Sérgio Alves de Carvalho

Expression Profiling of Virulence and Pathogenicity Genes of Xanthomonas axonopodis pv. citri

DNA macroarrays of 279 genes of Xanthomonas axonopodis pv. citri potentially associated with pathogenicity and virulence were used to compare the transcriptional alterations of this bacterium in response to two synthetic media. Data analysis indicated that 31 genes were up-regulated by synthetic medium XVM2, while only 7 genes were repressed. The results suggest that XVM2 could be used as an in vitro system to identify candidate genes involved in pathogenesis of X. axonopodis pv. citri.

Uso de subprodutos de carvão vegetal na formação do porta-enxerto limoeiro 'Cravo' em ambiente protegido

The effect of finely grounded charcoal and pyroligneous acid (PA) in mixture with a commercial growing media and PA sprayed on leaves on growth of Rangpur lime (Citrus limonia Osbeck) seedlings under screen house was studied. Treatments consisted in a combination of 3 levels of charcoal (0, 100 and 200 cm3 dm-3) mixture on the commercial growing media, 2 rates of PA mixtured (0 and 20 cm3 dm-3) to a growing media (240 cm3 per dm3 of growing media), and 3 rates of PA (0, 5 and 10 cm3 dm-3) sprayed on the leaves, with 4 replications. The experiment was carried out in a randomized complete block design, and analyzed as a factorial 3 x 2 x 3. Growth of seedlings was evaluated at 150 and 180 days after planting. The presence of charcoal at 100 cm3 dm-3 in the media did not influence on growth of seedlings. However, the mixture with 200 cm3 dm-3 of charcoal reduced production of total dry mass and high of the plants. The addition of PA (solution 20 cm3 dm-3) to the media reduced plant growth, and sprayed (solutions 5 and 10 cm3 dm-3) on the leaves reduced seedlings diameter.

Chemical Characterization of Citrus sinensis Grafted on C. limonia and the Effect of Some Isolated Compounds on the Growth of Xylella fastidiosa

Citrus sinensis grafted on C. limonia produces a considerable number of compounds that are not common in both plants developed from germination of seeds. The chemical profile of scion and rootstock differ notably for absence in the form of flavonoids and coumarins containing C5 prenyl groups attached to the carbon atoms of aromatic and heterocyclic systems or to oxygen. Only linear pyranocoumarins xanthyletin and xanthoxyletin were found in scion. This observation indicates that the prenylated compounds once biosynthesized in the roots could have been translocated to other organs. Xylella fastidiosa colonizes the xylem of plants causing diseases on several economically important crops such as citrus variegated chlorosis (CVC). A number of flavonoids, coumarins, alkaloids, dihydrocinnamic acid derivative, anacardic acid, triterpenes, and limonoids were tested for in vitro activity on the growth of Xylella fastidiosa. Azadirachtin A was the most active. Hesperidin, which occurs in great amounts in cells of the mesophyll of the affected leaves with CVC, showed a moderate activity suggesting that it can act as a good barrier for small-size colonies from X. fastidiosa.

Quantification and localization of hesperidin and rutin in Citrus sinensis grafted on C. limonia after Xylella fastidiosa infection by HPLC-UV and MALDI imaging mass spectrometry.

A high performance liquid chromatography-ultraviolet (HPLC-UV) method was developed for quantifying hesperidin and rutin levels in leaves and stems of Citrus limonia, with a good linearity over a range of 1.0-80.0 and 1.0-50.0 μg mL(-1) respectively, with r(2)>0.999 for all curves. The limits of detection (LOD) for both flavonoids were 0.6 and 0.5 μg mL(-1), respectively, with quantification (LOQ) being 2.0 and 1.0 μg mL(-1), respectively. The quantification method was applied to Citrus sinensis grafted onto C. limonia with and without CVC (citrus variegated chlorosis) symptoms after Xylella fastidiosa infection. The total content of rutin was low and practically constant in all analyses in comparison with hesperidin, which showed a significant increase in its amount in symptomatic leaves. Scanning electron microscopy studies on leaves with CVC symptoms showed vessel occlusion by biofilm, and a crystallized material was noted. Considering the difficulty in isolating these crystals for analysis, tissue sections were analyzed by matrix-assisted laser desorption/ionization mass spectrometry imaging (MALDI MSI) to confirm the presence of hesperidin at the site of infection. The images constructed from MS/MS data with a specific diagnostic fragment ion (m/z 483) also showed higher ion intensities for it in infected plants than in healthy ones, mainly in the vessel regions. These data suggest that hesperidin plays a role in the plant-pathogen interaction, probably as a phytoanticipin. This method was also applied to C. sinensis and C. limonia seedlings, and comparison with the graft results showed that the rootstock had an increased hesperidin content ∼3.6 fold greater in the graft stem than in the stem of C. sinensis seedlings. Increase in hesperidin content by rootstock can be related to induced internal defense mechanisms.

Development and Validation of Standard Area Diagrams as Assessment Aids for Estimating the Severity of Citrus Canker on Unripe Oranges

Asiatic citrus canker (ACC) is an important disease of citrus in Brazil and elsewhere in the world. Infection with the causal pathogen, Xanthomonas citri subsp. citri, can cause severe disease on the fruit. Visual estimation of severity is the usual method used to quantify ACC on diseased fruit. The objective of this research was to construct and validate standard area diagram (SAD) sets as assessment aids for raters to improve the accuracy and reliability of visual estimates of ACC on unripe (green) fruit of sweet orange. Two SAD sets were constructed. A five-diagram SAD set had five severities depicted (0.5, 2.0, 8.0, 27.0, and 40.0%) and a six-diagram SAD set had six severities depicted (0.5, 1.0, 3.0, 9.0, 20.0, and 40.0%). Fifteen raters evaluated 40 images of cankered, unripe fruit. Both the five- and six-diagram SAD sets significantly improved the accuracy and reliability of estimates. Agreement, measured by Lins concordance correlation coefficient, was 0.220 to 0.913 when not using SADs, 0.814 to 0.955 when using five-diagram SAD sets, and 0.863 to 0.925 when using six-diagram SAD sets. The five-diagram SAD set was significantly more accurate and reliable compared with the six-diagram set. Possible reasons for this are discussed. Based on the results, the five-diagram SAD set is preferable to use. Although the SAD set was developed for sweet orange, it doubtless has applicability to other citrus, including grapefruit. These SAD sets should be useful for research endeavors where accurate and reliable estimates of the severity of ACC are required.

Viroides em citros

Os viroides sao os menores fitopatogenos conhecidos. Constituidos por uma molecula de RNA de fita simples, circular, que nao e encapsidada e nao codifica proteinas, sao capazes de se replicar de maneira autonoma nas celulas do hospedeiro. Os viroides de citros pertencem a familia Pospiviroidae (cujos membros apresentam uma regiao central conservada, replicam-se no nucleo das celulas hospedeiras e nao apresentam atividade ribozimatica) com cinco especies: Citrus exocortis viroid, CEVd (Pospiviroid), Hop stunt viroid, HSVd (Hostuviroid), Citrus bark cracking viroid, CBCVd (Cocadviroid) e Citrus bent leaf viroid, CBLVd e Citrus dwarfing viroid, CDVd (Apscaviroid). Alem disso, Citrus viroid original source (CVd-OS) e mais recentemente, Citrus viroid V (CVd-V) foram propostas como especies tentativas do genero Apscaviroid. Os viroides de citros sao transmitidos via enxertia e disseminados principalmente pela propagacao de material contaminado. Sabe-se que os viroides de citros infectam praticamente todas as especies do genero Citrus e afins. Porem, ha somente duas doencas importantes descritas em citros, induzidas por viroides: (i) a exocorte; (ii) e a xiloporose (ou cachexia). Apesar dos viroides induzirem sintomas severos, ou simplesmente afetarem o tamanho das arvores, muitas especies de citros sao assintomaticas, sendo o controle baseado em medidas preventivas, como utilizacao de gemas livres de viroides aliadas a metodos confiaveis de indexacao. A proposta desta revisao e apresentar ao leitor os recentes avancos nas pesquisas com viroides de citros, principalmente na taxonomia, distribuicao geografica, metodos de deteccao, limpeza e indexacao, epidemiologia e controle, alem do historico e importância desses patogenos para a citricultura mundial.

Disponibilidade de boro em substrato para produção de porta-enxertos de citros em fase de sementeira

ABSTRACTBORON AVAILABILITY IN THE GROWTH MEDIA FOR PRODUCTION OF CITRUSROOTSTOCK SEEDLINGS The production of citrus nursery trees under protected environment using growth media requireshealthy management practices. Problems associated with deficiency and/or nutrient excess have frequentlycaused economic losses in the nursery. Citrus grow differently depending on boron (B) availability inthe growth media, even tough B toxicity commonly cause damage to plants. This research aimed toevaluate the effects of B supply in the growth media on nutrient uptake and seedling growth of tworootstock citrus varieties. A factorial design was set up with two factors: two rootstocks (Rangpur limeand Swingle citrumelo) and four B levels added to the substrate (0, 3, 6, e 12 g m -3 B) as boric acid, with3 replications. Boron availability in the substrate was evaluated at beginning the experiment, 55 and 110days after rootstock seeding, as well as seedling growth and B uptake by seedlings in the same timeintervals. Boron concentration in the treated substrate varied from 0.1 to 4.7 mg L

Produtividade de laranjeira Folha Murcha enxertada em limoeiro Cravo sob adensamento de plantio

) para cada espacamento proposto. No espacamento mais adensado (6,5×2,0 m), as plantas apresentaram menor volume de copa; em contrapartida, maior produtividade da laranjeira Folha Murcha foi observada nesse tratamento. Houve queda de producao nos anos mais avancados de cultivo, independentemente da densidade de plantio. O espacamento 6,5×2,0 acarreta alta produtividade para laranjeira Folha Murcha, tornando a tecnica do adensamento apta a adocao pelo citricultor. Palavras-chave: Citrus sinensis, Citrus limonia, densidade de plantio, sistema de producao, produtividade. Yield of Folha Murcha sweet orange grafted on Rangpur lime under high density planting Abstract The Brazilian citrus production comes through changes in technological standard, specially the high density of plantations. Within this context, was implemented in 1982, the experiment for evaluation of five plant spacings (2.0, 3.0, 4.0, 5.0 and 6.0 meters) to Folha Murcha sweet orange grafted on Rangpur. The test was conducted without irrigation, settling the line spacing of 6.5 meters. Complementing the data obtained in the years of deployment and stabilization of the orchard (1982 to 1993), are presented and discussed in this paper results of assessments carried between the 12 th and 22 nd years of cultivation (1994-2004). It was calculated canopy volume from the data of height and diameter of the plants in 1995. In addition, evaluated the production of fruits per plant and yield (t ha -1 ) for each spacing proposed. In the high density (6.5×2.0 m) the plants had lower canopy volume, on the other hand increased yield of the Folha Murcha sweet orange was observed in this treatment. There was production fall, in the final years of cultivation, regardless of plant density. The 6,5×2,0 m spacing leads to high productivity of Folha Murcha sweet Orange, making high density an apt technique to the adoption by the citrus producer.