Induction of transgenic hairy roots in soybean genotypes by Agrobacterium rhizogenes‑mediated transformation

The objective of this work was to perform the screening of soybean genotypes as to their ability to respond to the induction of hairy roots by Agrobacterium rhizogenes‑mediated transformation. Four Brazilian soybean cultivars (BRSMG 68 Vencedora, BRS 137, Embrapa 48, and MG/BR 46 Conquista) and two North American ones adapted to Brazilian cropping conditions (Bragg and IAS‑5) were screened for their capacity to respond to A. rhizogenes in protocols for in vitro hairy root culture and ex vitro composite plant production. Four‑day‑old seedlings with uniform size were injected with A. rhizogenes harboring the plasmid p35S‑GFP. Seedlings expressing green fluorescent protein (GFP) in at least one hairy root were used to determine the transformation frequency. Using an axenic in vitro protocol, excised cotyledons from four‑day‑old seedlings were infected with A. rhizogenes harboring the pCAMBIA1301 plasmid, containing the gusA reporter gene. The transformation frequency and the number of days for hairy root emergence after bacterial infection (DAI) were evaluated. The transformation frequency and DAI varied according to the genotype. Cultivars MG/BR 46 Conquista and BRSMG 68 Vencedora are more susceptible to A. rhizogenes and can be recommended for transformation experiments.

Obtaining interspecific hybrids, and molecular analysis by microsatellite markers in grapevine

The objective of this work was to assess the potential of interspecific hybridization of Vitis labruscana and Muscadinia rotundifolia by using artificial cross-pollinations. Microsatellite markers were used to confirm interspecific hybridizations and the identity of the parental genotypes. In crosses in which M. rotundifolia was used as the female parent, no true hybrids were obtained. In the reciprocal crosses, 114 seedlings were identified as true V. labruscana x M. rotundifolia hybrids. Self pollination occurred in direct and in reciprocal crosses. The crossings between 'Bordo' x 'Carlos', 'Magnolia', 'Regale' and' Roanoke', and between' Isabel' x 'Bountiful', 'Carlos', 'Magnolia', 'Regale' and 'Roanoke' were confirmed. The 15 markers evaluated showed that two M. rotundifolia parental genotypes had the same fingerprint profile, indicating a like lyplanting error. The success of hybridization depends mainly on the species and on the cultivar used as the female parent. Microsatellite markers are efficient to confirm the paternity of interspecific F1 hybrids and to determine the correct identity of M. rotundifolia cultivars.

Watermelon transformation with Zucchini yellow mosaic virus coat protein gene and comparison with parental cultivar

The objective of this work was to transfer Zucchini yellow mosaic virus coat protein (ZYMV-CP) and neomycin phosphotransferase II (NPT II) genes to the watermelon 'Crimson Sweet'(CS) genome, and to compare the transgenic progenies T1 and T2 with the nontransformed parental cultivar for morphological, pomological, growth and yield characteristics. The ZYMV-CP gene was transferred by Agrobacterium tumefaciens. The presence of the gene in transgenic T0, T1 and T2 plants was determined by polymerase chain reaction, and the results were confirmed by Southern blot. Two experiments were performed, one in the winter-spring and the other in the summer-autumn. In both experiments, the hypocotyl length of transgenic seedlings was significantly higher than that of nontransgenic parental ones. In the second experiment, the differences between transgenic and nontransgenic individuals were significant concerning fruit rind thickness, flesh firmness, fruit peduncle length, size of pistil scar, and a* values for fruit stripe or flesh color. Transferring ZYMV-CP gene to CS genome affected only a few characteristics from the 80 evaluated ones. The changes in rind thickness, flesh firmness and flesh color a* values are favorable, while the increase in the size of pistil scar is undesirable. The transgenic watermelon line having ZYMV-CP gene and the parental cultivar CS are very similar.

Antibacterial activity of essential oils on Xanthomonas vesicatoria and control of bacterial spot in tomato

The objective of this work was to evaluate the effects of plant essential oils (EOs) on the growth of Xanthomonas vesicatoria, on bacterial morphology and ultrastructure, and on the severity of tomato bacterial spot. EOs from citronella, clove, cinnamon, lemongrass, eucalyptus, thyme, and tea tree were evaluated in vitro at concentrations of 0.1, 1.0, 10, and 100% in 1.0% powdered milk. The effect of EOs, at 0.1%, on the severity of tomato bacterial spot was evaluated in tomato seedlings under greenhouse conditions. The effects of citronella, lemongrass, clove, and tea tree EOs, at 0.1%, on X. vesicatoria cells were evaluated by transmission electron microscopy. All EOs showed direct toxic effect on the bacteria at a 10%-concentration in vitro. Under greenhouse conditions, the EOs of clove, citronella, tea tree, and lemongrass reduced disease severity. EOs of clove and tea tree, and streptomycin sulfate promoted loss of electron-dense material and alterations in the cytoplasm, whereas EO of tea tree promoted cytoplasm vacuolation, and those of citronella, lemongrass, clove, and tea tree caused damage to the bacterial cell wall. The EOs at a concentration of 0.1% reduce the severity of the disease.

Dinâmica de nutrientes na solução do solo em pomar fertirrigado de citros

O objetivo deste trabalho foi avaliar a dinâmica de nutrientes na solução do solo após a aplicação, via fertirrigação, de nitrogênio, fósforo e potássio a laranjeiras. O experimento foi realizado entre setembro de 2007 e outubro de 2009, em pomares de laranjeiras 'Valência' e 'Hamlin', enxertadas sobre citrumeleiro 'Swingle'. Foram avaliadas cinco doses de N, P2O5 e K2O (0, 25, 50, 100 e 200% da dose recomendada). A solução do solo foi extraída a 30 e 60 cm de profundidade, com o auxílio de extratores com cápsulas porosas. Foram realizadas 11 avaliações durante o período experimental, com as extrações iniciadas após 12 horas das fertirrigações. O aumento das doses reduziu o pH (pH~3,5, na maior dose), e aumentou a condutividade elétrica (CE~1,5 dS m-1, na maior dose) e os teores de NH4, NO3, P, K, Mn e Zn na solução do solo, nas duas profundidades amostradas. Nos meses com maior precipitação pluvial, houve perda potencial de nutrientes por lixiviação, pois maiores concentrações de NO3, K e B foram observadas à profundidade de 60 cm. A análise da solução do solo, obtida por extratores com cápsula de cerâmica porosa, pode ser considerada ferramenta auxiliar para monitorar e avaliar a disponibilidade de nutrientes às plantas.

Leaf-cutting ant attack in initial pine plantations and growth of defoliated plants

The objective of this work was to evaluate the natural attack by Acromyrmex crassispinus in initial Pinus taeda plantations without control measures against ants, as well as the effect of defoliation in seedlings of P. taeda. Evaluations of the attack of leaf-cutting ants on P. taeda plantations were done monthly in the first six months, then 9 and 12 months after planting. The percentages of plants that were naturally attacked by ants were registered. The effect of defoliation was evaluated by artificial defoliation, simulating the natural patterns of attack by A. crassispinus on P. taeda seedlings. The natural attack of A. crassispinus was greater during the first months after planting, being more intense in the first 30 days. Artificial defoliation indicated that there were no significant losses in diameter and height in plants with less than 75% defoliation. However, there were significant losses in diameter and height in plants with 100% defoliation, independently of the cut of the apical meristem, and also plant death. The control of leaf-cutting ants in P. taeda plantings, in which A. crassispinus is the most frequent leaf-cutting ant, should be intense only at the beginning of planting, since the most severe attacks occur during this time.

Genetic similarity in a hybrid population of 'Montenegrina' and 'King' mandarins

The objective of this work was to confirm the hybrids obtained in plants originated from the crossing between the mandarins Citrus deliciosa 'Montenegrina' and C. nobilis 'King', and to estimate the genetic similarity among hybrids, and between each hybrid and its parents. Twenty‑three pairs of microsatellite primers were tested. Fourteen of these pairs showed polymorphic bands between parents. Primers CCSM 129 and CCSME 52 were sufficient to identify the 12 nucellar clones observed in the studied population. Genetic similarity analysis of the population (hybrids and parents) showed 0.56 average similarity. Besides the 12 clones of 'Montenegrina' identified, 25 hybrids were found of which D18, C32, D06, C05 and D09 are the more similar to 'Montenegrina'.

Seleção de cultivares de laranja doce de maturação precoce por índices de desempenho

O objetivo deste trabalho foi selecionar cultivares de laranja doce de maturação precoce, adequadas para o mercado de frutas in natura e para o processamento industrial, por meio de índices de desempenho. Índices de desempenho para citros foram estabelecidos com base em dados coletados em experimento conduzido na região sudoeste do Estado de São Paulo, envolvendo 12 cultivares de laranja doce de maturação precoce. Resultados pioneiros foram obtidos na identificação de cultivares superiores. Em comparação com a laranja 'Hamlin', cultivar padrão de maturação precoce, identificaram-se as laranjas 'Valência 2' e 'Salustiana' com potencial para o mercado de frutas in natura, e a laranja 'Westin', para o processamento industrial.

Genetic transformation of sweet oranges with the D4E1 gene driven by the AtPP2 promoter

The objective of this work was to produce transgenic 'Pêra' and 'Valência' sweet orange plants using the D4E1 gene driven by the Arabidopsis thaliana phloem protein (AtPP2) promoter and to quantify transgene expression in different transformation events. Genetic transformation experiments were carried out with epicotyl segments co‑cultivated with Agrobacterium tumefaciens. Six plants from 'Pêra' sweet orange and seven plants from 'Valência' sweet orange were confirmed as different transgenic events by means of the polymerase chain reaction (PCR) and the Southern blot techniques. Transgene expression was quantified using real‑time quantitative PCR. D4E1 gene expression levels vary from 5 up to 50 times among different transformation events.

Antecipação do período de diagnose foliar em laranjeira 'Pêra' no Amazonas

O objetivo deste trabalho foi avaliar a possibilidade de se antecipar o período de realização da diagnose foliar em laranjeira 'Pêra'. Vinte e sete pomares representativos da região produtora de laranja do Estado do Amazonas foram monitorados durante o ano agrícola de 2011/2012. Foram realizadas diagnoses da composição nutricional (CND) em amostras foliares retiradas durante a floração e quando a árvore apresentava frutos com três e seis meses de idade (época tradicional para o monitoramento nutricional). Pomares com produtividade superior a 25 Mg ha‑1 foram selecionados para o estabelecimento dos padrões de referência. O estado nutricional da laranja variou com o estádio fenológico no qual se realizou a amostragem foliar, o que fez com que fosse necessário estabelecer normas CND para cada período. Com a antecipação da diagnose para o período de floração, observou-se aumento nas concentrações foliares de N, P, K e Cu diminuição e nas de Ca. A antecipação da diagnose foliar em laranja 'Pêra' depende da geração de padrões nutricionais CND específicos para cada época de amostragem.

Reaction of sweet orange cultivars expressing the attacin A gene to 'Candidatus Liberibacter asiaticus' infection

The objective of this work was to evaluate the reaction of four sweet orange cultivars expressing the attacin A gene to 'Candidatus Liberibacter asiaticus' (Las) infection, a bacterium associated to huanglongbing (HLB) disease. Transgenic sweet orange plants of Hamlin, Natal, Pêra, and Valência cultivars, as well as nontransgenic controls received inocula by grafting budwood sections of HLB-infected branches. Disease progression was evaluated through observations of leaf symptoms and by polymerase chain reaction (PCR) analysis, eight months after inoculation. A completely randomized design was used, with four experiments (one for each cultivar) performed simultaneously. Bacteria title was estimated by quantitative PCR (qPCR). HLB symptoms and Las titers were present in nontransgenic and transgenic plants expressing the attacin A gene of the four sweet orange cultivars, eight months after bacteria inoculation. Five transgenic lines (transformation events) of 'Pêra' sweet orange expressing the attacin A gene have significantly lower Las titers in comparison with nontransgenic plants of this cultivar.

Diversidade genética de Xylella fastidiosa em regiões produtoras de citros na Bahia

O objetivo deste trabalho foi avaliar, por meio de marcadores SSR, a diversidade genética de Xylella fastidiosa no Estado da Bahia. Foram estudadas duas das principais regiões produtoras de citros no Estado, o Litoral Norte e o Recôncavo Sul. Para fins comparativos, utilizaram-se dez amostras provenientes do Estado de São Paulo. Foram empregados os seguintes iniciadores: ASSR20, OSSR9, OSSR17, CSSR4, CSSR12 e CSSR20, dos quais os quatro últimos permitiram identificar 22 loci polimórficos. As populações de X. fastidiosa presentes em citros no Estado da Bahia apresentam elevada diversidade genética, com base nos marcadores SSR, com pools gênicos distintos e agrupamento geográfico. No Litoral Norte, as populações do isolado apresentam maior diversidade genética do que as da região do Recôncavo Sul da Bahia.

Carboidratos foliares durante a floração e os estádios iniciais de crescimento de frutilhos em tangerineira 'Ponkan'

O objetivo deste trabalho foi avaliar os teores de carboidratos foliares em tangerineira 'Ponkan' (Citrus reticulata), durante o pleno florescimento e os estádios iniciais de crescimento dos frutilhos, e identificar a melhor época para realização do raleio químico. O experimento foi realizado durante dois anos de produção (2009/2010 e 2010/2011), com quatro épocas de amostragem de folhas: 0, 30, 60 e 90 dias após o pleno florescimento. Utilizou-se o delineamento de blocos ao acaso, com parcelas subdivididas no tempo, quatro repetições e quatro plantas por parcela. Para determinar a utilização dos carboidratos pelas plantas, foram avaliados os teores de açúcares solúveis, açúcares redutores e amido na matéria seca das folhas. Nas mesmas épocas de amostragem foliar, o tamanho dos frutilhos também foi determinado. Os teores foliares de carboidratos solúveis aumentam e os de amido diminuem entre 35 e 50 dias após o pleno florescimento. No final da fase de fixação dos frutilhos, a partir dos 50 dias após o pleno florescimento, os teores de açúcares solúveis nas folhas diminuem. A melhor época para a realização do raleio químico em tangerineira 'Ponkan' é logo após a fase de queda fisiológica, quando os frutilhos atingirem cerca de 18 mm de diâmetro, aos 50 dias após o pleno florescimento.

Estimativa de repetibilidade para caracteres de qualidade de frutos de laranjeira-doce

O objetivo deste trabalho foi determinar o coeficiente de repetibilidade de características de qualidade do fruto de laranjeira-doce (Citrus sinensis) e o número mínimo de avaliações capaz de proporcionar níveis de certeza da predição do valor real dos genótipos. Foram avaliados, em cinco safras, 39 genótipos de laranjeira-doce, coletados em nove municípios do Estado do Acre. Utilizou-se o delineamento de blocos ao acaso, com três repetições. Foram avaliados: massa média de frutos, rendimento de suco, espessura de casca, sólidos solúveis (SS), acidez total (AT), relação SS/AT e índice tecnológico. Para a estimativa dos coeficientes de repetibilidade, foram utilizados os métodos da análise de variância, dos componentes principais e da análise estrutural. Todos os caracteres avaliados mostraram variabilidade, exceto o rendimento de suco. Os caracteres avaliados mostraram padrão cíclico, o que foi mais bem captado pelas metodologias multivariadas de estimativa do coeficiente de repetibilidade. São necessárias 15 avaliações para determinar, com 90% de certeza, os caracteres espessura de casca e sólidos solúveis, e 11, 6, 3, 2 e 1 avaliações, respectivamente para massa média de fruto, acidez total, índice tecnológico, rendimento de suco e relação SS/AT.

Plant growth and leaf-spot severity on eucalypt at different CO2concentrations in the air

The objective of this work was to evaluate the effects of increased air-CO2 concentration on plant growth and on leaf-spot caused by Cylindrocladium candelabrumin Eucalyptus urophylla. Seedlings were cultivated for 30 days at 451, 645, 904, and 1,147 µmol mol-1 CO2 ; then, they were inoculated with the pathogen and kept under the same conditions for seven days. Increased CO2concentration increased plant height and shoot dry matter mass, and decreased disease incidence and severity. Stem diameter was not affected by the treatments. Increased concentrations of atmospheric CO2 favorably affect eucalypt growth and reduce leaf-spot severity.