RESUMEN
Xylella fastidiosa subsp. pauca, once confined to South America and infecting mainly citrus and coffee plants, has been found to be associated with other hosts and in other geographic regions. We present high-quality draft genome sequences of X. fastidiosa subsp. pauca strains J1a12, B111, U24D, and XRB isolated from citrus plants in Brazil, strain Fb7 isolated from a citrus plant in Argentina and strains 3124, Pr8x, and Hib4 isolated, respectively, from coffee, plum, and hibiscus plants in Brazil. Sequencing was performed using Roche 454-GS FLX, MiSeq-Illumina or Pacific Biosciences platforms. These high-quality genome assemblies will be useful for further studies about the genomic diversity, evolution, and biology of X. fastidiosa.
Asunto(s)
Citrus , Hibiscus , Prunus domestica , Xylella , Argentina , Brasil , Café , Enfermedades de las Plantas , Xylella/genéticaRESUMEN
Xylella fastidiosa subsp. pauca , once confined to South America and infecting mainly citrus and coffee plants, has been found to be associated with other hosts and in other geographic regions. We present high-quality draft genome sequences of X. fastidiosa subsp. pauca strains J1a12, B111, U24D and XRB isolated from citrus plants in Brazil, strain Fb7 isolated from a citrus plant in Argentina and strains 3124, Pr8x and Hib4 isolated, respectively, from coffee, plum and hibiscus plants in Brazil. Sequencing was performed using Roche 454-GS FLX, MiSeq-Illumina or Pacific Biosciences platforms. These high-quality genome assemblies will be useful for further studies about the genomic diversity, evolution, and biology of X. fastidiosa.
RESUMEN
Plant acclimation to recurrent stress involves profound alterations in multiple genetic, metabolic and physiological processes. Stressful conditions usually implicate imbalance in reactive oxygen species (ROS) production and removal rates, which may lead to oxidative stress. However, the primary cellular targets of oxidative stress and their relevance in plant acclimation to abiotic stresses remains poorly characterized. By comparing redox proteomic and sugar profiles in citrus Valencia (VO) scions grafted onto two rootstocks with different soil water extraction capacities - Rangpur Lime (RL) and Sunki Maravilha (SM) - here we demonstrate that both ROS-mediated post-translational protein modification and changes in sugar composition are associated with acclimation to recurrent drought in citrus. The redox proteomic analysis of the distinct scion/rootstock combinations exposed to one (WD1), two (WD2) or three (WD3) water deficit episodes revealed a total of 32 and 55 redox protein spots present in VO/RL and VO/SM plants, respectively. Mass spectrometry analysis of these protein spots revealed essential targets of ROS-mediated posttranslational protein modification in citrus plants challenged by recurrent drought. The oxidation of cysteine thiol groups into glyceraldehyde 3-phosphate dehydrogenase (GAPDH) was shown to increase in WD3 samples of the VO/RL combination, whereas the opposite was observed for the VO/SM combination. Similarly, recurrent drought promoted the oxidation of catalase thiol groups in VO/SM, but not in VO/RL. Carbohydrate profiling revealed that glucose, fructose and galactose may also contribute to the phenotypic differences observed between the citrus genotypes exposed to drought. These findings reveal for the first time that recurrent drought differentially affects the profile of redox proteomics of citrus, suggesting that this alteration may be part of the stress memory in perennial plants.
Asunto(s)
Citrus/metabolismo , Citrus/fisiología , Sequías , Proteínas de Plantas/metabolismo , Agua , Carbohidratos/química , Electroforesis en Gel Bidimensional , Oxidación-Reducción , Fenotipo , Hojas de la Planta/metabolismo , Análisis de Componente Principal , Coloración y EtiquetadoRESUMEN
Rootstocks influence certain characteristics of the scion, such as vigor, fruit yield and quality, and tolerance to biotic and abiotic factors. In conventional citrus breeding programs, hybridization is the main method used for obtaining rootstock varieties. Between Sept and Oct 2006, controlled pollinations were carried out resulting in nine progenies, totaling 461 hybrid seedlings. The female parents were the Santa Cruz Rangpur lime (SCRL) and the Sunki of Florida mandarin (SFM), while the male parents, in addition to this mandarin, were the Indio and Riverside citrandarins, the Thomasville citrangequat (THOM), the Swingle citrumelo, the Argentina citrange (AGC) and the Benecke trifoliate orange (BKTO). Fruit setting rates of these crosses were measured and the hybrids generated were planted in the field in 2008 and evaluated from 2010 to 2014, for the variables of plant mortality, visual vigor, plant height and trunk diameter. The descriptive statistics mean, variation range, coefficient of variation and Pearson skewness coefficient were calculated. Data were submitted to analysis of variance and the means of the treatments were grouped by the Scott-Knott test, both at 5 % significance. The progenies, having the Sunki mandarin as the female parent, presented higher fruit setting rates and lower plant mortality rates than those obtained from the female parent Rangpur lime, while zero plant mortality was observed in the SFM × BKTO cross. The crosses SCRL × SFM, SFM × BKTO, SFM × AGC and SFM × THOM have the potential to generate vigorous hybrids, resulting in rootstock varieties adapted to the landscape unit of Coastal Tableland.(AU)
Asunto(s)
Citrus , Poncirus , Hibridación Genética , Fitomejoramiento/métodosRESUMEN
Rootstocks influence certain characteristics of the scion, such as vigor, fruit yield and quality, and tolerance to biotic and abiotic factors. In conventional citrus breeding programs, hybridization is the main method used for obtaining rootstock varieties. Between Sept and Oct 2006, controlled pollinations were carried out resulting in nine progenies, totaling 461 hybrid seedlings. The female parents were the Santa Cruz Rangpur lime (SCRL) and the Sunki of Florida mandarin (SFM), while the male parents, in addition to this mandarin, were the Indio and Riverside citrandarins, the Thomasville citrangequat (THOM), the Swingle citrumelo, the Argentina citrange (AGC) and the Benecke trifoliate orange (BKTO). Fruit setting rates of these crosses were measured and the hybrids generated were planted in the field in 2008 and evaluated from 2010 to 2014, for the variables of plant mortality, visual vigor, plant height and trunk diameter. The descriptive statistics mean, variation range, coefficient of variation and Pearson skewness coefficient were calculated. Data were submitted to analysis of variance and the means of the treatments were grouped by the Scott-Knott test, both at 5 % significance. The progenies, having the Sunki mandarin as the female parent, presented higher fruit setting rates and lower plant mortality rates than those obtained from the female parent Rangpur lime, while zero plant mortality was observed in the SFM × BKTO cross. The crosses SCRL × SFM, SFM × BKTO, SFM × AGC and SFM × THOM have the potential to generate vigorous hybrids, resulting in rootstock varieties adapted to the landscape unit of Coastal Tableland.
Asunto(s)
Citrus , Hibridación Genética , Fitomejoramiento/métodos , PoncirusRESUMEN
The present study evaluated the physiological, molecular and hormonal parameters from scion/rootstock interaction of citrus plants during recurrent water deficit. Responses of the Valencia (VO) scion variety grafted on two rootstocks with different soil water extraction capacities, Rangpur Lime (RL) and Sunki Maravilha (SM), during three successive periods of water deficit: plants exposed to a single episode of water deficit (WD1) and plants exposed to two (WD2) and three (WD3) recurrent periods of WD were compared. The combinations VO/RL and VO/SM presented polymorphic alterations of epigenetic marks and hormonal (i.e. abscisic acid, auxins and salicylicacid) profiles, which were particularly prominent when VO/SM plantswere exposed toWD3 treatment. Upon successive drought events, the VO/SM combination presented acclimatization characteristics that enable higher tolerance to water deficit by increasing transpiration (E), stomatal conductance (g s ) and photosynthetic rate (A), which in turn may have facilitated the whole plant survival. Besides providing comprehensive data on the scion/rootstock interactions upon successive stress events, this study brings the first dataset suggesting that epigenetic alterations in citrus plants triggered by recurrent water deficit lead to improved drought tolerance in this crop species.
Asunto(s)
Citrus/metabolismo , Sequías , Epigénesis Genética , Reguladores del Crecimiento de las Plantas/metabolismo , Estrés Fisiológico/fisiología , Antioxidantes/metabolismo , Citrus/genética , Metilación de ADN , Deshidratación/genética , Deshidratación/metabolismo , Regulación de la Expresión Génica de las Plantas , Fotosíntesis/genética , Fotosíntesis/fisiología , Hojas de la Planta/metabolismo , Proteínas de Plantas/metabolismo , Raíces de Plantas/genética , Raíces de Plantas/metabolismo , Estomas de Plantas/metabolismo , Transpiración de Plantas/genética , Transpiración de Plantas/fisiología , Distribución Aleatoria , Estrés Fisiológico/genética , Agua/metabolismoRESUMEN
Water scarcity can elicit drastic changes in plant metabolic and hormonal regulation, which may be of fundamental importance to stress tolerance. The study of plant the metabolic alterations in response to water deficit, especially the effects of the rootstocks level, is important to elucidate the mechanisms associated to drought tolerance. To verify the influence of rootstock and grafting on the tolerance to drought in citrus plants, we analyzed the growth, phytohormone levels and flavonoid profiles in grafted and ungrafted citrus plants subjected to different soil water regimes on plant status (well-watered, moderate drought and severe drought and rehydrated) under field conditions. The experiments were conducted under field conditions in the Brazilian Agricultural Research Corporation (EMBRAPA), Cruz das Almas, BA, Brazil. Water deficit reduced the total leaf area per plant in all canopy/rootstock combinations. Self-grafting reduce root volume, area and length when compared to ungrafted plants. Drought-induced increases in salicylic acid and abscisic acid associated with concomitant reductions in indoleacetic acid were observed in most canopy/rootstock combinations. However, plants with 'Sunki Maravilha' rootstocks exhibited the most pronounced changes in hormonal levels upon drought stress. Associated to these hormonal changes, drought also significantly affected flavonoid content and profile in both leaves and roots of the distinct citrus combinations. Glycosylated (GFs) and polimethoxylated flavonoids were predominantly found in leaves, whereas prenylated coumarins were found in the roots. Leaf levels of GFs (vicenin, F11, rutin and rhoifolin) were particularly modulated by drought in plants with 'Rangpur Santa Cruz' lime rootstock, whereas root levels of prenylated coumarins were most regulated by drought in plants with the 'Sunki Maravilha' root system. Taken together, these data indicate that the impacts of water deficit restriction on growth, hormonal balance and flavonoid profiles significantly varies depending on the canopy/rootstock combinations.
Asunto(s)
Citrus/crecimiento & desarrollo , Flavonoides/metabolismo , Reguladores del Crecimiento de las Plantas/metabolismo , Raíces de Plantas/crecimiento & desarrollo , Deshidratación/metabolismoRESUMEN
Two citrus rootstocks, Rangpur lime (RL) and Sunki Maravilha mandarin (SM), were analyzed either ungrafted or grafted with their reciprocal graft combinations or with shoot scions of two commercial citrus varieties: Valencia orange (VO) and Tahiti acid lime (TAL). All graft combinations were subjected to distinct watering regimes: well-watered, severe drought and rehydration. Growth and water relation parameters, gas exchange as well as sugar and hormone profiles were determined. Data indicated that RL adopted a dehydration avoidance strategy and maintained growth, whereas SM adopted a dehydration tolerance strategy focused on plant survival. Compared with RL, the leaves and roots of SM exhibited higher concentrations of abscisic acid and salicylic acid, which induced drought tolerance, and accumulation of carbohydrates such as trehalose and raffinose, which are important reactive oxygen species scavengers. SM rootstocks were able to transfer their survival strategy to the grafted shoot scions (RL, VO, TAL). Because of their contrasting survival strategies, RL reached the permanent wilting point more quickly than SM whereas SM recovered from prolonged droughts more efficiently than RL. This is one of the most complete studies of drought tolerance mechanisms in citrus crops and is the first to use reciprocal grafting to clarify scion/rootstock interactions.
Asunto(s)
Citrus/metabolismo , Raíces de Plantas/metabolismo , Brotes de la Planta/metabolismo , Estrés Fisiológico , DeshidrataciónRESUMEN
Persian lime (PL) [Citrus latifolia (Yu. Tanaka) Tanaka] is an important species both for domestic fresh fruit consumption in Brazil as well as the export market, since the country is one of the largest producers in the world despite the fact that, in commercial plantations, it is still not uncommon to find trees with low productivity and high plant vigor of unknown origin. Selections of Persian lime CNPMF2000, CNPMF2001, CNPMF01, CNPMF02, IAC5, IAC5.1, Bearss, Persian58, and 5059, were therefore grafted onto Swingle citrumelo [C. paradisi Macfad. cv. Duncan × Poncirus trifoliata (L.) Raf.] rootstocks and evaluated in Cruz das Almas, Bahia, Brazil in a field experiment conducted in a completely randomized block design with five replications and two trees per plot. The biometric attributes (canopy height, diameter and volume), yield parameters (yield during the off-season harvest period, yield per plant, production efficiency), and fruit quality traits, were evaluated. The CNPMF2001, CNPMF01, CNPMF02, IAC5, and Bearss selections had 5-11 % shorter trees than the other cultivars. CNPMF01, CNPMF02, Persian58, and 5059 presented higher yield efficiency values, between 3.1-3.4 kg m3, and higher yield levels during the off-season harvest periods. The Bearss, Persian58, CNPMF2000, IAC5.1 and 5059 selections had more acid fruits and the latter three, smaller fruits. Based on their horticultural performance up to eight years of age, CNPMF01, CNPMF02, Persian58 and 5059 selections were the most promising varieties of Persian lime.
Asunto(s)
Citrus/anatomía & histología , Citrus/crecimiento & desarrolloRESUMEN
Persian lime (PL) [Citrus latifolia (Yu. Tanaka) Tanaka] is an important species both for domestic fresh fruit consumption in Brazil as well as the export market, since the country is one of the largest producers in the world despite the fact that, in commercial plantations, it is still not uncommon to find trees with low productivity and high plant vigor of unknown origin. Selections of Persian lime CNPMF2000, CNPMF2001, CNPMF01, CNPMF02, IAC5, IAC5.1, Bearss, Persian58, and 5059, were therefore grafted onto Swingle citrumelo [C. paradisi Macfad. cv. Duncan × Poncirus trifoliata (L.) Raf.] rootstocks and evaluated in Cruz das Almas, Bahia, Brazil in a field experiment conducted in a completely randomized block design with five replications and two trees per plot. The biometric attributes (canopy height, diameter and volume), yield parameters (yield during the off-season harvest period, yield per plant, production efficiency), and fruit quality traits, were evaluated. The CNPMF2001, CNPMF01, CNPMF02, IAC5, and Bearss selections had 5-11 % shorter trees than the other cultivars. CNPMF01, CNPMF02, Persian58, and 5059 presented higher yield efficiency values, between 3.1-3.4 kg m3, and higher yield levels during the off-season harvest periods. The Bearss, Persian58, CNPMF2000, IAC5.1 and 5059 selections had more acid fruits and the latter three, smaller fruits. Based on their horticultural performance up to eight years of age, CNPMF01, CNPMF02, Persian58 and 5059 selections were the most promising varieties of Persian lime.(AU)
Asunto(s)
Citrus/crecimiento & desarrollo , Citrus/anatomía & histologíaRESUMEN
Citrus Tristeza disease, caused by CTV (Citrus tristeza virus), committs citrus plantations around the world and specifically attacks phloem tissues of the plant. The virus exists as a mixture of more or less severe variants, which may or may not cause symptoms of Tristeza. The objective of this study was to analyze the changes caused by CTV in the proteome of stems of sweet orange, as well as in the activity and gene expression of antioxidant enzymes. The CTV-infected sweet orange displayed mild symptoms, which were characterized by the presence of sparse stem pitting throughout their stems. The presence of virus was confirmed by RT-PCR. Proteomic analysis by 2DE-PAGE-MS / MS revealed the identity of 40 proteins differentially expressed between CTV- infected and -non-infected samples. Of these, 33 were up-regulated and 7 were down-regulated in CTV-infected samples. Among the proteins identified stands out a specific from the virus, the coat protein. Other proteins identified are involved with oxidative stress and for this their enzymatic activity was measured. The activity of superoxide dismutase (SOD) was higher in CTV-infected samples, as catalase (CAT) showed higher activity in uninfected samples. The activity of guaiacol peroxidase (GPX) did not vary significantly between samples. However, ascorbate peroxidase (APX) was more active in the infected samples. The relative expression of the genes encoding CAT, SOD, APX and GPX was analyzed by quantitative real time PCR (RT-qPCR). The CTV-infected samples showed greater accumulation of transcripts, except for the CAT gene. This gene showed higher expression in the uninfected samples. Taken together, it can be concluded that the CTV affects the protein profile and activity and gene expression of antioxidant enzymes in plants infected by this virus.
Asunto(s)
Citrus sinensis/virología , Closterovirus/fisiología , Enzimas/metabolismo , Proteínas de Plantas/metabolismo , Proteoma/metabolismo , Proteómica/métodos , Ascorbato Peroxidasas/genética , Ascorbato Peroxidasas/metabolismo , Catalasa/genética , Catalasa/metabolismo , Citrus sinensis/genética , Citrus sinensis/metabolismo , Closterovirus/genética , Electroforesis en Gel Bidimensional , Enzimas/genética , Perfilación de la Expresión Génica/métodos , Regulación de la Expresión Génica de las Plantas , Regulación Viral de la Expresión Génica , Interacciones Huésped-Patógeno/genética , Peroxidasa/genética , Peroxidasa/metabolismo , Enfermedades de las Plantas/genética , Enfermedades de las Plantas/virología , Proteínas de Plantas/genética , Proteoma/genética , Reacción en Cadena de la Polimerasa de Transcriptasa Inversa , Superóxido Dismutasa/genética , Superóxido Dismutasa/metabolismo , Espectrometría de Masas en TándemRESUMEN
BACKGROUND: The pathogenesis related protein PR10 (TcPR-10), obtained from the Theobroma cacao-Moniliophthora perniciosa interaction library, presents antifungal activity against M. perniciosa and acts in vitro as a ribonuclease. However, despite its biotechnological potential, the TcPR-10 has the P-loop motif similar to those of some allergenic proteins such as Bet v 1 (Betula verrucosa) and Pru av 1 (Prunus avium). The insertion of mutations in this motif can produce proteins with reduced allergenic power. The objective of the present work was to evaluate the allergenic potential of the wild type and mutant recombinant TcPR-10 using bioinformatics tools and immunological assays. METHODOLOGY/PRINCIPAL FINDINGS: Mutant substitutions (T10P, I30V, H45S) were inserted in the TcPR-10 gene by site-directed mutagenesis, cloned into pET28a and expressed in Escherichia coli BL21(DE3) cells. Changes in molecular surface caused by the mutant substitutions was evaluated by comparative protein modeling using the three-dimensional structure of the major cherry allergen, Pru av 1 as a template. The immunological assays were carried out in 8-12 week old female BALB/c mice. The mice were sensitized with the proteins (wild type and mutants) via subcutaneous and challenged intranasal for induction of allergic airway inflammation. CONCLUSIONS/SIGNIFICANCE: We showed that the wild TcPR-10 protein has allergenic potential, whereas the insertion of mutations produced proteins with reduced capacity of IgE production and cellular infiltration in the lungs. On the other hand, in vitro assays show that the TcPR-10 mutants still present antifungal and ribonuclease activity against M. perniciosa RNA. In conclusion, the mutant proteins present less allergenic potential than the wild TcPR-10, without the loss of interesting biotechnological properties.
Asunto(s)
Alérgenos/inmunología , Antígenos de Plantas/inmunología , Cacao , Proteínas de Plantas/inmunología , Algoritmos , Alérgenos/química , Secuencia de Aminoácidos , Animales , Antifúngicos/farmacología , Antígenos de Plantas/química , Basidiomycota/citología , Basidiomycota/efectos de los fármacos , Lavado Broncoalveolar , Cacao/química , Cacao/inmunología , Recuento de Células , Biología Computacional , Bases de Datos de Proteínas , Femenino , Interacciones Hidrofóbicas e Hidrofílicas/efectos de los fármacos , Inmunoglobulina E/inmunología , Pulmón/efectos de los fármacos , Pulmón/inmunología , Ratones , Ratones Endogámicos BALB C , Viabilidad Microbiana/efectos de los fármacos , Modelos Moleculares , Datos de Secuencia Molecular , Proteínas Mutantes/química , Proteínas Mutantes/inmunología , Proteínas de Plantas/química , Proteínas de Plantas/farmacología , Ribonucleasas/metabolismo , Alineación de Secuencia , Homología Estructural de ProteínaRESUMEN
This study reports on expression analysis associated with molecular systems biology of cacao-Moniliophthora perniciosa interaction. Gene expression data were obtained for two cacao genotypes (TSH1188, resistant; Catongo, susceptible) challenged or not with the fungus M. perniciosa and collected at three time points through disease. Using expression analysis, we identified 154 and 227 genes that are differentially expressed in TSH1188 and Catongo, respectively. The expression of some of these genes was confirmed by RT-qPCR. Physical protein-protein interaction (PPPI) networks of Arabidopsis thaliana orthologous proteins corresponding to resistant and susceptible interactions were obtained followed by cluster and gene ontology analyses. The integrated analysis of gene expression and systems biology allowed designing a general scheme of major mechanisms associated with witches' broom disease resistance/susceptibility. In this sense, the TSH1188 cultivar shows strong production of ROS and elicitors at the beginning of the interaction with M. perniciosa followed by resistance signal propagation and ROS detoxification. On the other hand, the Catongo genotype displays defense mechanisms that include the synthesis of some defense molecules but without success in regards to elimination of the fungus. This phase is followed by the activation of protein metabolism which is achieved with the production of proteasome associated with autophagy as a precursor mechanism of PCD. This work also identifies candidate genes for further functional studies and for genetic mapping and marker assisted selection.
Asunto(s)
Cacao/genética , Cacao/microbiología , Resistencia a la Enfermedad/genética , Perfilación de la Expresión Génica/métodos , Redes Reguladoras de Genes/genética , Enfermedades de las Plantas/inmunología , Biología de Sistemas/métodos , Basidiomycota/fisiología , Análisis por Conglomerados , Resistencia a la Enfermedad/inmunología , Susceptibilidad a Enfermedades/inmunología , Regulación de la Expresión Génica de las Plantas , Genes de Plantas/genética , Enfermedades de las Plantas/genética , Enfermedades de las Plantas/microbiología , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Mapas de Interacción de Proteínas , ARN Mensajero/genética , ARN Mensajero/metabolismo , Transcriptoma/genéticaRESUMEN
Witches' broom disease, caused by Crinipellis perniciosa, is one of the major fungal diseases causing severe losses to cacao tree (Theobroma cacao L.) plantations in South America. One of the challenges associated with the understanding of the cacao and Crinipellis interaction in genomic studies is the isolation of intact nucleic acids. In this report, we describe a new, successful, and reliable procedure for the isolation of RNA from tissues of cacao tree, both infected and uninfected by Crinipellis. This protocol overcomes the problems associated with the very high amount of polyphenols and polysaccharides present in cacao organs that are not easily removed by conventional extraction procedures. The protocol requires few reagents, uses ultracentrifugation and inexpensive consumables, and can be easily applied in any laboratory. This method produced high-quality RNA that was suitable for subsequent purposes, such as reverse transcription PCR and cDNA library construction. We also report the first evidence of RNA isolation from cacao organs infected by C. perniciosa such as meristems and fruits.
Asunto(s)
Basidiomycota/patogenicidad , Cacao/metabolismo , Cacao/microbiología , Reacción en Cadena de la Polimerasa/métodos , ARN/aislamiento & purificación , Cacao/genética , Frutas/genética , Frutas/metabolismo , Frutas/microbiología , Tallos de la Planta/genética , Tallos de la Planta/metabolismo , Tallos de la Planta/microbiología , Ultracentrifugación/métodosRESUMEN
The genetic reduction of linolenic acid levels increases the quality and stability of soybean oil. The objective of this study was to determine the inheritance and evaluate the nature and magnitude of gene effects on soybean seed linolenic acid level. Means and variances of F-1, F-2, and F-3 generations were made from the cross between accession BARC-12 (low linolenic acid content) and the commercial Brazilian cultivar CAC-1 (normal linolenic acid content). The results demonstrated that linolenic acid content in soybean is under the genetic control of a small number of genes. The additive model explained the means for the three generations and for the parents. Non-allelic gene interactions had little effect on the determination of genotypic values for the individuals. The generation means and population variation analyses demonstrated that the dominance deviations contribute little to the trait. These results showed that backcross breeding programs can be used to introduce the low linolenic acid content trait into soybean seeds, since it is possible to identify with very high accuracy the desired genotypes in segregating populations