milk increased antioxidant activity, crude protein and fiber contents, whereas biofertilizer increased kale yield, lipid and antioxidant contents. Milk-based products and biofertilizer were further evaluated in greenhouse trials to determinate the activity of defense-related enzymes and lignin content. Biofertilizer treatment resulted in increased phenylalanine ammonia lyase, catalase, peroxidase activities and lignin content. Hence, the application of milk-based products and biofertilizer are promising to control black rot of crucifers and also improves food quality by boosting nutritional values and antioxidant activity.
Assuntos
Antioxidantes/metabolismo , Brassica/efeitos dos fármacos , Suplementos Nutricionais/análise , Xanthomonas campestris/patogenicidade , Brassica/química , Brassica/microbiologia , Compostos de Cálcio/farmacologia , Cobre/farmacologia , Qualidade dos Alimentos , Valor Nutritivo , Doenças das Plantas/prevenção & controle , Extratos Vegetais/farmacologia , Sulfetos/farmacologia , Soro do Leite/químicaRESUMO
Several minimal media capable of inducing pathogenicity genes have been used to study plant-pathogen interactions. An in planta assay to study a closer interaction between the bacteria and the host was also developed and has been employed by our group. In order to determine whether growth medium could be improved to better approximate in planta conditions beyond that offered by the defined minimal medium XVM1, we compared the expression of 20 Xanthomonas campestris pv. campestris (Xcc) genes by quantitative reverse transcription - polymerase chain reaction (qRT-PCR) under in vivo (bacteria recovered from the plant) and in vitro (rich medium NYG, minimal medium XVM1 and XVM1 + leaf extract) growth systems. The results showed a higher expression level of the genes in the in planta system when compared to growth in culture media. In planta growth is closest to a real interaction condition and captures the complexity of the plant cell environment; however, this system has some limitations. The main finding of our work is that the addition of plant extract to XVM1 medium results in a gene expression profile that better matches the in planta profile, when compared with the XVM1 medium alone, giving support to the use of plant extract to study pathogenicity mechanisms in Xanthomonas.
Assuntos
Doenças das Plantas/microbiologia , Patologia Vegetal , Virulência/genética , Xanthomonas campestris/genética , Xanthomonas campestris/patogenicidade , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Brassica/microbiologia , Meios de Cultura , Perfilação da Expressão Gênica , Regulação Bacteriana da Expressão Gênica/genética , RNA Bacteriano/análise , RNA Bacteriano/genética , RNA Bacteriano/metabolismo , RNA Mensageiro/análise , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Xanthomonas campestris/metabolismoRESUMO
Xanthomonas campestris pv. campestris (Xcc) is the causal agent of black rot, a highly destructive disease that affects all brassicas. This work aimed to study the interaction Xcc-Brassica oleracea using an in vivo system in an attempt to identify proteins involved in pathogenicity. We used label-free shotgun 2D-nanoUPLC/MSE to analyze Xcc proteins in three conditions: in the interaction with susceptible (REK) and resistant (REU) plants and in culture medium (control condition). A model of Xcc-susceptible host interaction is proposed and shows that Xcc increases the abundance of several crucial proteins for infection and cell protection. In this study, we also confirmed the differential expression by qPCR analysis of selected genes. This is the first report showing a large-scale identification of proteins in an in vivo host plant condition. Considering that most studies involving phytopathogens are in vitro (growth in culture medium or in plant extract), this work contributes with relevant information related to the plant-pathogen interaction in planta.
Assuntos
Proteínas de Bactérias/metabolismo , Brassica/metabolismo , Brassica/microbiologia , Fatores de Virulência/metabolismo , Xanthomonas campestris/patogenicidade , Proteínas de Bactérias/genética , Brassica/crescimento & desenvolvimento , Interações Hospedeiro-Patógeno , Doenças das Plantas/microbiologia , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/metabolismo , Folhas de Planta/microbiologia , Proteoma/metabolismo , Fatores de Virulência/genéticaRESUMO
Transgenic expression of the pepper Bs2 gene confers resistance to Xanthomonas campestris pv. vesicatoria (Xcv) pathogenic strains which contain the avrBs2 avirulence gene in susceptible pepper and tomato varieties. The avrBs2 gene is highly conserved among members of the Xanthomonas genus, and the avrBs2 of Xcv shares 96% homology with the avrBs2 of Xanthomonas citri subsp. citri (Xcc), the causal agent of citrus canker disease. A previous study showed that the transient expression of pepper Bs2 in lemon leaves reduced canker formation and induced plant defence mechanisms. In this work, the effect of the stable expression of Bs2 gene on citrus canker resistance was evaluated in transgenic plants of Citrus sinensis cv. Pineapple. Interestingly, Agrobacterium-mediated transformation of epicotyls was unsuccessful when a constitutive promoter (2× CaMV 35S) was used in the plasmid construction, but seven transgenic lines were obtained with a genetic construction harbouring Bs2 under the control of a pathogen-inducible promoter, from glutathione S-transferase gene from potato. A reduction of disease symptoms of up to 70% was observed in transgenic lines expressing Bs2 with respect to non-transformed control plants. This reduction was directly dependent on the Xcc avrBs2 gene since no effect was observed when a mutant strain of Xcc with a disruption in avrBs2 gene was used for inoculations. Additionally, a canker symptom reduction was correlated with levels of the Bs2 expression in transgenic plants, as assessed by real-time qPCR, and accompanied by the production of reactive oxygen species. These results indicate that the pepper Bs2 resistance gene is also functional in a family other than the Solanaceae, and could be considered for canker control.
Assuntos
Capsicum/genética , Citrus sinensis/genética , Citrus sinensis/microbiologia , Doenças das Plantas/microbiologia , Xanthomonas campestris/patogenicidade , Agrobacterium tumefaciens/genética , Resistência à Doença/genética , Regulação da Expressão Gênica de Plantas , Doenças das Plantas/genética , Brotos de Planta/genética , Plantas Geneticamente Modificadas , Regiões Promotoras Genéticas , Transformação GenéticaRESUMO
Xanthan, the main exopolysaccharide (EPS) synthesized by Xanthomonas spp., contributes to bacterial stress tolerance and enhances attachment to plant surfaces by helping in biofilm formation. Therefore, xanthan is essential for successful colonization and growth in planta and has also been proposed to be involved in the promotion of pathogenesis by calcium ion chelation and, hence, in the suppression of the plant defense responses in which this cation acts as a signal. The aim of this work was to study the relationship between xanthan structure and its role as a virulence factor. We analyzed four Xanthomonas campestris pv. campestris mutants that synthesize structural variants of xanthan. We found that the lack of acetyl groups that decorate the internal mannose residues, ketal-pyruvate groups, and external mannose residues affects bacterial adhesion and biofilm architecture. In addition, the mutants that synthesized EPS without pyruvilation or without the external mannose residues did not develop disease symptoms in Arabidopsis thaliana. We also observed that the presence of the external mannose residues and, hence, pyruvilation is required for xanthan to suppress callose deposition as well as to interfere with stomatal defense. In conclusion, pyruvilation of xanthan seems to be essential for Xanthomonas campestris pv. campestris virulence.
Assuntos
Arabidopsis/microbiologia , Biofilmes/crescimento & desenvolvimento , Glucanos/metabolismo , Doenças das Plantas/microbiologia , Polissacarídeos Bacterianos/química , Xanthomonas campestris/patogenicidade , Interações Hospedeiro-Patógeno , Mutação , Folhas de Planta/microbiologia , Estômatos de Plantas/microbiologia , Polissacarídeos Bacterianos/genética , Polissacarídeos Bacterianos/metabolismo , Ácido Pirúvico/química , Virulência , Fatores de Virulência/química , Fatores de Virulência/genética , Fatores de Virulência/metabolismo , Xanthomonas campestris/genética , Xanthomonas campestris/crescimento & desenvolvimento , Xanthomonas campestris/fisiologiaRESUMO
UNLABELLED: Cruciferous plants are important edible vegetables widely consumed around the world, including cabbage, cauli-flower and broccoli. The main disease that affects crucifer plants is black rot, caused by Xanthomonas campestris pv. campestris (Xcc). In order to better understand this specific plant-pathogen interaction, proteins responsive to Xcc infection in resistant (União) and susceptible (Kenzan) Brassica oleracea cultivars were investigated by 2-DE followed by mass spectrometry. A total of 47 variable spots were identified and revealed that in the susceptible interaction there is a clear reduction in the abundance of proteins involved in energetic metabolism and defense. It was interesting to observe that in the resistant interaction, these proteins showed an opposite behavior. Based on our results, we conclude that resistance is correlated with the ability of the plant to keep sufficient photosynthesis metabolism activity to provide energy supplies necessary for an active defense. As a follow-up study, qRT-PCR analysis of selected genes was performed and revealed that most genes showed an up-regulation trend from 5 to 15days after inoculation (DAI), showing highest transcript levels at 15DAI. These results revealed the gradual accumulation of transcripts providing a more detailed view of the changes occurring during different stages of the plant-pathogen interaction. BIOLOGICAL SIGNIFICANCE: In this study we have compared cultivars of Brassica oleracea (cabbage), susceptible and resistant to black rot, by using the classical 2-DE approach. We have found that resistance is correlated with the ability of the plant to keep sufficient photosynthesis metabolism activity to provide energy supplies necessary for an active defense.
Assuntos
Brassica/microbiologia , Interações Hospedeiro-Patógeno/imunologia , Xanthomonas campestris/fisiologia , Brassica/química , Brassica/imunologia , Brassica/metabolismo , Eletroforese em Gel Bidimensional , Metabolismo Energético , Espectrometria de Massas , Fotossíntese , Proteômica/métodos , Regulação para Cima , Xanthomonas campestris/patogenicidadeRESUMO
The antimicrobial activity of samples of Northern Argentine propolis (Tucumán, Santiago del Estero and Chaco) against phytopathogenic bacteria was assessed and the most active samples were identified. Minimal inhibitory concentration (MIC) values were determined by agar macrodilution and broth microdilution assays. Strong antibacterial activity was detected against Erwinia carotovora spp carotovora CECT 225, Pseudomonas syringae pvar tomato CECT 126, Pseudomonas corrugata CECT 124 and Xanthomonas campestris pvar vesicatoria CECT 792. The most active propolis extract (Tucumán, T1) was selected to bioguide isolation and identified for antimicrobial compound (2',4'-dihydroxychalcone). The antibacterial chalcone was more active than the propolis ethanolic extract (MIC values of 0.5-1 µg ml(-1) and 9.5-15 µg ml(-1), respectively). Phytotoxicity assays were realized and the propolis extracts did not retard germination of lettuce seeds or the growth of onion roots. Propolis solutions applied as sprays on tomato fruits infected with P. syringae reduced the severity of disease. Application of the Argentine propolis extracts diluted with water may be promising for the management of post harvest diseases of fruits.