Tesi sul tema "Citrus"

O cancro cítrico é uma doença de importância mundial para o cultivo comercial de citros. O desenvolvimento de medidas alternativas de controle que contribuam para o manejo da doença é de grande importância. O presente estudo teve como objetivo verificar se o tratamento de plantas cítricas com inseticidas neonicotinóides é capaz de controlar a incidência de cancro cítrico na planta, assim como, verificar se as plantas cítricas apresentam diferenças no vigor em decorrência do tratamento com estes produtos. A atividade antimicrobiana de acetamipride, imidaclopride (IMI) e tiametoxam (TMX) foi testada in vitro com seis isolados de Xanthomonas citri subsp. citri (Xcc). Alíquotas das suspensões bacterianas (108 UFC/ml) foram depositadas em meio Agar Nutriente acrescido dos produtos nas concentrações de 0 a 3000 µg/ml de princípio ativo (p.a.). Em casa-de-vegetação, plantas de laranja Valência foram tratadas com quatro doses de IMI, por rega no solo, em intervalos de tempo variando entre 0 e 10 dias, entre a inoculação de Xcc, por infiltração com seringa, e o tratamento com o neonicotinóide. Plantas controle foram tratadas com água e inoculadas com a bactéria. Foi avaliada a incidência por contagem de lesões de cancro cítrico por cm2 de área foliar, a dinâmica populacional da bactéria por reisolamento e o teor de macronutrientes e micronutrientes das plantas tratadas e inoculadas. A campo, foram conduzidos experimentos nos municípios de Paranavaí e São João do Caiuá, PR, em plantas de laranja Natal (quatro meses) e Valência (oito meses), respectivamente. As plantas foram tratadas com cinco doses de IMI no tronco ou no solo, e uma dose de TMX e de clotianidina no solo. Plantas controle foram tratadas com água. Foram avaliadas bimestralmente as incidências de cancro cítrico, de larva minadora dos citros (LMC) e a desfolha. O vigor vegetativo foi avaliado pelo volume de copa, pelo diâmetro de caule e pela altura de planta. Os neonicotinóides não apresentaram efeito antimicrobiano contra nenhum isolado de Xcc in vitro. O tratamento das plantas com IMI, além de reduzir o número de lesões de cancro cítrico e a população bacteriana na planta, alterou as características dessas lesões, independente da dose, época do ano ou do intervalo de tempo entre o tratamento da planta e a inoculação com Xcc. De maneira geral, nos experimentos a campo, foi constatada diminuição de desfolha e das incidências de cancro cítrico e de LMC para todos os neonicotinóides testados, destacando-se a menor dose de IMI aplicada no tronco da planta e a maior dose de IMI aplicada no solo. Os neonicotinóides testados reduziram a incidência de cancro cítrico, aumentaram os teores de N e K nas folhas e o vigor vegetativo das plantas cítricas. Assim, os neonicotinóides podem ser considerados no manejo do cancro cítrico, não somente por sua ação inseticida, mas também como bioativadores das plantas cítricas.
Citrus canker is a disease of global importance for citrus production. The development of alternative measures to control the disease is of great importance. This study aimed to determine if the aplication of neonicotinoid insecticides on citrus plants is capable to control the incidence of citrus canker by induced resistance, as well as to verify if citrus plants differ in vigor due to the treatment with these products. The antimicrobial activity of acetamiprid, imidacloprid (IMI) and thiamethoxan (TMX) was tested in vitro against six isolates of Xanthomonas citri subsp. citri (Xcc). Aliquots of bacterial suspensions (108 CFU / ml) were placed on nutrient agar medium containing the neonicotinoids in concentrations from 0 to 3000 µg/ml of active ingrediente (a.i). In greenhouse, plants of Valencia orange were treated with four doses of IMI by soil drench. Intervals of time ranging from 0 and 10 days, between the inoculation of Xcc by syringe infiltration and treatment with the neonicotinoid were tested. Check plants were treated with water and inoculated with the bacteria. The plants were evaluated by counting the incidence of citrus canker lesions per cm2 of leaf area, the population dynamics of bacteria by reisolations, and macronutrient and micronutrient content of the inoculated and treated plants. Field experiments were carried out in the counties of Paranavaí and São João do Caiuá, PR, on four months old Natal orange plants and eight months old Valencia orange plants, respectively. The plants were treated with five doses of IMI in the trunk or by soil drench. Doses of TMX and clothianidin by soil drench. Check plants were treated with water. The plants were evaluated every two months by determining the incidence of citrus canker, defoliation and citrus leafminer (CLM) incidence. The vigor of the plants was assessed by the volume of canopy, trunk diameter and plant height. The neonicotinoids showed no antimicrobial effect against the strain of Xcc in vitro. The treatment of plants with IMI in addition to reducing the number of lesions of citrus canker and bacterial population in the plant, also changed the characteristics of these injuries, regardless of dose, time of the year or interval between treatment and inoculation of plants with Xcc. In general, experiments in the field showed decrease in the incidence of citrus canker for all neonicotinoids tested, especially the lower dose of IMI applied in the trunk of the plants and the highest dose of IMI by soil drench. The neonicotinoids tested reduced the incidence of citrus canker, increased the levels of N and K and the vigor of citrus plants. Thus the neonicotinoids may be considered in the management of citrus canker not only by its activity as insecticide to control CLM, but also by the bioactivity on citrus plants.

Mealybugs (Planococcus citri) and thrips (Scirtothrips aurantii) are common and extremely damaging citrus crop pests which have proven difficult to control via conventional methods, such as chemical pesticides and insect growth regulators. The objective of this study was to determine the efficacy of entomopathogenic fungi against these pests in laboratory bioassays. Isolates of Metarhizium anisopliae and Beauveria bassiana from citrus orchards in the Eastern Cape, South Africa were maintained on Sabouraud Dextrose 4% Agar supplemented with Dodine, chloramphenicol and rifampicin at 25°C. Infectivity of the fungal isolates was initially assessed using 5th instar false codling moth, Thaumatotibia leucotreta, larvae. Mealybug bioassays were performed in 24 well plates using 1 x 107 ml-1 conidial suspensions and kept at 26°C for 5 days with a photoperiod of 12 L:12 D. A Beauveria commercial product and an un-inoculated control were also screened for comparison. Isolates GAR 17 B3 (B. bassiana) and FCM AR 23 B3 (M. anisopliae) both resulted in 67.5% mealybug crawler mortality and GB AR 23 13 3 (B. bassiana) resulted in 64% crawler mortality. These 3 isolates were further tested in dose-dependent assays. Probit analyses were conducted on the dose-dependent assays data using PROBAN to determine LC₅₀ values. For both the mealybug adult and crawlers FCM AR 23 B3 required the lowest concentration to achieve LC₅₀ at 4.96 x 10⁶ conidia ml-1 and 5.29 x 10⁵ conidia ml-1, respectively. Bioassays on adult thrips were conducted in munger cells with leaf buds inoculated with the conidial suspensions. Isolate GAR 17 B3 had the highest mortality rate at 70% on thrips while FCM AR 23 B3 resulted in 60% mortality. Identification of the isolates, FCM AR 23 B3, GAR 17 B3 and GB AR 23 13 3, were confirmed to be correct using both microscopic and molecularly techniques. ITS sequences were compared to other sequences from GenBank and confirmed phylogenetically using MEGA6. Mealybug infection was investigated using scanning electron microscopy, mycosis was confirmed but the infection process could not be followed due to the extensive waxy cuticle. These results indicate that there is potential for the isolates FCM AR 23 B3 and GAR 17 B3 to be developed as biological control agents for the control of citrus mealybug and thrips. Further research would be required to determine their ability to perform under field conditions.

Orientadores: Marcos Antonio Machado, Alessandra Alves de Souza
Tese (doutorado) - Universidade Estadual de Campinas, Instituto de Biologia
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Resumo: Avaliou-se as alterações transcricionais em laranja 'Pera' (Citrus sinensis L. Osb.) promovidas por quitosana (CHI) e ácido salicílico (SA), utilizando RNA-seq, e o efeito destes compostos no controle do cancro cítrico (Xanthomonas citri subsp. citri) e da clorose variegada dos citros (CVC - Xylella fastidiosa). As plantas foram tratadas com CHI ou SA e após 48h e 24h, respectivamente, foram coletadas amostras foliares para avaliar seus transcriptomas. Para a avaliação dos eliciadores sobre o cancro cítrico e a CVC, as plantas foram tratadas com CHI ou SA e após 48h e 24h, respectivamente, inoculadas com as duas bactérias separadamente. A partir de 24h da inoculação, foram coletadas amostras foliares para avaliar a curva de crescimento de ambas as bactérias, a redução da severidade e/ou incidência das doenças e respostas de defesa da planta por RT-qPCR. Com os resultados do transcriptoma, observou-se que mais genes foram induzidos pelo tratamento com SA do que com CHI. O tratamento com SA aumentou a expressão de genes que participam da via de sinalização do SA na planta (WRKY50, PR2 e PR-9) e genes da biossíntese do etileno e ácido jasmônico (ACS 12, fator de transcrição contendo domínio AP2 e OPR3). Além disso, promoveu a indução de genes relacionados ao metabolismo secundário, processos redox e estresse biótico. No tratamento com CHI, foi observada maior indução de genes relacionados ao metabolismo secundário. Para ambos os tratamentos, a via da auxina foi reprimida. No experimento para controle do cancro cítrico, observou-se que ambos os eliciadores promoveram reduções na severidade e incidência da doença. Entretanto, a CHI pareceu não interferir diretamente na formação do biofilme pela bactéria, mas pode ter dificultado a multiplicação de X. citri na planta. O SA retardou a entrada da bactéria na planta e, aparentemente, inibiu mais a formação do biofilme bacteriano do que a CHI. Comparações da expressão gênica entre os eliciadores reforçam a ideia de que a CHI tem maior potencial de induzir resistência ao cancro cítrico do que SA. No experimento para o controle da CVC, observou-se que a CHI induziu importantes genes da via do SA (NPR1, TGA, EDS1) e etileno (EIN-3, PR-4) 24h após a inoculação. Aplicações exógenas de SA potencializaram o seu efeito endógeno na planta, pois houve indução de NPR1, TGA e PRs. Entretanto, não foi possível estabelecer uma relação clara entre a multiplicação de X. fastidiosa, a incidência da doença e o uso da CHI e SA em laranja 'Pera', já que na maioria das avaliações não houve redução da população bacteriana em amostras foliares e não houve redução da incidência em plantas tratadas. Com base nos resultados, observou-se que CHI e SA induziram diversos genes envolvidos em respostas de defesa em laranja 'Pera'. Entretanto, essas respostas podem ser moduladas diferencialmente a depender do patógeno que afeta a planta, pois os eliciadores foram eficientes no controle da X. citri, um patógeno que coloniza o mesófilo da planta, entretanto não foram efetivos no controle da X. fastidiosa, um patógeno que coloniza o xilema da planta, embora respostas de defesa tenham sido expressas nos momentos iniciais (24h) após a inoculação com X. fastidiosa
Abstract: This study was carried out to evaluate transcriptional modification in sweet orange 'Pera' (Citrus sinensis L. Osb.), promoted by chitosan (CHI) and salicylic acid (SA), using RNA-seq, and the effect of these compounds on citrus canker (Xanthomonas citri subsp. citri) and citrus variegated chlorosis (CVC - Xylella fastidiosa). Plants were treated with CHI or SA and after 48h and 24h, respectively, leaf samples were collected to assess the transcriptome. In the experiments for disease assessment, the plants were treated with CHI or SA and after 48h and 24h, respectively, inoculated. Starting from 24h after inoculation, leaf samples were collected to evaluate the multiplication of the pathogens (X. citri and X. fastidiosa), reduction of the severity and / or incidence and plant defense responses by RT-qPCR. Based upon the transcriptome results, it was observed that more genes were induced by SA than by CHI. SA treatment increased the expression of genes that participate in the SA signaling pathway in the plant (WRKY50, PR2 and-PR9), and genes involved in the biosynthesis of ethylene and jasmonic acid (ACS 12, transcription factor containing AP2 and OPR3 domain). Besides these, SA promoted induction of genes of secondary metabolism, redox processes and biotic stress. The treatment with CHI exhibited higher induction of genes related to secondary metabolism. For both treatments, the auxin pathway was suppressed. In the experiment for the control of citrus canker, it was observed that both elicitors reduced the severity and incidence of the disease. However, CHI seems not to interfere directly in biofilm formation, but may have hindered the multiplication of X. citri in the plant. The SA slowed down the entry of the bacteria into the plant and, apparently, inhibited the formation of biofilm more efficiently than the CHI. Comparisons of gene expression between elicitors reinforce the idea that CHI has higher potential to induce resistance to citrus canker than SA. In the experiment for the control of CVC, it was observed that the CHI induced important genes of the SA (NPR1, TGA, EDS1) and ethylene (EIN-3, PR-4) pathways 24h after inoculation. Exogenous applications of SA potentiated its endogenous effect in the plant, since there was induction of EDS-1, NPR1, TGA and PRs. However, it was not possible to establish a clear relationship between the multiplication of X. fastidiosa, the incidence of the disease and the use of CHI and SA in 'Pera' sweet orange, since most of the assessments did not show reduction of bacterial populations in leaf samples and there was no reduction of the incidence in treated plants. Based upon the results of this study, it was observed that CHI and SA induced several genes involved in defense responses in 'Pera' sweet orange. However, these responses can be modulated differentially depending on the pathogen that affects the plant. This fact was demonstrated in this study, as elicitors were effective in controlling X. citri, a pathogen that colonizes the mesophyll of the plant, but were not effective in controlling X. fastidiosa, a pathogen that colonizes the xylem of the plant, although defense responses were expressed in the early stages (24 h) after inoculation with X. fastidiosa
Doutorado
Bioquimica
Doutora em Biologia Funcional e Molecular

4 pp.
Publication contains an introduction to the rationale for pruning as well as sections on when to prune, what part of the tree to prune, techniques for best pruning, and how to protect the tree following pruning. Six figures are included.

Orientador: Silvio Aparecido Lopes
Resumo: Huanglongbing (HLB) é a doença mais devastadora dos citros. Está associada a bactérias de floema disseminadas eficientemente pelo psilídeo Diaphorina citri, inseto altamente prolífico e com ampla gama de hospedeiros dentre as rutáceas. Diversas espécies desta família ocorrem em matas onde o inseto poderia estar se reproduzindo e reinfestando pomares além de servir como fonte de multiplicação de Candidatus Liberibacter asiaticus (CLas), dificultando o controle do HLB. Nesse sentido foram realizados dois estudos, sendo o primeiro para avaliar a sobrevivência de D. citri em folhas jovens recém-diferenciadas e totalmente expandida e a reprodução de D. citri em brotos em 20 espécies de rutáceas. E o segundo para avaliar a reação dessas espécies após inoculação do patógeno por meio de enxertia de tecidos infectados e uso de adultos infectivos de D. citri. No primeiro, classificou-se as espécies quanto a favorabilidade em reproduzir D. citri em quatro grupos de plantas: Grupo I (altamente favorável ao inseto) incluiu Citrus ×aurantium 'Valência', Citrus limonia, Murraya paniculata (syn. Murraya exotica L.) (Aurantioideae: Aurantieae) e Bergera koenigii (Aurantioideae: Clauseneae). As espécies do Grupo II (de intermédiio a baixa favorabilidade) incluiu Citrus (Poncirus) trifoliata ‘Pomeroy’, Citrus wintersii, Swinglea glutinosa (Aurantieae) e Clausena lansium (Clauseneae). O grupo III (não favorável) incluiu Aegle marmelos, Atalantia buxifolia e Citrus (‘Microcitrus’) sp. (Aurantieae... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: Huanglongbing (HLB) is the most devastating disease of citrus. It is associated with phloem bacteria efficiently disseminated by the psilid Diaphorina citri, a highly prolific insect with a wide range of hosts among the rutaceous. Several species of this family occur in forests where the insect could be reproducing and reinfesting orchards besides serving as a source of multiplication of Candidatus Liberibacter asiaticus (CLas), hindering the control of HLB. In this sense, two investigations were carried out, the first one to evaluate the survival of D. citri on newly differentiated and fully-expanded soft leaves and the shoots reproduction of D. citri in 20 species of rutaceous. And the second to evaluate the reaction of these species after inoculation of the pathogen by grafting of infected tissues and use of infective adults of D. citri. In the first, this allowed suitability of the plants as hosts of the psyllid to be separated into 4 groups of plants. Group I (highly suitable to the insect) species included Citrus ×aurantium ‘Valencia’, Citrus limonia, and Murraya paniculata (syn. Murraya exotica L.) (Aurantioideae: Aurantieae), and Bergera koenigii (Aurantioideae: Clauseneae). Group II (intermediate to low suitability) species included Citrus (Poncirus) trifoliata ‘Pomeroy’, Citrus wintersii, and Swinglea glutinosa (Aurantieae), and Clausena lansium (Clauseneae). Group III (not suitable) included Aegle marmelos, Atalantia buxifolia, and Citrus (‘Microcitrus’) sp. (Auran... (Complete abstract click electronic access below)
Doutor

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
O ácaro Brevipalpus yothersi Baker é vetor da leprose dos citros, principal doença viral da citricultura mundial. Informações sobre a biologia de B. yothersi são essenciais para compreender a dinâmica populacional do ácaro no campo e inferir se mudanças no manejo do pomar em função da região pode alterar a biologia do ácaro. O objetivo do trabalho foi determinar a biologia e elaborar a tabela de vida de fertilidade de B. yothersi coletados em diferentes regiões citrícolas do estado de São Paulo. Os experimentos foram realizados no Laboratório de Acarologia, pertencente à Faculdade de Ciências Agrárias e Veterinárias - FCAV/UNESP, Jaboticabal - SP. Os ácaros foram coletados em pomares cítricos das regiões de Barretos, Jales e Santa Cruz do Rio Pardo, posteriormente, em laboratório, foram multiplicados em frutos de laranja. Os parâmetros biológicos avaliados foram duração das fases de desenvolvimento, oviposição, período de incubação, viabilidade dos ovos, longevidade, taxa líquida de reprodução (Ro), tempo médio de geração (T), taxa intrínseca de crescimento populacional (rm) e taxa finita de crescimento populacional (λ). Estes parâmetros foram avaliados em dois experimentos, o primeiro consistiu na biologia de B. yothersi em frutos isentos de resíduos de produtos fitossanitários à 23±1ºC e o segundo sob frutos com resíduo de espirodiclofeno à 25±1ºC. As observações foram realizadas diariamente, pela manhã e ao fim da tarde. A duração do desenvolvimento, longevidade, período de pré-oviposição, taxa de oviposição e número de ovos de B. yothersi apresentaram diferenças entre as populações. O ciclo biológico de B. yothersi criados sobre frutos de laranja varia de 18 a 24 dias, considerando os intervalos de 22º a 26 ºC, 50 a 70% de umidade relativa e fotofase de 14 horas. As diferenças biológicas entre populações de B. yothersi indicam que há diferenças na dinâmica populacional do ácaro no campo, necessitando de manejos adequados para cada região para melhor controle da leprose.
The mite Brevipalpus yothersi Baker is the vector of the citrus leprosis, major viral disease of citrus worldwide. Information about B. yothersi's biology are essential to understanding the population dynamics of the mite in the field and infer whether changes in orchard management by region can change the mite biology. The objective was to determine the biology and prepare the fertility life table of B. yothersi collected in different citrus regions of São Paulo state. The experiments were performed in Acarology Laboratory, belonging to the Faculty of Agricultural and Veterinary Sciences - FCAV/UNESP, Jaboticabal - SP. The mites were collected in citrus orchards in the regions of Barretos, Jales and Santa Cruz do Rio Pardo, later in the laboratory were multiplied in orange fruits. The biological parameters assessed were duration of the stages of development, oviposition, incubation period, egg viability, longevity, net reproductive rate (Ro), mean generation time (T), intrinsic rate of increase (rm) and finite rate increase (λ). These parameters were evaluated in two experiments, the first consisted the biology of B. yothersi in fruits free of residues of pesticides at 23 ± 1°C and the second consisting of the biology of B. yothersi under fruit with spirodiclofen residue at 25 ± 1°C . The observations were performed daily, in the morning and in the afternoon. The duration of the development, longevity, pre-oviposition period, oviposition rate and number of B. yothersi eggs showed differences between populations. The life cycle of B. yothersi created on orange fruit ranges from 18 to 24 days, considering the ranges of 22 to 26°C, 50-70% relative humidity and 14 hours. The biological differences between populations of B. yothersi indicate that there are differences in the population dynamics of the mite in the field, requiring adequate management practices for each region for better control of leprosis.

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
A citricultura é uma das principais atividades do agronegócio brasileiro. Entretanto, inúmeras pragas e doenças atacam os citros, causando grandes prejuízos econômicos. O cancro cítrico, causado pela bactéria Xanthomonas citri subsp. citri (Xac), é um grave problema para o setor, não havendo ainda um método eficaz para o seu controle. Neste estudo, utilizando RNASeq, foram analisados os perfis transcricionais de Xac inoculada em duas espécies de citros contrastantes à doença: laranja doce ‘Pêra Rio’ (Citrus sinensis L. Osbeck), menos suscetível e lima ácida ‘Galego’ (Citrus aurantifolia Swingle), altamente suscetível, às 48 e 72 horas após a infecção (hai), com o objetivo de identificar genes de Xac envolvidos no processo de infecção. Foram identificados 80 genes de Xac diferencialmente expressos (GDEs) no hospedeiro laranja doce ‘Pêra Rio’, sendo 41 e 39 nos tempos de 48 e 72 hai, respectivamente. Em lima ácida ‘Galego’ foram identificados 82 GDEs, sendo 40 no tempo de 48 hai e 42 em 72 hai. Alguns destes genes diferencialmente expressos foram avaliados pela técnica de PCR quantitativa em tempo real, sendo estes hpa1, hrpE, hrpW, virK, ahpC, katE, katG, cydA e cydB, os quais estão envolvidos na patogenicidade e virulência, na defesa ao estresse oxidativo e na fosforilação oxidativa. Os genes de patogenicidade e virulência foram induzidos em Xac em ambos os hospedeiros, enquanto que os genes relacionados à cadeia respiratória foram inibidos em ambos os hospedeiros, com maior inibição em lima ácida ‘Galego’. No entanto, os genes relacionados ao estresse oxidativo apresentaram um perfil de expressão maior em Xac na interação com laranja doce ‘Pêra Rio’ do que em lima ácida ‘Galego’, principalmente ahpC e katG. A determinação da concentração de H2O2 nas folhas revelou que laranja doce ‘Pêra Rio’, espécie menos suscetível ao cancro cítrico, possui maior quantidade de H2O2 do que lima ácida ‘Galego’, espécie altamente suscetível. Isso sugere que a menor susceptibilidade ao cancro cítrico da laranja ‘Pêra Rio’ pode estar relacionada com a maior quantidade de H2O2 presente nesta espécie, o que leva a bactéria a ativar seu arsenal de enzimas para combater o estresse oxidativo do meio, retardando a infecção.
The citrus industry is one of the main activities of Brazilian agribusiness. However, many pests and diseases attack citrus, causing great economic losses. The citrus canker, caused by Xanthomonas citri subsp. citri (Xac), is a major problem for the sector, there is not yet an effective method for its control. In this study, the transcriptional profiles of Xac inoculated in two species of contrasting citrus disease were analyzed using RNA-Seq : sweet orange 'Pêra Rio' (Citrus sinensis L. Osbeck), moderately tolerant and Mexican Lime 'Galego' (Citrus aurantifolia Swingle) highly susceptible at 48 and 72 hours after infection (hai) aiming to identify Xac genes involved in the infection process. We identified 80 Xac differentially expressed genes (DGE) in sweet orange 'Pera Rio', 41 and 39 at 48 and 72 hai, respectively. In Mexican Lime 'Galego' 82 DGE were identified, 40 at 48 and 42 at 72 hai. Some of these differentially expressed genes were evaluated by real time quantitative PCR : hpa1, hrpE, hrpW, Virk, ahpC, KatE, katG, cyda and cydB, which are involved in pathogenicity and virulence, oxidative stress defense and oxidative phosphorylation. The pathogenicity and virulence genes were induced in Xac in both hosts, whereas the respiratory chain-related genes were inhibited in both hosts with greater inhibition in Mexican lime 'Galego'. However, genes related to oxidative stress showed a higher expression profile in Xac interaction with sweet orange 'Pera Rio' than with Mexican lime 'Galego', mainly ahpC and katG. The determination of H2O2 concentration in leaves revealed a higher amount of H2O2 in sweet orange 'Pêra Rio' moderately tolerant to citrus canker, than in Mexican Lime 'Galego' highly susceptible to citrus canker. The results suggests that the lower susceptibility to citrus canker orange 'Pera Rio' may be related to the greater amount of H2O2 present in this specie, which leads the bacteria to activate their arsenal of enzymes to fight oxidative stress environment, slowing the infection.

Orientador: Jesus Aparecido Ferro
Coorientador: Roberto Hirochi Herai
Coorientador: Juliana da Silva Vantini
Banca: José Belasque Júnior
Banca: Priscila Lupino gatão
Resumo: A citricultura é uma das principais atividades do agronegócio brasileiro. Entretanto, inúmeras pragas e doenças atacam os citros, causando grandes prejuízos econômicos. O cancro cítrico, causado pela bactéria Xanthomonas citri subsp. citri (Xac), é um grave problema para o setor, não havendo ainda um método eficaz para o seu controle. Neste estudo, utilizando RNASeq, foram analisados os perfis transcricionais de Xac inoculada em duas espécies de citros contrastantes à doença: laranja doce 'Pêra Rio' (Citrus sinensis L. Osbeck), menos suscetível e lima ácida 'Galego' (Citrus aurantifolia Swingle), altamente suscetível, às 48 e 72 horas após a infecção (hai), com o objetivo de identificar genes de Xac envolvidos no processo de infecção. Foram identificados 80 genes de Xac diferencialmente expressos (GDEs) no hospedeiro laranja doce 'Pêra Rio', sendo 41 e 39 nos tempos de 48 e 72 hai, respectivamente. Em lima ácida 'Galego' foram identificados 82 GDEs, sendo 40 no tempo de 48 hai e 42 em 72 hai. Alguns destes genes diferencialmente expressos foram avaliados pela técnica de PCR quantitativa em tempo real, sendo estes hpa1, hrpE, hrpW, virK, ahpC, katE, katG, cydA e cydB, os quais estão envolvidos na patogenicidade e virulência, na defesa ao estresse oxidativo e na fosforilação oxidativa. Os genes de patogenicidade e virulência foram induzidos em Xac em ambos os hospedeiros, enquanto que os genes relacionados à cadeia respiratória foram inibidos em ambos os hospedeiros, com maior ... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: The citrus industry is one of the main activities of Brazilian agribusiness. However, many pests and diseases attack citrus, causing great economic losses. The citrus canker, caused by Xanthomonas citri subsp. citri (Xac), is a major problem for the sector, there is not yet an effective method for its control. In this study, the transcriptional profiles of Xac inoculated in two species of contrasting citrus disease were analyzed using RNA-Seq : sweet orange 'Pêra Rio' (Citrus sinensis L. Osbeck), moderately tolerant and Mexican Lime 'Galego' (Citrus aurantifolia Swingle) highly susceptible at 48 and 72 hours after infection (hai) aiming to identify Xac genes involved in the infection process. We identified 80 Xac differentially expressed genes (DGE) in sweet orange 'Pera Rio', 41 and 39 at 48 and 72 hai, respectively. In Mexican Lime 'Galego' 82 DGE were identified, 40 at 48 and 42 at 72 hai. Some of these differentially expressed genes were evaluated by real time quantitative PCR : hpa1, hrpE, hrpW, Virk, ahpC, KatE, katG, cyda and cydB, which are involved in pathogenicity and virulence, oxidative stress defense and oxidative phosphorylation. The pathogenicity and virulence genes were induced in Xac in both hosts, whereas the respiratory chain-related genes were inhibited in both hosts with greater inhibition in Mexican lime 'Galego'. However, genes related to oxidative stress showed a higher expression profile in Xac interaction with sweet orange 'Pera Rio' than with Mexica... (Complete abstract click electronic access below)
Mestre

Orientador: Alessandro de Mello Varani
Resumo: As plantas como organismos sésseis requerem a síntese e o acúmulo de uma ampla variedade de moléculas envolvidas no crescimento, desenvolvimento e processos relacionados à defesa. Os transportadores Proteínas de Extrusão Multi- Antimicrobianas (MATE), Cassette de Ligação de ATP (ABC) e Superfamília dos Facilitadores Maioritários (MFS) são as principais famílias de transportadores de membrana em plantas, desempenhando um papel central nos processos relacionados à defesa nas interações planta-patógenos. Por exemplo, protegem as células das espécies de Citros sob a infecção de Xanthomonas citri subsp. citri (Xac), o agente etiológico do Cancro Cítrico tipo A, uma das doenças de Citros mais devastadoras envolvidas em sérios impactos econômicos e ambientais. Aqui, identificamos genes e transcritos das famílias MATE, ABC e MFS usando o genoma disponível de Citrus sinensis (v2.0 HZAU) e o Transcriptoma Referência de Citros (CRT) re-anotado da base de dados CitrusKB (http://bioinfo.deinfo.uepg.br). Foram identificados 67 genes MATE, 91 MFS e 143 ABC no genoma de C. sinensis e 82 transcritos MATE, 139 MFS e 226 ABC no CRT. Os transcritos foram mapeados no genoma de C. sinensis, revelando uma alta taxa de genes parálogos e putativos eventos de splicing alternativo (AS), cujos perfis de expressão gênica e potenciais papéis na interação Citros-Xac foram propostos. As cópias de genes em tandem e cópias dispersas juntamente com genes que possivelmente sofreram eventos de AS representam fon... (Resumo completo, clicar acesso eletrônico abaixo)
Abstract: Plants as sessile organisms require the synthesis and accumulation of a large array of molecules involved in growth, development, and defense-related processes. The Multi-Antimicrobial Extrusion Protein (MATE), ATP-Binding Cassette (ABC) and Major Facilitator Superfamily (MFS) transporters are the largest families of membrane transporters in plants, playing a central role in the defense-related processes in plant- pathogen interactions. For instance, protecting Citrus species cells under the infection of Xanthomonas citri subsp. citri (Xac), the etiologic agent of the Citrus Canker type A, one of the most devastating Citrus diseases involved with serious economic and environmental impacts. Herein, we identified genes and transcripts from MATE, ABC, and MFS families using the available Citrus sinensis genome (v2.0 HZAU) and the re-annotated Citrus Reference Transcriptome (CRT) from CitrusKB Knowledge Base (http://bioinfo.deinfo.uepg.br). We identified 67 MATE, 91 MFS, and 143 ABC genes in the C. sinensis genome and 82 MATE, 139 MFS, and 226 ABC transcripts in the CRT. The transcripts were mapped in the C. sinensis genome revealing a high rate of paralogs genes and probably alternative splicing (AS) events, whose expression profiles and potential roles in the Citrus-Xac interaction were proposed. The tandem and dispersed copies along with genes that underwent AS events represents sources of transporters’ genes diversity and complexity. Moreover, we also highlighted potential bi... (Complete abstract click electronic access below)
Mestre

5 pp.
Water is essential for citrus trees because it is an integral component of the biochemical reactions that occur within the plant, and also because it is the carrier that moves plant nutrients and other substances throughout the tree. This publication examines irrigating citrus trees. Topics include; water stress, estimating tree water use, timing water applications, dealing with salty water, calculating how much water is applied, as well as application methods and mulches. And it also includes a table which can be applied to spreadsheets, to calculate the exact amount of water a citrus tree needs.

3 pp.
Citrus budding is a plant propagation technique that any homeowner can do. Once the technique is learned, homeowners can add citrus tree. This publication addresses the budding techniques of citrus trees. Topics include the preparation prior to budding, selecting budsticks, storing budsticks, selecting and preparing the budding location, cutting the bud, inserting the bud in the t cut, and forcing the bud.

Thesis (MScAgric)--Stellenbosch University, 2012.
ENGLISH ABSTRACT: Planococcus citri (Risso) (Hemiptera: Pseudococcidae), the citrus mealybug, is a highly destructive pest of citrus in South Africa. The ability of mealybugs to develop resistance to chemicals, as well as their cryptic nature and protective wax coverings, all individually and combined, impair the ability of insecticides to control them. Furthermore, chemicals deplete natural enemy populations that usually establish control over mealybug populations well before harvest in March to May. The inability of chemicals to control P. citri successfully and the growing public awareness of the detrimental environmental effects and health risks that are associated with pesticides are pressuring citrus growers to find alternative methods to chemical control. Entomopathogenic nematodes of the Rhabditida order, belonging either to the families Heterorhabditidae or Steinernematidae, have proven to be valuable biocontrol agents of a variety of insect pest species. These nematodes are, however, not yet commercially available in South Africa. Various laboratory bioassays were conducted to determine the potential of entomopathogenic nematodes to control P. citri. Adult female P. citri were screened for susceptibility to six indigenous nematode species. Planococcus citri was found to be most susceptible to Steinernema yirgalemense and Heterorhabditis zealandica, causing 97% and 91% mortality, respectively. Both H. zealandica and S. yirgalemense completed their life cycles inside adult female P. citri. Under optimal conditions, H. zealandica and S. yirgalemense, at an application rate of 170 infective juveniles (IJs) / insect (LD90), were able to control P. citri efficiently with a discriminating dosage of 11(LD50). The water activity (aw) bioassay indicated that S. yirgalemense was two times more tolerant to lower levels of free water than H. zealandica, with aw50 = 0.96 and aw90 = 0.99, compared to H. zealandica with aw50 = 0.98 and aw90 = 1.01. Furthermore, S. yirgalemense proved able to locate and infect P. citri at a faster rate than H. zealandica. Nematode activity was not significantly affected when exposed to 15°C, 20°C and 25°C. Results of the exposure trial also showed that the first 2−4 h post-application was the most decisive time for establishing successful infection of mealybugs. Further experiments were conducted to determine the ability of two polymer products, Zeba® and Xanthan gum, and a surfactant, Nu-Film-P®, to improve nematode application suspensions. Despite not being as effective as Xanthan gum, Zeba® still retarded sedimentation significantly. The addition of 0.02% Xanthan gum did not improve the ability of nematodes to control P. citri at 60% and 80% relative humidity containing H. zealandica and 0.03% Zeba® increased mortality by 14% at 60% RH and by 22% at 80% RH. The same polymer formulation was tested with S. yirgalemense, with mortality being found to have increased by 21% at 60% RH and by 27% at 80% RH. The combined addition of Nu-Film-P® and Zeba® was able to retard sedimentation significantly, increasing the average number of nematodes deposited on 2-cm2 leaf discs. To illustrate those factors that should be taken into consideration before applying nematodes in an integrated pest management (IPM) programme for citrus, the compatibility of H. zealandica and S. yirgalemense with biological control agents and agrochemicals to which they are most likely to be exposed was determined. Bioassays showed the coccinellid beetle, Cryptolaemus montrouzieri, to be susceptible to both nematode species. Beetle larvae proved to be highly susceptible, with H. zealandica obtaining 80%, and S. yirgalemense obtaining 92% control. Adult beetles were found to be twice as susceptible to S. yirgalemense, with 64% mortality recorded, as to H. zealandica, with 30% mortality recorded. Tolerance of H. zealandica and S. yirgalemense infective juveniles to the affect of aqueous solutions of an insecticide (Cyperphos 500 E.C.®), two biopesticides (CryptogranTM and HelicovirTM), and two adjuvants (Nu-Film-P® and Zeba®) on the infectivity and survival of H. zealandica and S. yirgalemense was evaluated. Heterorhabditis zealandica proved to be highly compatible with all products tested. Significant increase in mortality of S. yirgalemense was recorded after 12 h exposure to CryptogranTM, HelicovirTM and Cyperphos 500 E.C.®, and after 6 h exposure to Nu-Film-P®. However, there was no decrease in the ability of S. yirgalemense to infect the insect hosts. Towards selecting the appropriate nematode species for field studies, the ability of S. yirgalemense and H. zealandica to control P. citri was first evaluated under less harsh simulated glasshouse conditions. The ability of both the above-mentioned nematode species to control P. citri, with and without the addition of 0.03% Zeba® and 0.06% Nu-Film-P®, was evaluated in a growth chamber at 75 ± 8% relative humidity and 22°C. All treatments resulted in significantly higher mortality of adult female P. citri than did the control. The addition of 0.03% Zeba® and 0.06% Nu-Film-P® to an H. zealandica and an S. yirgalemense suspension increased resultant mortality from 26% to 30%, and from 34% to 45%, respectively. This increase in mortality was, however, not significant. The ability of this formulation to prolong the ability of S. yirgalemense to infect P. citri and to prolong nematode survival was also evaluated under the same conditions and showed the formulation to improve both infectivity and survival for up to 2–3 h post-application. In the semi-field trial, S. yirgalemense was unable to obtain significant control of P. citri without the addition of 0.03% Zeba® to nematode application suspensions. However, with the addition of 0.03% Zeba® S. yirgalemnse was able to obtain up to 53% control. The study established that the polymer product Zeba® improves the ability of S. yirgalemense to infect P. citri by retarding desiccation and by buffering nematodes from suboptimal environmental conditions.
AFRIKAANSE OPSOMMING: Planococcus citri (Risso) (Hemiptera: Pseudococcidae), die sitrus witluis, is ʼn baie skadelike pes van sitrus in Suid Afrika. Die vermoë van witluise om weerstand teen chemiese middels te ontwikkel, hul kriptiese lewenswyse en die beskermende waslaag wat hul liggame omhul inhibeer gesamentlik en individueel die vermoë van insektisiede om witluis bevolkings te beheer. Chemiese middels verminder ook die natuurlike vyande wat gewoonlik witluis bevolkings beheer voor die sitrus oestydperk in Maart tot Mei. Die onvermoë van chemiese middels om P. citri suksesvol te beheer en verhoogde bewustheid van die publiek rond om die vernietigende omgewings impak en gesondheidsrisiko’s verbonde aan chemiese insek beheer, noodsaak sitrus produsente om alternatiewe beheermetodes te ontwikkel. Entomopatogeniese nematodes, van die orde Rhabditida wat aan die families Heterorhabditidae of Steinernematidae behoort, is bekend as effektiewe biologiese beheeragente van ʼn verskeidenheid insek pes spesies. Hierdie nematodes is egter tans nie kommersieel beskikbaar in Suid- Afrika nie. Om ten einde die vermoë van plaaslike nematode spesies te bepaal om P. citri te beheer, is verskeie biotoetse in die laboratorium uitgevoer. ʼn Vinnige siftings proses is uitgevoer om vas te stel watter plaaslike nematode spesies die hoogste persentasie mortaliteit van P. citri wyfies veroorsaak. Daar is bevind dat P. citri die mees vatbaarste is vir Steinernema yirgalemense en Heterorhabditis zealandica wat 97% en 91% mortaliteit respektiewelik veroorsaak het. Die ontwikkeling van beide H. zealandica en S. yirgalemense na die infektering van volwasse P. citri wyfies is gevolg en daar is bevind dat beide nematode spesies hul lewensiklusse kon voltooi in insek kadawers. Onder optimale toestande was H. zealandica en S. yirgalemense in staat om P. citri effektief te beheer, as hul teen ʼn konsentrasie van 170 infektiewe larwes (JIs) per insek (LD90) toegedien word, met ʼn diskriminerende toedienings dosis van 11 (LD50). ʼn Water aktiwiteit biotoets het gewys dat S. yirgalemense twee keer so verdraagsaam is teenoor laer vlakke van vrye water as wat H. zealandica is met aw50 = 0.96 en aw90 = 0.99, in vergelyking met H. zealandica met aw50 = 0.98 en aw90 = 1.01. Resultate van ʼn blootstellings toets het ook gewys dat S. yirgalemense die vermoë het om P. citri vinniger op te spoor en te infekteer as H. zealandica. Die blootstellings toets was uitgevoer teen 15°C, 20°C en 25°C en daar is bevind dat die aktiwiteit van H. zealandica nie beduidend verhoog het met ʼn verhoging in temperatuur nie. Resultate vir die blootstellings toets het ook gewys dat die eerste 2 tot 4 uur na toediening van nematodes die mees kritieke tyd is om suksesvolle infektering van witluise te bevestig. Verdere eksperimente is uitgevoer om te bepaal of die byvoeging van twee polimeer produkte nl. Zeba® en Xanthan gum en ʼn benatter Nu-Film-P®, nematode toedienings suspensies kan verbeter. Deur 0.02% Xanthan gum by nematode toedienings suspensies te voeg is die vermoë van nematodes om P. citri by 60% en 80% relatiewe humiditeit (RH) te beheer nie beduidend verhoog nie, terwyl die byvoeging van 0.03% Zeba® wel ʼn beduidende verhoging in mortaliteit veroorsaak het. ʼn Suspensie van H. zealandica en 0.03% Zeba® het beheer met 14% by 60% RH en met 22% by 80% RH verhoog. Die invloed van dieselfde polimeer formulasie was ook getoets op S. yirgalemense en mortaliteit het verhoog met 21% by 60% RH en met 27% by 80% RH. Ten spyte daarvan dat “Xanthan gum” nie so effektief was om afsakking van nematodes te verhoed nie, het die byvoeginging van 0.03% Zeba® by toedienings suspensies steeds ʼn beduidende invloed gehad. Die gekombineerde byvoeging Nu-Film-P® en Zeba® was in staat om die gemiddelde aantal nematodes gedeponeer op 2-cm2 blaar skyfies te verhoog. Om te wys watter faktore in ag geneem moet word voor EPNs as deel van ʼn geïntegreerde pes beheer program toegedien word, is die verenigbaarheid van H. zealandica en S. yirgalemense vir biologiese beheer agente en landbouchemikalieë, waaraan hulle heel waarskynlik blootgestel gaan word, bepaal. Biotoetse het gewys dat die liewenheersbesie, Cryptolaemus montrouzieri, vatbaar is vir beide nematode spesies. Die larwale fase was hoogs vatbaar met mortaliteit van 80% en 92% verkry deur die toedeining van H. zealandica en S. yirgalemense respektiewelik. Daar is ook bepaal dat volwasse besies twee maal so vatbaar is vir S. yirgalemense wat 64% mortaliteit veroorsaak het, as vir H. zealandica met 30% mortaliteit aangeteken. Verdraagsaamheid van IJs van H. zealandica en S. yirgalemense met oplossings van ʼn insektisied (Cyperphos 500 E.C.®), twee bio-plaagdoders (CryptogranTM en HelicovirTM), en twee byvoegmiddels (Nu-Film-P® en Zeba®), vir infektiwiteit en oorlewing is bepaal. Resultate het gewys dat H. zealandica hoogs verenigbaar is met alle produkte wat getoets is. ʼn Beduidende verhoging in mortaliteit van S. yirgalemense is aangeteken na 12 ure se blootstelling aan CryptogranTM, HelicovirTM en Cyperphos 500 E.C.®, en na 6 ure se blootstelling aan Nu-Film-P®. Resultate het egter ook gewys dat die vermoë van S. yirgalemense om om gashere te infekteer nie deur beinvloed word nie. Om ten einde te bepaal watter een van H. zealandica of S. yirgalemense die beter isolaat sou wees om te gebruik in veldproewe, is hul vermoë om P. citri te beheer eers evalueer onder minder ongunstige gesimuleerde glashuis toestande. Die vermoë van beide nematode spesies om P. citri te beheer is bepaal in ʼn groeikamer by 75 ± 8% (RH) met en sonder die byvoeging van 0.03% Zeba® en 0.06% Nu-Film-P®. Mortaliteit verkry deur alle behandelings was beduidend hoër as die kontrole. Deur 0.03% Zeba® en 0.06% Nu-Film-P® by toedienings suspensies van H. zealandica en S. yirgalemense te voeg is die gevolglike mortaliteit van P. citri verhoog van 26% na 30% en van 34% na 45%, respektiewelik. Hierdie verhoging in mortaliteit was egter nie beduidend nie. Die vermoë van die bo-genoemde formulasie om oorlewing van S. yirgalemense en infeksie tydperk van P. citri te verleng was bepaal onder dieselfde toestande en daar is gevind dat die formulasie beide die infektiwiteit en oorlewing van nematodes met 2 tot 3 uur na toediening verleng het. Tydens die semi-veldproef was S. yirgalemense nie in staat om beduidend hoër mortaliteit van P. citri te verkry, in vergelyking met die kontrole, sonder die byvoeging van 0.03% Zeba® by toedienings suspensies nie. Met die byvoeging van Zeba® is daar tot 53% beheer verkry. Die studie het onweerlegbaar gewys dat die polimeer produk Zeba® die vermoë van S. yirgalemense om P. citri te infekteer verbeter deur die uitdroging van nematodes te vertraag en deur hul te beskerm teen ongunstige omgewings toestande.

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O cancro cítrico, causado pela bactéria Xanthomonas citri subsp. citri (Xac), é uma das principais doenças que acometem a citricultura mundial e ataca uma ampla gama de espécies comerciais de citros, causando perdas significativas nos países produtores. A espécie de laranja doce Hamlin (Citrus sinensis) é suscetível ao cancro cítrico, enquanto a espécie de tangerina Satsuma (Citrus unshiu) é resistente. Para compreender os mecanismos moleculares relacionados aos sistemas de defesa ativados pela planta é importante identificar as alterações transcricionais de cada espécie vegetal sob estresse fitopatogênico, a fim de desvendar os elementos moleculares que são específicos de cada espécie. O objetivo do presente trabalho foi realizar uma análise comparativa temporal do transcriptoma de duas espécies cítricas contrastantes em resposta à Xac, pela técnica do RNA-Seq (Illumina). Um total de 5.673 e 6.231 transcritos diferencialmente expressos foi induzido nos tempos 24, 48 e 72 horas após a inoculação de Xac em Satsuma e Hamlin, respectivamente, enquanto 3.982 e 7.944 transcritos foram reprimidos. Deste total, 52 transcritos foram induzidos em comum, nas duas espécies, em todos os tempos de inoculação. Estes genes estão relacionados com a defesa basal da planta contra o ataque de Xac, pois apresentam genes que participam na percepção e reconhecimento do patógeno, genes que codificam fatores de transcrição e genes que participam na defesa da planta, como glucanases e proteinases. Entre os genes induzidos exclusivamente na espécie resistente Satsuma destacou-se uma proteinase aspártica. Esta proteína apresentou a maior expressão gênica no tempo 24 horas e pode estar envolvida na resistência desta espécie, visto que na espécie suscetível Hamlin, a expressão desta proteína foi menos expressiva e tardia, no tempo 48 horas. Outra resposta oposta entre as espécie foi na expressão de genes relacionados à ...
Citrus canker, caused by Xanthomonas citri subsp. citri (Xac), is a one of the most important disease affecting citrus production worldwide and attacks a wide range of commercial species of citrus trees, causing significant losses in producing countries. Hamlin sweet orange (Citrus sinensis) is canker-sensitive, while Satsuma mandarin (Citrus unshiu) is canker-resistant. To understand the molecular mechanisms underlying the differences in responses to Xac, transcriptional profiles of these two genotypes following Xac attack were compared by RNA-Seq (Illumina). The purpose of this study was to examine simultaneous changes in gene expression profile during the early stages (24, 48 and 72 hpi) of citrus canker infection in Satsuma and Hamlin. A total of 5673 and 6231 up-regulated transcripts were identified at 24, 48 and 72 hpi in Satsuma and Hamlin, respectively, while 3982 and 7944 were down-regulated. Of these, 52 transcripts were up-regulated in common between both genotypes. These genes in common are related to basic defense against Xac, because there are genes involved in patogen perception and recognition, transcription factors and genes related to plant defense, such as glucanases and proteinases. Among up-regulated genes expressed only in Satsuma, aspartic proteinase was highlighted. This protein presented the highest gene expression 24 hpi and it can be involved in Satsuma resistance, since the expression of this protein was less pronounced and delayed in Hamlin. Another opposite response between these two genotypes was the expression of genes related to cell wall. Such genes were pectato lyase, extensin, cellulose sinthase, and xiloglucano endotransglycosilase. The genes were up-regulated in Satsuma, while in Hamlin, they were down-regulated. For genes related to plant defense, both genotypes up- regulated pathogenesis-related proteins, especially 72 hpi. However, the expression of these genes did not prevent the symptoms in ...

Orientador: Jesus Aparecido Ferro
Coorientador: Rui Pereira Leite Júnior
Banca: Alessandro de Mello Varani
Banca: José Belasque Júnior
Resumo: O cancro cítrico, causado pela bactéria Xanthomonas citri subsp. citri (Xac), é uma das principais doenças que acometem a citricultura mundial e ataca uma ampla gama de espécies comerciais de citros, causando perdas significativas nos países produtores. A espécie de laranja doce Hamlin (Citrus sinensis) é suscetível ao cancro cítrico, enquanto a espécie de tangerina Satsuma (Citrus unshiu) é resistente. Para compreender os mecanismos moleculares relacionados aos sistemas de defesa ativados pela planta é importante identificar as alterações transcricionais de cada espécie vegetal sob estresse fitopatogênico, a fim de desvendar os elementos moleculares que são específicos de cada espécie. O objetivo do presente trabalho foi realizar uma análise comparativa temporal do transcriptoma de duas espécies cítricas contrastantes em resposta à Xac, pela técnica do RNA-Seq (Illumina). Um total de 5.673 e 6.231 transcritos diferencialmente expressos foi induzido nos tempos 24, 48 e 72 horas após a inoculação de Xac em Satsuma e Hamlin, respectivamente, enquanto 3.982 e 7.944 transcritos foram reprimidos. Deste total, 52 transcritos foram induzidos em comum, nas duas espécies, em todos os tempos de inoculação. Estes genes estão relacionados com a defesa basal da planta contra o ataque de Xac, pois apresentam genes que participam na percepção e reconhecimento do patógeno, genes que codificam fatores de transcrição e genes que participam na defesa da planta, como glucanases e proteinases. Entre os genes induzidos exclusivamente na espécie resistente Satsuma destacou-se uma proteinase aspártica. Esta proteína apresentou a maior expressão gênica no tempo 24 horas e pode estar envolvida na resistência desta espécie, visto que na espécie suscetível Hamlin, a expressão desta proteína foi menos expressiva e tardia, no tempo 48 horas. Outra resposta oposta entre as espécie foi na expressão de genes relacionados à ...
Abstract: Citrus canker, caused by Xanthomonas citri subsp. citri (Xac), is a one of the most important disease affecting citrus production worldwide and attacks a wide range of commercial species of citrus trees, causing significant losses in producing countries. Hamlin sweet orange (Citrus sinensis) is canker-sensitive, while Satsuma mandarin (Citrus unshiu) is canker-resistant. To understand the molecular mechanisms underlying the differences in responses to Xac, transcriptional profiles of these two genotypes following Xac attack were compared by RNA-Seq (Illumina). The purpose of this study was to examine simultaneous changes in gene expression profile during the early stages (24, 48 and 72 hpi) of citrus canker infection in Satsuma and Hamlin. A total of 5673 and 6231 up-regulated transcripts were identified at 24, 48 and 72 hpi in Satsuma and Hamlin, respectively, while 3982 and 7944 were down-regulated. Of these, 52 transcripts were up-regulated in common between both genotypes. These genes in common are related to basic defense against Xac, because there are genes involved in patogen perception and recognition, transcription factors and genes related to plant defense, such as glucanases and proteinases. Among up-regulated genes expressed only in Satsuma, aspartic proteinase was highlighted. This protein presented the highest gene expression 24 hpi and it can be involved in Satsuma resistance, since the expression of this protein was less pronounced and delayed in Hamlin. Another opposite response between these two genotypes was the expression of genes related to cell wall. Such genes were pectato lyase, extensin, cellulose sinthase, and xiloglucano endotransglycosilase. The genes were up-regulated in Satsuma, while in Hamlin, they were down-regulated. For genes related to plant defense, both genotypes up- regulated pathogenesis-related proteins, especially 72 hpi. However, the expression of these genes did not prevent the symptoms in ...
Mestre

From a world prospective, the continuous application of chemical pesticides has serious long-term effects on human health and environmental pollution, and can result in resistant pathogen strains. However, postharvest diseases cause major losses on the markets and need to be controlled effectively. The search for biopesticides using microbial antagonists and natural plant products has subsequently become more important as viable alternatives to control postharvest diseases. Currently, little information exists in terms of citrus production practices, disease management measures and postharvest losses in Ethiopia. The aim of this study was therefore to determine what the current situation in the country is in terms of production, disease management and postharvest disease incidence, disease management practices in Ethiopia and to develop an effective and safe disease control strategy for the industry. Citrus production in Ethiopia is mainly done by Government enterprises with little technical expertise. Disease control strategies are ineffective with postharvest losses exceeding 46%. The most important postharvest pathogen identified was Penicillium digitatum. In development of biopesticides, three yeast antagonists [Cryptococcus laurentii (strain MeJtw 10-2 and strain TiL 4-3) and Candida sake (TiL 4-2)] and plant leaf extracts of Acacia seyal and Withania somnifera were found to have some potential to control Penicillium in in vitro and in vivo trials and ensure fruit quality. The modes of action of the yeast antagonists were not based on antibiosis. Instead, it involved competitive colonization where the antagonists inhibited P. digitatum spore germination and reduced mycelial growth by 75-100%. Extracts from the two plant species showed broad-spectrum antimicrobial activity against a range of several fungal and bacterial pathogens. The semi-commercial application of the antagonists and plant extracts improve fruit quality and the integration of these biopesticides were found effective in semi commercial trials and may provide a commercial solution for the citrus industry.
Thesis (PhD (Crop Protection))--University of Pretoria, 2007.
Microbiology and Plant Pathology
unrestricted

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)
Fundecitrus
A distribuição espacial do cancro cítrico em talhões com laranjeira ‘Natal’ enxertada em limoeiro ‘Cravo’ com 3 anos de idade foi estudada utilizando a geoestatística. Foram utilizados 13 talhões mapeados pelo Fundecitrus (Fundo de Defesa da Citricultura), com 595 ou 1080 plantas cada, infectados naturalmente com Xanthomonas citri subsp. citri, em uma única propriedade, no Estado de São Paulo. A severidade da doença em cada planta foi avaliada por meio de uma escala com os seguintes níveis: 0 - sem folhas com sintomas; 1 - de 1 a 5 folhas com sintomas; 2 - de 6 a 10 folhas com sintomas; 3 - de 11 a 20 folhas com sintomas; 4 - de 21 a 50 folhas com sintomas e 5 - > 50 folhas com sintomas. Nestes talhões, as coordenadas correspondentes à posição das plantas contaminadas foram obtidas com um GPS, permitindo mapear a doença no talhão. A distribuição espacial foi avaliada por ajuste de semivariogramas e interpolação dos dados por krigagem. A distribuição de plantas com cancro cítrico no talhão mostrou-se agregada, com raio de agregação de 30 a 45 m. Os mapas de krigagem mostraram que os focos da doença ocorreram mais frequentemente nos limites dos talhões. Quando a nota média de severidade da doença foi menor que 0,04, o semivariograma apresentou efeito pepita puro, indicando que não houve dependência espacial
The spatial distribution of citrus canker in areas with orange 'Natal' grafted on ‘Rangpur’ lime with three years of age were studied using geostatistics. We used 13 plots mapped by Fundecitrus (Citrus Defense Fund), with 595 or 1080 individual plants, naturally infected with Xanthomonas citri subsp. citri that were on the same property in the State of São Paulo. Disease severity in each plant was assessed using a scale with the following levels: 0 - no leaf with symptoms, 1 – 1 - 5 leaves with symptoms; 2 – 6 -10 leaves with symptoms; 3 – 11 - 20 leaves with symptoms; 4 – 21 - 50 leaves with symptoms and 5 - > 50 leaves with symptoms. In these plots, the coordinates corresponding to the position of symptomatic of infected plants was obtained with a GPS, allowing mapping the symptomatic plants in the block. The spatial distribution was evaluated by fitting of semivariograms and kriging interpolation of the data. The distribution of plants with citrus canker in the block was aggregated, with aggregate radius of 30 - 45 meters. Kriging maps showed that the foci of diseased plants stood on the edge of the blocks. When the average grade of severity was less than 0.04, the semivariogram showed pure nugget effect, indicating that no spatial dependence

Thesis (MScAgric (Horticulture))--University of Stellenbosch, 2010.
ENGLISH ABSTRACT: Creasing, also known as albedo breakdown, is a preharvest disorder that affects the albedo of citrus fruit causing creases on the surface of the fruit. It is a recurrent problem in Navel and Valencia oranges and can cause individual orchard losses which often exceed 50%. Although the contributing factors are known, the physiological basis of creasing development is unresolved and the current control measures do not prevent creasing satisfactory. Hence, better control measures and further understanding of the physiology of creasing development is required. The objective of this two-year study was to determine if the position of fruit in a tree, light and carbohydrate manipulation techniques, and albedo mineral nutrients influence creasing development. Furthermore, the most effective application timing of gibberellic acid (GA3) with the least negative effect on fruit rind colour development and the effectiveness of cytokinins, other products and different root biostimulants to reduce creasing incidence were evaluated. The position of fruit in the tree and light influenced the development of creasing and the distribution of mineral nutrients in the albedo. Creasing incidence was higher on the south side than on the north side of the tree and fruit from the inside sub-sectors had a greater creasing incidence compared to fruit from the outside sub-sectors. The shady part of outside fruit was more creased compared to the sunny part of the fruit and covering fruit with brown paper bags increased creasing severity. The light manipulation techniques used on the leaves and fruit increased the nitrogen (N), phosphorus (P), potassium (K) and manganese (Mn) concentrations in the albedo and differences in the albedo mineral nutrients amongst the sub-sectors evaluated were observed, but creasing severity or creasing incidence was not significantly correlated with the albedo mineral concentrations at harvest. Albedo mineral concentrations earlier in the season may play a role in creasing development, as creasing severity was significantly correlated with copper (Cu), K, and Mn concentrations in the albedo during stage II of fruit development. Creasing incidence and albedo mineral concentrations were not affected by any of the carbohydrate manipulation techniques used in this study. The incidence and severity of creasing was significantly reduced, with a minor negative effect on fruit rind colour development, by the application of GA3, from mid November to mid January. Localised fruit application of CPPU [N-(2-chloro-4-pyridyl)-N-phenylurea], MaxCel (6- Benzyladenine) and CPPU in combination with calcium after physiological fruit drop reduced the incidence and severity of creasing, although creasing incidence was not significantly different from the control. The application of Messenger®, AVG (aminoethoxyvinylglycine) and different root biostimulants did not reduce creasing incidence. The results showed that cytokinins could reduce creasing incidence and justify further studies on application and uptake efficiency. The use of different root biostimulants are not recommended, but it is suggested that treatment effects may be more pronounced over a longer period.
AFRIKAANSE OPSOMMING: Kraakskil is ‘n vooroes abnormalitiet wat die albedo van sitrusvrugte affekteer, deur krake op die oppervlak van vrugte te veroorsaak. Dit is ‘n algemene probleem in Navel en Valencia lemoene en kan boordverliese van tot 50% of soms hoër veroorsaak. Alhoewel die bydraende faktore bekend is, is die fisiologiese basis van kraakskil ontwikkeling onopgelos en die beskikbare beheermaatreëls is nie bevredigend nie. Dus, beter beheermaatreëls en ‘n beter begrip van die fisiologie van kraakskil ontwikkeling is nodig. Die doel van die twee-jaar studie was om te bepaal of die posisie van vrugte in ‘n boom, lig en koolhidraat manipulasie tegnieke en minerale elemente in die albedo, kraakskil ontwikkeling beïnvloed. Die mees effektiewe toedieningstyd van gibberelliensuur (GA3) sonder ‘n negatiewe effek op vrugkleur is bepaal en die effektiwiteit van sitokiniene, ander produkte en verskillende wortel biostimulante om kraakskil voorkoms te verminder, is geëvalueer. Die posisie van vrugte in ‘n boom en lig het kraakskil ontwikkeling en die verspreiding van minerale element in die albedo beïnvloed. Kraakskil voorkoms was hoër aan die suidekant van die boom as aan die noordekant en vrugte in die binnekant van die boom het ‘n groter kraakskil voorkoms as vrugte in die buitekant van die boom gehad. Die skadukant van buitevrugte het meer kraakskil gehad as die sonkant en die toemaak van vrugte met ‘n bruin papiersak het die graad van kraaksil verhoog. Die lig manipulasie tegnieke wat op die blare en vrugte gebruik is, het die stikstof (N), fosfaat (P), kalium (K) en mangaan (Mn) konsentasies in die albedo verhoog en verskille in die albedo minerale elemente tussen sub-sektore is waargeneem, maar betekenisvolle korrelasies is nie tussen die graad en voorkoms van kraakskil en die albedo minerale element konsentrasies by oestyd waargeneem nie. Albedo minerale element konsentrasies vroeër in die seisoen mag ‘n rol speel by kraakskil ontwikkeling, omdat die graad van kraakskil betekenisvol gekorreleer was met albedo koper (Cu), K, en Mn konsentrasies tydens fase II van vrugontwikkeling. Kraakskil voorkoms en albedo minerale element konsentrasies is nie deur enige van die koolhidraat manipulasie tegnieke geaffekteer nie. Die voorkoms en graad van kraakskil is betekenisvol verlaag, met ‘n geringe negatiewe effek op vrugkleur, deur die toediening van GA3 vanaf mid November tot mid Januarie. Gelokaliseerde vrugtoedienings van CPPU [N-(2-chloro-4-piridiel)-N-phenielureum], MaxCel (6- Bensieladenien) en CPPU saam met kalsium na fisiologiese vrugval het die voorkoms en graad van kraakskil verlaag, alhoewel kraakskil voorkoms nie betekenisvol van die kontrole verskil het nie. Die toediening van Messenger®, AVG (amino etoksievinielglisien) en veskillende wortel biostimulante het nie kraakskil voorkoms verlaag nie. Die resultate het getoon dat sitokiniene kraakskil voorkoms kan verlaag en verdere studies op die toediening en opname effektiwiteit word aanbeveel. Die gebruik van verskillende wortel biostimulante word nie aanbeveel nie, maar die effek behoort meer sigbaar te wees na ‘n langer periode van behandeling.

Contexte: Les pectines est sont des macromolécules complexes, composées essentiellement d 'homogalacturonanes et de rhamnogalacturonanes I. Ils représentent environ 90% des pectines dans la plupart des plantes. Les pectines peuvent être isolées des parois par divers procédés d'extraction et leurs caractéristiques chimiques, physicochimiques et structurales sont influencées non seulement par ces méthodes mais également par l'origine végétale de la matière première. Les pectines extraites ont d'abord été analysées de façon globale pour leur composition et leurs caractéristiques macromoléculaires. Dans un deuxième temps, les domaines pectiques homogalacturonanes et rhamnogalacturonanes I ont été spécifiquement isolés et purifiés. Cette approche visait à appréhender les différences structurales des pectines d'agrumes suivant leur mode d'extraction et la source végétale. Les proportions relatives des différents domaines pectiques, la longueur des domaines homogalacturonanes et la structure fine des domaines rhamnogalacturonanes ont été particulièrement étudiées. Matériel et méthodes: Des écorcés d'agrumes (orange, citron, lime et pamplemousse) ont été utilisées comme matière première. Extraction de la pectine: Des échantillons de pectine ont été obtenus dans une usine pilote ( extractions sévère et douce par l'acide nitrique et extraction sévère par l'acide oxalique) et au laboratoire (extraction douce par l'acide oxalique). La composition osidique et les caractéristiques macromoléculaires (masse molaire et viscosité intrinsèque) des pectines ont été étudiées. Isolement et purification des homogalacturonanes: des échantillons de pectine désestérifiée ont été soumis à une hydrolyse acide douce à différents intervalles de temps. Les caractéristiques macromoléculaires des fractions insolubles à l'acide représentant les domaines homogalacturonanes ont été analysées. . En outre, la composition osidique des fractions solubles à l'acide, représentant les sucres liés au rhamnogalacturonane I détachés de la pectine par des conditions acides, a été analysée. Isolement et purification des rhamnogalacturonanes I: Les domaines rhamnogalacturonane I ont été isolés à partir des écorces et à partir des pectines extraites. Le procédé d'isolement utilisé repose sur une dégradation alcaline des domaines homogalacturonane. Les fractions purifiées de rhamnogalacturonane I ont été caractérisées par leur composition monosaccharidique et par la nature des liaisons entre les oses. Principaux résultats: Le pH d'extraction et le type d'acide utilisé (nitrique ou oxalique) influencent la structure et les caractéristiques macromoléculaires des pectines. L'acide oxalique a permis d'extraire des pectines de masse molaire élevée. Les pectines extraites à l'acide oxalique (extraction douce) étaient enrichies en longues chaînes d'homogalacturonane et contenaient du rhamnogalacturonane I avec des chaînes latérales conservées. Les pectines extraites à l'acide nitrique avaient une faible masse molaire et contenaient des rhamnogalacturonanes comprenant de nombreuses chaînes latérales courtes. En terme de source végétale, les pectines de pamplemousse contenaient moins de chaînes latérales par rapport aux pectines d'orange, de lime et de citron. En outre, les pectines d'orange et de pamplemousse étaient particulièrement riches en rhamnogalacturonane I non ramifié. Après 24 h d'hydrolyse acide ménagée (conditions optimales), la répartition de l'acide galacturonique entre fraction insoluble à l'acide et fraction soluble à l'acide a été utilisée pour estimer la proportion entre homogalacturonane et rhamnogalacturonane I, respectivement. Les ratios varient de 70/30 à 95/05, selon la source d'agrumes et les conditions d'extraction des pectines. En ce qui concerne la caractérisation du rhamnogalacturonane I purifié, une dégradation des chaînes latérales contenant de l'arabinose a été observée pour les extractions par l'acide nitrique. L'analyse des liaisons glycosidiques à révélé la présence d'arabinanes et d'arabinogalactanes de type I et Il
Background: Pectin is a complex macromolecule, made mostly of homogalacturonan and rhamnogalacturonan I. Together, they represent approximately 90% of pectin in most plants. Pectins can be isolated from the walls by various extraction processes. Their chemical and physicochemical characteristics are influenced not only by these extraction methods, but also by the source of the raw material. Extracted pectin samples were first analyzed for their composition and macromolecular characteristics. Secondly, homogalacturonans and rhamnogalacturonans I were specifically isolated and purified. This approach aimed to understand the structural differences in citrus pectin due to extraction conditions and plant source. The relative proportions of different pectic domains, the length of homogalacturonans the fine structure of rhamnogalacturonans were particularly studied. Materials and methods: Citrus peel (orange, lemon, lime, and grapefruit) was used as raw material. Pectin extraction: Pectin samples were obtained on a pilot plant scale (harsh nitric acid, mild nitric acid, and harsh oxalic acid extraction) and on a laboratory scale (mild oxalic acid extraction). Pectin composition (acidic and neutral sugars) and physicochemical properties (molar mass and intrinsic viscosity) were investigated. Homogalacturonan isolation: De-esterified pectin samples were subjected to mild acid hydrolysis for different time intervals. Macromolecular characteristics (molecular weight and intrinsic viscosity) were analysed in acid-insoluble fractions, representing homogalacturonan domains. In addition, sugar compositions of acid- soluble fractions (representing rhamnogalacturonan I related sugars detached from pectin due to the acidic conditions) were analysed. Rhamnogalacturonan I isolation: Rhamnogalacturonan I domains were both isolated from peels and from extracted pectin samples. The isolation method was based on alkaline degradation of homogalacturonan. Purified rhamnogalacturonan I fractions were characterized by their monosaccharide compositions and linkage analyses. Key Results: The pH and extraction agent (nitric acid or oxalic acid) influenced the chemical structure and macromolecular characteristics. Oxalic acid allowed the recovery of pectin samples of high molecular weight. Mild oxalic acid-extracted pectins were rich in long homogalacturonan stretches and contained rhamnogalacturonan I stretches with conserved side chains. Nitric acid-extracted pectins had a low molecular weight and contained rhamnogalacturonan I stretches made of numerous short side chains. In terms of plant source, grapefruit pectin was found to have less side chains compared to orange, lime and lemon. Moreover, orange and grapefruit pectin samples were both particularly rich in rhamnogalacturonan I backbone. After 24 h of mild acid hydrolysis of pectin samples (defined as optimum conditions), the repartition of total galacturonic acid into acid-insoluble and acid-soluble fractions was used as a measure to estimate the proportion between pectic main domains, homogalacturonan and rharnnogalacturonan I, respectively. Ratios varied from 70/30 to 95/05, depending on citrus source and pectin extraction conditions. Regarding purified rhamnogalacturonan I characterisation, arabinose-containing side chains were deconstructed when nitric acid (mild or harsh) pectin extraction conditions were applied. Linkage analyses revealed the presence of arabinans and of type I and II arabinogalactans

6 pp.
Low Desert Citrus Varieties; Irrigating Citrus Trees
Diagnosing Home Citrus Problems includes information on all the commonly encountered dooryard citrus problems encountered in Arizona. Problems/disorders are grouped into three catagories: problems with fruit, problems with leaves, and problems with stems, branches and entire tree. Symptoms, causes and control measures are given for each disorder.

6 pp.
When choosing a variety of citrus to plant in your yard consider: what you like to eat; when you want to harvest; and how cold it gets in your yard. There are many varieties of species of citrus, each with its own characteristics. This publication lists the characteristics of some of the most popular varieties of citrus.

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Conselho Nacional de Desenvolvimento Científico e Tecnológico
The citrus blackfly (Aleurocanthus woglumi) an important pest of the citrus of Asian origin, was detected in Brazil in 2001. The aim of this research was to evaluate the biology and infestation levels of A. woglumi in three host plants (orange Pêra, C. sinensis, acid lime Tahiti, Citrus latifolia and mango, M. indica) and biometry evaluations. For this, experiments were assembled in the period of January to June of 2006 at laboratory conditions. It was verified that acid lime Tahiti had been shown as the most suitable host for A. woglumi. Orange Pêra and mango are similar, being the citrus genus the most favorable to the complete development of A. woglumi. Mango is an appropriate host for A. woglumi. The duration of embrionary development was 15 days on average for the three hosts. The fourth nymph stadium was the longest when compared to the other stages of evolutive cycle. The higher viabilities were on third nymph stages, eggs and second nymph stages. The duration of egg-adult cycle was 70 days on average (2 months and 10 days) for the three hosts evaluated.
A mosca-negra-dos-citros (Aleurocanthus woglumi Ashby) uma importante praga dos citros de origem asiática, foi detectada no Brasil em 2001. O objetivo deste estudo foi avaliar a biologia e os níveis de infestação de A. woglumi em três plantas hospedeiras (laranja Pêra, C. sinensis; lima ácida Tahiti, Citrus latifolia e manga, M. indica) e avaliações de biometria. Para isto, foram montados experimentos no período de janeiro a junho de 2006 em condições de laboratório. Foi verificado que lima ácida Tahiti se mostrou o hospedeiro mais favorável para A. woglumi. Laranja Pêra e manga assemelham-se; sendo que o gênero Citrus se mostrou mais favorável ao desenvolvimento completo de A. woglumi. Manga é um hospedeiro adequado para A. woglumi. A duração do desenvolvimento embrionário foi de 15 dias em média para os três hospedeiros. O estádio de ninfa 4 (pupário) foi o mais longo quando comparado com as outras fases do período desenvolvimento. As maiores viabilidades foram nas fases de ninfa 3, ovos e ninfa 2. A duração do ciclo ovo-adulto foi de 70 dias em média (2 meses e 10 dias) para os três hospedeiros avaliados.

Um dos grandes desafios da citricultura brasileira, como grande produtora de lima ácida \'Tahiti\', tem sido a conservação do fruto in natura com destino à exportação. A prevenção da mudança na coloração de verde para amarelo durante o armazenamento constitui o foco de grandes esforços na área de pesquisas em pós-colheita. O presente trabalho teve como objetivo: (1) avaliar a aplicação de concentrações de etileno visando determinar a concentração mínima necessária para ativar a mudança da cor na casca dos frutos; (2) avaliar o tratamento térmico por imersão em água quente, visando utilizá-lo como alternativa para retardar o amarelecimento. No capítulo 1 foram realizados dois experimentos, sendo avaliados 4 tratamentos no experimento 1: 0 μL L-1, 3 μL L-1 , 6 μL L-1 e 12 μL L-1 de etileno, armazenados por 18 dias a 22 ± 2 °C e 85 ± 5% de umidade relativa (UR), utilizando frutos beneficiados, mas não tratados com ácido giberélico (GA). Se observou baixa perda de massa fresca e uma aceleração no desverdecimento a partir do dia 10 nos tratamentos de 3, 6 e 12 μL L-1 de etileno. No experimento 2 foram avaliados seis tratamentos: 0, 1, 2, 3, 6 e 12 μL L-1 de etileno, armazenados por 21 dias a 25 ± 2 °C e 85 ± 5% de UR. Foram utilizados frutos beneficiados, e tratados com ácido giberélico (GA). Os resultados mostraram pouco desverdecimento segundo análise no colorímetro, sendo esse resultado mais perceptível na análise por nota de cor para concentrações de 3 μL L-1 de etileno e baixa perda de massa fresca. O beneficiamento com GA e cera pode ter afetado a ação do etileno. No Capítulo 2 foi conduzido um experimento utilizando frutos colhidos diretamente do campo e avaliado o tratamento térmico por imersão em água quente e posterior armazenamento em presença ou ausência de 3 μL L-1 de etileno. Foram estabelecidos 6 tratamentos: controle sem etileno, controle + 3 μL L-1 de etileno, 3 minutos a 50 ± 2 °C sem etileno, 3 minutos a 50 ± 2 °C + 3 μL L-1 de etileno, 5 minutos a 50 ± 2 °C sem etileno, e 5 minutos a 50 ± 2 °C + 3 μL L-1 de etileno. Os frutos foram armazenados por 30 dias a 25 ± 2 °C e 90 ± 5% UR. Foi observado que o tratamento hidrotérmico por 5 minutos a 50 ± 2 °C ajudou na manutenção da cor verde na casca dos frutos, também houve redução significativa na taxa respiratória e na perda de massa fresca dos frutos. Esses resultados sugerem que o tratamento térmico auxilia na manutenção da qualidade de frutos de lima ácida \'Tahiti\', uma vez que tem efeito no retardo de processos envolvidos no amadurecimento e senescencia.
One of the great challenges of the Brazilian citrus industry, as a major producer of \'Tahiti\' acid lime, has been the conservation of the fruit in natura with destiny to export. Preventing the change in color from green to yellow during storage is the focus of great efforts in post-harvest research. The objective of the present work was: (1) to evaluate the application of ethylene concentrations in order to determine the minimum concentration necessary to activate the color change in the fruit peels; (2) to evaluate the heat treatment by immersion in hot water, in order to use it as an alternative to delay the yellowing. In Chapter 1, two experiments were carried out and four treatments were evaluated in experiment 1: 0 μL L-1, 3 μL L-1, 6 μL L-1 and 12 μL L-1 of ethylene, stored for 18 days at 22 ± 2 ° C and 85 ± 5% of relative humidity (RH), using fruits benefited, but not treated with gibberellic acid (GA). It was observed low fresh mass loss and an acceleration in de-greening from day 10 in the treatments of 3, 6 and 12 μL L-1 of ethylene. In the experiment 2, six treatments were evaluated: 0, 1, 2, 3, 6 and 12 μL L-1 of ethylene, stored for 21 days at 25 ± 2 ° C and 85 ± 5% RH. The fruit was benefited and treated with gibberellic acid (GA). The results showed little de-greening according to the colorimeter analysis, being this result more noticeable in the analysis by color rating for concentrations of 3 μL L-1 of ethylene and low fresh mass loss. Processing with GA and wax may have affected the action of ethylene. In Chapter 2 an experiment was conducted using fruits harvested directly from the field and the heat treatment was evaluated by immersion in hot water and subsequent storage in the presence or absence of 3 μL L-1 of ethylene. Six treatments were performed: control without ethylene, control + 3 μL L-1 ethylene, 3 minutes at 50 ± 2 ° C without ethylene, 3 minutes at 50 ± 2 ° C + 3 μL L-1 ethylene, 5 minutes at 50 ± 2 ° C without ethylene, and 5 minutes at 50 ± 2 ° C + 3 μL L-1 ethylene. The fruits were stored for 30 days at 25 ± 2 ° C and 90 ± 5% RH. It was observed that the hydrothermal treatment for 5 minutes at 50 ± 2 ° C helped to maintain the green color in the fruits peel, also there was a significant reduction in the respiratory rate and the loss of fresh fruit mass. These results suggest that the hydrothermal treatment helps to maintain the quality of \'Tahiti\' acid lime fruits, since it has an effect on the delay of processes involved in ripening and senescence

Isolates of Citrus Tristeza Virus (CTV) in Belize may be causing inverse stem pitting, stunting, and slow decline on sweet and sour orange rootstock combination on field trees. One isolate on Cleopatra mandarin rootstock expressed no field symptoms but may be a possible source of a detrimental isolate to more susceptible citrus rootstock combinations. Field plants and field source inoculated indicator plants were used in this study. The first objective was to determine whether field trees of sweet Valencia orange trees that expressed symptoms of CTV would test positive to a polyclonal (mild and severe symptoms) and a monoclonal (severe decline) antibody. Of the 81 trees tested, 56% tested positive to a polyclonal antibody G-604. None of the trees tested positive to the monoclonal antibody MCA-13. The second objective was to determine the biological and serological reactivity on host indicator plants of some of the most symptomatic trees having highest optical density value readings from the ELISA test. Mexican lime showed mild to moderate vein clearing on three of the isolates tested, while sweet orange showed mild to moderate translucent veins on two of the isolates tested. Four isolates from the indicator host plants tested positive to polyclonal antibody G-604 but none tested positive to MCA-13. The third objective was to determine if there were any differences among the nucleotide sequences of three CTV isolates. The CTV coat protein gene (CPG) of isolates which reacted to a polyclonal antibody G-604 but not to monoclonal antibody (MCA-13) was selectively amplified by polymerase chain reaction from infected tissue extracts and sequenced using negative sense CN 407 primer. All three isolates revealed between ninety-eight and ninety percent significant identities to known CTV isolates previously characterized from citrus growing regions around the world.

A partir da década de 1950, o limoeiro \'Cravo\' (Citrus limonia Osbeck) passou a ser o principal porta-enxerto para citros no Estado de São Paulo. Devido à sua suscetibilidade ao declínio dos citros, a sua utilização diminuiu na década de 1980. Em 2001, com a constatação da doença morte súbita dos citros (MSC) na região norte e noroeste do Estado de São Paulo e Triângulo Mineiro, o emprego do limoeiro \'Cravo\' como porta-enxerto se tornou praticamente inviável, já que o mesmo se mostrou suscetível à MSC. A utilização de porta-enxertos tolerantes no cultivo de laranjeiras doces (C. sinensis (L.) Osbeck), até o momento, é a medida de manejo mais indicada para a MSC, porém, esses porta-enxertos apresentam baixa tolerância à seca, inviabilizando o cultivo sem irrigação em regiões que apresentam baixa disponibilidade hídrica no solo e altas temperaturas em determinadas épocas do ano. Com base nessas premissas, o objetivo do presente estudo foi determinar o desempenho horticultural de laranjeira \'Valência\' (C. sinensis (L.) Osbeck) sobre onze porta-enxertos na região norte do Estado de São Paulo, baseando-se nas características de produção e qualidade de fruto, desenvolvimento das plantas, tolerância à MSC e à seca. O experimento foi instalado no município de Colômbia, região norte do Estado de São Paulo. As diferentes combinações (copa / porta-enxerto) foram plantadas em março de 2007. Em abril de 2012, foram pré-selecionados combinações sobre onze porta-enxertos dentre os 42 que foram plantados originalmente. Para as diferentes combinações copa/porta-enxerto selecionadas, foram avaliadas as variáveis produção e qualidade dos frutos, desenvolvimento da planta, eficiência produtiva, taxa média de crescimento, tolerância à MSC (avaliação visual, detecção de Citrus sudden death-associated virus (CSDaV) e detecção do Citrus tristeza vírus (CTV)) e tolerância à seca (avaliação visual, potencial de água em folha e estimativa do índice de área foliar). Os resultados encontrados no presente estudo relatam a superioridade na produção de frutos de laranjeira \'Valencia\' quando enxertada em pelos menos um porta-enxerto pré-selecionado em relação ao limoeiro \'Cravo\' \"CNPMF 03\". Em relação às demais variáveis, também foi atribuído a duas combinações de copa/porta-enxerto qualidade superior dos frutos de laranja \'Valência\' e a outras três combinações tolerância satisfatória à seca comparadas aos clones de limoeiro \'Cravo\'. Foi observado aptidão para cultivo adensado de laranjeira \'Valência\' em dois porta-enxertos. Também foram encontradas diferenças em relação à presença do CSDaV nos diferentes porta-enxertos. Tangerina \'Sunki\' x P. trifoliata \'English\' e citrange \'C-13\' \"S\" apresentaram potencial para utilização como porta-enxertos para laranjeira \'Valência\' na região norte do Estado de São Paulo como alternativa ao limoeiro \'Cravo\'.
Since the 1950s, the Rangpur lime (Citrus limonia Osbeck) became the main rootstock for brazilian citrus industry in São Paulo State. Because of its susceptibility to citrus blight, the use of this rootstock decreased in the 1980s. In 2001, with the first report of citrus sudden death disease (CSD) in the north and northwest of São Paulo State and Minas Gerais Triangle, the use of Rangpur lime as a rootstock became practically impossible, because of its susceptibility to CSD. The use of tolerant rootstocks in sweet orange groves (Citrus sinensis (L.) Osbeck) yet is the most adequate management to CSD. However, these rootstocks have low tolerance to drought stress, limiting the citrus industry in areas with high temperatures at certain seasons of year, and with low water availability for irrigation. Based on these assumptions, the aim of this study was to determine the horticultural performance of \'Valencia\' sweet orange (C. sinensis (L.) Osbeck) onto eleven rootstocks in the northern region of São Paulo state, based on the characteristics of production and fruit quality, plant growth, tolerance to CSD and drought tolerance. The experiment was carried out in Colombia, north of the São Paulo State. The different combinations (canopy / rootstock) were planted in March 2007. In April 2012, eleven rootstocks were pre-selected from the 42 that were originally planted. For the different selected canopy / rootstock combinations, the following variables were evaluated: yield and fruit quality, plant development, yield efficiency, growth rate, tolerance to CSD (visual evaluation, detection of Citrus sudden death-associated virus (CSDaV) and detection of Citrus tristeza virus (CTV)) and drought tolerance (visual evaluation, leaf water potential and leaf area index). The results found in this study showed higher production quality of \'Valencia\' sweet orange fruits when grafted onto, at least, one pre-selected rootstock compared to \"CNPMF 03\" Rangpur lime. For the other variables, it was also attributed to two combinations scion / rootstock superior quality of \'Valencia\' sweet orange fruits and in the other three combinations satisfactory tolerance to water stress compared to Rangpur lime. It was observed aptitude for high density planting of \'Valencia\' sweet orange onto two rootstocks. There were also found differences regarding the presence of CSDaV on different rootstocks. The \'Sunki\' mandarin x English\' P. trifoliata and the \'C-13 \'\"S\" citrange showed potential to be used as rootstocks for \'Valencia\' sweet orange in the northern region of São Paulo state as an alternative to Rangpur lime.

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A citricultura é uma das principais atividades do agronegócio brasileiro, porém o aumento de doenças na última década tem causado grandes prejuízos a toda a cadeia produtiva. A doença cancro cítrico, causada pela bactéria Xanthomonas citri subsp. citri (Xac) é um grave problema para o setor, não havendo até o momento método eficaz para o seu controle. Nesse estudo, utilizando RNASeq, foram analisados os perfis transcricionais de dois genótipos hospedeiros contrastantes à doença: laranja doce Pêra-Rio (PR) moderadamente resistente, e Lima Ácida Galego (LG), altamente suscetível, 24, 48 e 72 horas após a infecção com Xac, no intuito de identificar genes das plantas envolvidos na interação patógeno-hospedeiro. Foram encontrados 6.330, 3.478 e 6.795 genes diferencialmente expressos (GDEs) na espécie moderadamente resistente PR, 24, 48 e 72 horas após a inoculação com Xac, respectivamente, quando comparados com seus controles. Na espécie altamente suscetível Limão Galego foram identificados 1.491, 5.621 e 2.145 GDEs após 24, 48 e 72 horas da inoculação com Xac, respectivamente. Através do programa Blast2GO, genes e vias metabólicas de fotossíntese, sinalização celular, síntese hormonais, fatores de transcrição, entre outros, foram encontrados. Esse estudo revelou diferenças associadas à resistência e desenvolvimento a nível molecular em PR e LG em resposta ao cancro cítrico, demonstrando que na espécie moderadamente resistente há uma maior ativação dos mecanismos de defesa. Tais estudos podem ser utilizados para o desenvolvimento de plantas de citros com adequado nível de resistência ao cancro cítrico
The citrus agribusiness is very important to the Brazilian economy, but the increase of diseases in the last decade has caused great economic losses to the sector. The citrus canker, caused by the bacterium Xanthomonas citri subsp. citri (Xac), is a serious disease that attacks all citrus species economically important worldwide and there is not an effective method for its control. In this study, RNASeq was used to analyze the transcriptional profiles of two contrasting citrus genotypes regarding citrus canker susceptibility: sweet orange Pêra Rio (PR), moderately resistant, and Mexican lime ‘Galego´ (ML), highly susceptible. Gene expression were performed in a HiScanSQ System (Illumina) using total RNA isolated from leaves collected 24, 48 and 72 hours after Xac inoculation, with leaves inoculated with water been used as control. It were found 6,330, 3,478 and 6,795 differentially expressed genes (DGEs) in moderately resistant PR specie at 24, 48 and 72 hours after Xac inoculation, respectively, when compared with their controls. In the specie highly susceptible ML, it was identified 1,491, 5,621 and 2,145 DGEs after 24, 48 and 72 hours after Xac inoculation, respectively. Through the program Blast2GO, genes and metabolic pathways related to photosynthesis, cell signaling, hormone synthesis, transcription factors, among others, were found as involved in plant defense. This study revealed differences at the molecular level between PR and ML in response to citrus canker, showing that in the specie moderately resistant there is a greater activation of host defense mechanisms. Such informations can be used for the development of citrus plants with an adequate level of resistance to citrus canker

Orientador: Jesus Aparecido Ferro
Coorientador: José Belasque Júnior
Banca: Rui Leite Pereira
Banca: Poliana Fernanda Gachetto
Resumo: A citricultura é uma das principais atividades do agronegócio brasileiro, porém o aumento de doenças na última década tem causado grandes prejuízos a toda a cadeia produtiva. A doença cancro cítrico, causada pela bactéria Xanthomonas citri subsp. citri (Xac) é um grave problema para o setor, não havendo até o momento método eficaz para o seu controle. Nesse estudo, utilizando RNASeq, foram analisados os perfis transcricionais de dois genótipos hospedeiros contrastantes à doença: laranja doce Pêra-Rio (PR) moderadamente resistente, e Lima Ácida Galego (LG), altamente suscetível, 24, 48 e 72 horas após a infecção com Xac, no intuito de identificar genes das plantas envolvidos na interação patógeno-hospedeiro. Foram encontrados 6.330, 3.478 e 6.795 genes diferencialmente expressos (GDEs) na espécie moderadamente resistente PR, 24, 48 e 72 horas após a inoculação com Xac, respectivamente, quando comparados com seus controles. Na espécie altamente suscetível Limão Galego foram identificados 1.491, 5.621 e 2.145 GDEs após 24, 48 e 72 horas da inoculação com Xac, respectivamente. Através do programa Blast2GO, genes e vias metabólicas de fotossíntese, sinalização celular, síntese hormonais, fatores de transcrição, entre outros, foram encontrados. Esse estudo revelou diferenças associadas à resistência e desenvolvimento a nível molecular em PR e LG em resposta ao cancro cítrico, demonstrando que na espécie moderadamente resistente há uma maior ativação dos mecanismos de defesa. Tais estudos podem ser utilizados para o desenvolvimento de plantas de citros com adequado nível de resistência ao cancro cítrico
Abstract: The citrus agribusiness is very important to the Brazilian economy, but the increase of diseases in the last decade has caused great economic losses to the sector. The citrus canker, caused by the bacterium Xanthomonas citri subsp. citri (Xac), is a serious disease that attacks all citrus species economically important worldwide and there is not an effective method for its control. In this study, RNASeq was used to analyze the transcriptional profiles of two contrasting citrus genotypes regarding citrus canker susceptibility: sweet orange Pêra Rio (PR), moderately resistant, and Mexican lime 'Galego' (ML), highly susceptible. Gene expression were performed in a HiScanSQ System (Illumina) using total RNA isolated from leaves collected 24, 48 and 72 hours after Xac inoculation, with leaves inoculated with water been used as control. It were found 6,330, 3,478 and 6,795 differentially expressed genes (DGEs) in moderately resistant PR specie at 24, 48 and 72 hours after Xac inoculation, respectively, when compared with their controls. In the specie highly susceptible ML, it was identified 1,491, 5,621 and 2,145 DGEs after 24, 48 and 72 hours after Xac inoculation, respectively. Through the program Blast2GO, genes and metabolic pathways related to photosynthesis, cell signaling, hormone synthesis, transcription factors, among others, were found as involved in plant defense. This study revealed differences at the molecular level between PR and ML in response to citrus canker, showing that in the specie moderately resistant there is a greater activation of host defense mechanisms. Such informations can be used for the development of citrus plants with an adequate level of resistance to citrus canker
Mestre

Atualmente os maiores gastos da citricultura concentram-se na sanidade do pomar. Os custos fitossanitários, juntamente com fertilizantes, ultrapassam 60% do custo da produção. O controle de doenças como a Morte Súbita dos Citros (MSC), Declínio e o Greening, também conhecida como Huanglongbing, são feitos de forma visuais os que conferem ao método um elevado grau de subjetividade e imprecisão. O presente trabalho teve por finalidade desenvolver uma metodologia rápida e segura de diagnóstico para as doenças acima citadas utilizando espectroscopia de fluorescência e fluorescência induzida por laser (FIL). Foram utilizados neste estudo dois porta-enxertos, Cravo e Cleópatra, e quatro variedades de copa, Pêra, Natal, Valência e Hamilim. As doenças estudadas também foram classificadas em diferentes níveis de severidade. O índice 1 foi utilizado para baixa severidade da MSC, e 2 para o estágio mais avançado da doença. Para o Greening fez-se uma distinção entre folhas sintomáticas e assintomáticas. A doença Declínio dos Citros também fez parte deste estudo em função dos sintomas serem muito parecidos aos da fase inicial de MSC, sendo este um problema sério para as avaliações de contaminação no campo. Inicialmente, foram realizados estudos a respeito da emissão de fluorescência dos pigmentos fotossintéticos provenientes dos extratos de folhas doentes e sadias através de um fluorímetro de bancada. Para as análises foliares utilizou-se um sistema portátil de espectroscopia de fluorescência induzida por laser desenvolvido no LOLEIA. O estudo dos extratos revelaram que as maiores variações ocorrem nas concentrações de clorofila a e b e que os pigmentos acessórios como ? e ?- caretonos não fluorescem no comprimento de onda estudado. De uma maneira geral, é possível dizer que plantas doentes sofrem uma diminuição das clorofilas a e b. Os resultados das folhas foram analisados por um método estatístico de Análises de Componentes Principais (PCA) para facilitar a observação das alterações espectrais. Através da metodologia desenvolvida foi possível a identificação e a distinção das doenças com nível de acerto superior a 93%. Contudo, estes resultados contribuíram para o desenvolvimento de uma metodologia para diagnóstico das doenças de citros de uma forma rápida, precisa e economicamente viável.
Currently the largest portion of money applied to citriculture focus on health of the orchard. The phytosanitary costs, along with fertilizers, exceed 60% of the production cost. The control of diseases such as Sudden Death of Citrus (SDC), Decline and Greening, also known as Huanglongbing, is done by visual inspection, which confers to the method a high level of subjectivity and imprecision. This work was intended to develop a quick and safe methodology of diagnosis for the diseases mentioned above using fluorescence spectroscopy and laser-induced fluorescence (LIF). It was used in this study two rootstocks, Cravo and Cleopatra, and four varieties of crown: Pera, Natal, Valencia and Hamlin. The diseases studied were also classified into different levels of severity. The index 1 was used for low severity SDC, and 2 for a more advanced stage of this disease. For Greening has been made a distinction between symptomatic and asymptomatic leaves. The Decline of Citrus disease was also part of this study because its symptoms are very similar to those in the initial phase of SDC, and this is a serious problem for the evaluation of contamination in the field. Initially, studies have been made on the fluorescence of photosynthetic pigments from the extracts of ill and healthy leaves using a non-portable fluorimeter. For the leaf analyses we used a portable system of laser-induced fluorescence spectroscopy developed in LOLEIA. The study of extracts revealed that the largest cariations occur in the concentrations of chlorophyll a and b, and that acessory pigments like ? and ?-carotones do not fluoresce in the wavelength used. In general, it is possible to say that ill plants have a decrease of chlorophylls a e b. The leaf results were analyzed through the statistical method Principal Component Analysis (PCA) in order to facilitate the observation of spectral changes. Through the developed methodology it was possible to identify and distinguish the diseases with efficiency greater than 93%. However, these results contributed for the development of a methodology for diagnosis of citrus diseases in a quick, accurate and economically viable way.

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O objetivo do presente estudo foi avaliar a influência da largura da área de controle de plantas daninhas sobre o desenvolvimento, produtividade e qualidade da laranjeira ‘Pera Rio’. O experimento foi conduzido em um pomar de três anos de idade, localizado no município de Monte Azul Paulista, SP, correspondendo a safra agrícola de 2015/2016. Os tratamentos constaram em larguras de áreas de controle ou convivência com as plantas daninhas, próximas à linha do plantio, e as plantas daninhas foram controladas por meio de capina manual ou herbicida. As larguras das áreas de controle ou convivência com as plantas daninhas foram faixas de 1,0, 2,0 e 3,0 metros; coroamento das plantas em raios de 0,5 e 1,0 metro e o correspondente à projeção da copa, e duas testemunhas: com e sem o controle das plantas daninhas, totalizando 12 tratamentos, em três repetições, dispostos em delineamento experimental em blocos casualizados. As variáveis foram: altura da planta, diâmetro da copa, comprimento do ramo, teor relativo de clorofila total, controle, produtividade e qualidade dos frutos. Os parâmetros de desenvolvimento vegetativo foram avaliados previamente à instalação do experimento e também aos 12 e 18 meses após a avaliação prévia; as notas de controle das plantas daninhas foram atribuídas aos 30 dias após a aplicação, em sete ocasiões, e ao final do ciclo produtivo avaliou-se a produtividade e a qualidade dos frutos. Os dados foram submetidos à análise de variância pelo teste F, sendo as médias comparadas pelo teste de Duncan a 10% de probabilidade. A área de controle influenciou negativamente sobre a parte vegetativa das plantas (exceto a altura da planta). Também influenciou sobre a produtividade, tendo os tratamentos testemunha com controle, o coroamento em um raio de 1,0 metro no limpo e as faixas no limpo de 1,0 e 3,0 metros proporcionados as maiores produtividades. O coroamento das plantas e a faixa tratada de no mínimo 1,0 metro proporcionaram frutos com maior massa, diâmetro (longitudinal e transversal), rendimento de suco e industrial. A convivência da cultura com as plantas daninhas interferiu negativamente no desenvolvimento, produtividade e qualidade dos frutos.
The objective of this study was to evaluate the influence of the width of the control area of weeds on the development, quality and productivity of the orange 'Pera Rio'. The experiment was conducted in an orchard of three years old, located in the municipality of Monte Azul Paulista, SP, corresponding to the season of 2015/2016. The treatments were in control areas widths or coexistence with weeds, near the line of planting, and weeds were controlled by hand weeding or herbicide. The widths of the areas of control or coexistence with the weeds were bands of 1,0; 2,0 and 3,0 meters; crowning of the radius in plant 0,5, 1,0 meters and corresponding to the projection of the crown, and two witnesses: with and without weed control, totaling 12 treatments in three replications in a randomized block . The variables evaluated were: plant height, canopy diameter, branch length, relative chlorophyll content, control, productivity and fruit quality. The vegetative growth parameters were prior evaluated to the experiment and also at 12 and 18 months after the previous evaluation; notes of weed control were assigned to 30 days after application, on seven occasions, and at the end of the production cycle evaluated the productivity and fruit quality. Data were submitted to analysis of variance by F test, the averages were compared by Duncan test at 10% probability. The area control negatively influenced on the vegetative parts of plants (except plant height). Also influenced on productivity, and the witness treatments control, the crowning achievement in a radius of 1.0 meters in clean and clean the tracks in 1.0 and 3.0 meters provided the highest yields. The crowning achievement of the plants and the treated range of at least 1.0 meters provided fruits with greater mass, diameter (longitudinal and transverse), juice yield and industrial. The coexistence of culture with weeds negative effect on the development, productivity and fruit quality.

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Present investigation was carried out to study the root morphology of one year old seedling and the influence of different rootstocks in pre-bearing Kinnow mandarin under field condition on plant morphological, physiological, tissue nutrients and biochemical parameters at the Division of Fruits and Horticultural Technology, I.A.R.I., New Delhi during 2014-15. Results of the findings revealed that the seedling rootstock of rough lemon had a better root morphology with respect to root length (1511.27cm), projected area (158.42cm2 ), surface area (482.00 cm2 ), root volume (12.24 cm3 ) and the number of forks (7026.80) with a comparatively inferior root morphology in Carrizo. Comparative performance of budded Kinnow on different rootstocks showed that trees on Jatti Khatti were superior in terms of stimulated increase in plant height (49.42%) and canopy volume (22.56%) during the period April-September, compared to minimum enhancement in Kinnow trees on Troyer citrange during the same period. Kinnow trees on different rootstocks maintained congenial relationship with the scion closest to 1.0. Leaf sclerophylly in terms of leaf area, fresh mass, dry matter and density of foliar tissue was higher in Jatti Khatti but leaves more succulent on rough lemon. Kinnow leaves on Karna Khatta and rough lemon maintained significantly higher relative water content (88.61%). However, chlorophyll fractions was higher in leaves on rough lemon with reduced chlorophyll fractions on Carrizo rootstock. Higher physiological activity in terms of photosynthetic rate, stomatal conductance, and internal CO2 was recorded in scion leaf of Kinnow on rough lemon during summer and during autumn and winter on Jatti Khatti and rough lemon. Leaf N and P content was higher in leaf tissue on sour orange while K, Ca and Mg content was higher in leaf tissue on rough lemon. Sodium accumulation in leaves was higher in Rangpur lime as compared to Troyer which accumulated minimum sodium. Leaf tissue of Sour orange excelled in micronutrients and maintained higher Fe, Zn, Mn and Cu in their leaves. Kinnow scions on rough lemon exhibited higher enzyme activities with respect to total soluble protein, 64 superoxide dismutase, peroxidase, and glutathione reductase activity, while catalase activity was significantly higher on sour orange rootstock. Proline accumulation was also observed to be higher on sour orange rootstock. Scion leaf on Troyer rootstock accumulated 1.88 fold more phenol. Polyphenol oxidase activity was observed higher in scion leaves on Jatti Khatti with minimum activity on Troyer and Carrizo citrange rootstock. From the findings of this study, it is concluded that rough lemon rootstock imparted higher metabolic activity in Kinnow scion, while sour orange accumulated more nutrients in their tissue. Carrizo proved to be an inferior rootstock with most of the characters studied.
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This thesis was aimed at the extraction, isolation and structure elucidation of insecticidally active secondary metabolites of Citrus aurantium plant parts for potential use either as commercial insecticides or as lead compounds. Fruits, leaves and shoots of C. aurantium were extracted in methanol and the chemicals recovered were fractionated by liquid-liquid partitioning using organic solvents of increasing polarity (petroleum ether, dichloromethane, ethyl acetate). Petri dish exposure bioassays revealed that only the petroleum ether fraction of the fruit methanol extract was toxic against Bactrocera oleae adults. The bioactive chemicals present in the fruits were produced and/or accumulated in the peels. Petri dish exposure and topical application bioassays revealed that a petroleum ether peel extract was toxic against B. oleae and Ceratitis capitata adults. In several cases, differences in susceptibility were revealed between the two species and between the two sexes of the same species. First gravity column fractionation of the peel extract revealed that only the F2 fraction was active, while fraction Ft has a synergistic effect. Further purification of the F2 fraction on the second gravity column, followed by HPLC, resulted in the isolation of three major compounds. The chemical structure of these components was elucidated by spectroscopic methods (UV, FTIR, GC-MS and *H NMR) and was assigned as osthol, bergapten and 6',7'- epoxybergamottin. Of the three isolated compounds, only the 6',7'-epoxybergamottin was active and its toxicity was the same to the synthesised 6',7'-epoxybergamottin. However, the synergism revealed between isolated bergapten and 6',7'- epoxybergamottin was not confirmed when isolated bergapten was replaced by synthetic bergapten, indicating that minor components present in the isolated bergapten were responsible for the synergistic effect. Crude or semi-purified peel extracts of C. aurantium may have potential for insect control. Moreover, 6',7'-epoxybergamottin has apparently never been reported to have deleterious effects on insects. At this early stage, the potential of 6',7'- epoxybergamottin as a lead compound for a new class of insecticides remains to be determined.

15 pp.
Originally published: 2000
This publication discusses some diseases that are sufficiently important to the citrus in Arizona. Topics include: -Parasitic Diseases -fungi / virus diseases / virus or virus-like diseases -Mycoplasma Diseases -Nematode Diseases -Nonparasitic Diseases

Citrus tristeza virus (CTV) ocorre em quase todas as áreas produtoras de citros do mundo. O controle da doença se baseia, principalmente, no uso de porta-enxertos tolerantes e na premunização das copas. A obtenção de copas de laranjas doces ou de porta-enxerto de laranja azeda transgênicos resistentes ao CTV permitiria retornar a um uso mais intensivo deste excelente porta-enxerto. Com isso, esse trabalho buscou avaliar linhagens transgênicas de laranja doce (Citrus sinensis) e obter plantas transgênicas de laranja azeda (Citrus aurantium) para a resistência ao CTV, a fim de oferecer uma outra alternativa para o controle desta doença em citros. Foram avaliadas plantas transgênicas de laranja doce cv. Valência e cv. Hamlin contendo três diferentes construções gênicas. Uma contendo uma sequência sense (684 pb) do gene da capa protéica do CTV (pCTV-CP), outra contendo uma sequência conservada (559 pb) do CTV (pCTV-SC) e uma do tipo hairpin, contendo sequências sense e antisense do gene da capa protéica separadas por um íntron (pCTV-dsCP). Dez linhagens transgênicas de cada construção gênica e de cada cultivar foram previamente confirmadas por análises de Southern blot e RT-PCR, totalizando 60 linhagens transgênicas. Tais linhagens foram clonadas e enxertadas sobre limão Cravo (C. limonia) e laranja azeda (C. aurantium), totalizando 360 plantas. Essas plantas, juntamente com plantas não transgênicas utilizadas como controle, foram inoculadas com o CTV por meio de Toxoptera citricida virulífero. As técnicas de ELISA indireto utilizando anticorpo monoclonal contra a capa protéica do CTV ou de Real-time PCR utilizando primers amplificadores dos genes p20 e p23 do genoma do CTV foram utilizadas para detectar o vírus nas plantas avaliadas, 4 semanas após as inoculações. Ocorreu variação na resistência ao vírus nas diferentes construções transgênicas utilizadas e entre clones de uma mesma planta. Alguns clones não foram infectados com o vírus mesmo após a quarta inoculação, indicando uma possível resistência ao patógeno. Um total de 30 experimentos de transformação genética de laranja azeda foram realizados, utilizando como explantes segmentos internodal, de epicótilo e de cotilédone associado ao hipocótilo. O teste de GUS permitiu a identificação de duas gemas com reação positiva (0,13% de eficiência de transformação). Tais gemas foram enxertadas in vitro sobre citrange Carrizo, mas apenas uma gema se desenvolveu. A planta obtida foi aclimatizada em casa-de-vegetação.
Citrus tristeza virus (CTV) occurs in almost all citrus-growing areas of the world. Control of citrus tristeza relies mainly on the use of tolerant rootstocks and scion cross protection. Obtaining transgenic sweet oranges cultivars or sour orange resistant to CTV would allow a better use of this excellent rootstock. This way, the aim of this work was to evaluate transgenic sweet orange (Citrus sinensis) lines and to obtain transgenic sour orange (Citrus aurantium) for the resistance to CTV, in order to offer another alternative for the control of the disease in citrus. Transgenic sweet orange cv. Valencia and cv. Hamlin containing three different genetic constructs were evaluated. One gene construct contains a sense sequence (684 pb) of the coat protein gene of CTV (pCTV-CP), another contains a conserved sequence (559 pb) of CTV (pCTV-SC), and the last one a hairpin type, containing the sense and antisense sequences of the coat protein gene separated by an intron (pCTV-dsCP). Ten transgenic lines of each gene construct and each cultivar were previously confirmed by Southern blot and RT-PCR analysis, totalizing 60 transgenic lines. These lines were cloned and grafted into C. limonia and into C. aurantium, totaling 360 plants. The plants, along with non-transgenic plants used as control, were challenged four times with the CTV by means of viruliferous Toxoptera citricida. Indirect ELISA using monoclonal antibody against the CTV coat protein or the Real-time PCR using primers to amplify the CTV genes p20 and p23 were used to detect the virus in the tested plants, 4 weeks after inoculation. Variation in the virus resistance was observed among different transgenic constructs and different clones of the same plant. Some clones were not infected with the virus even after the fourth inoculation, indicating a possible resistance to the pathogen. A total of 30 genetic transformation experiments of sour orange were performed, using as explants internodal segments, epicotyl segments and cotyledon fragment with hypocotyl attached. GUS reaction detected two shoots positive (transformation efficiency of 0,13%). These shoots were in vitro grafted in Carrizo citrange, but only one shoot developed. The plant obtained was acclimatized in greenhouse.

Citrus thrips populations (adults and nymphs) were monitored through the spring of 1991-1992 in several locations throughout most of the commercial citrus production areas in Yuma County to determine if citrus thrips seasonality was similar to that previously reported in California. Study findings indicate that seasonality is similar throughout the winter and very early spring. Adult thrips numbers increase rapidly in groves due to attractive foliage, whether it is weeds or citrus. High nymph numbers did not always follow adult peaks, and were not statistically correlated. Predatory mites and rains may have affected 1992 results.

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The populational floatation of the aphids Aphis spiraecola (Patch, 1914), Aphis gossypii (Glover, 1887), Toxoptera aurantii (Boyer de Foscolombe, 1841) and Toxoptera citricidus (Kirkald, 1907) was studied and related to climate variations from May of 2004 to April of 2005, through the use of Pearsons s Correlation Analysis. The sampling was carried out every two weeks, through Möericke trap. The highest populational densities of A. gossypii occurred in August and September of 2004, and in the latter the populational peak was verified. From July to September of 2004, it was verified the most intense incidences of T. aurantii, and the highest peak occurred in August of 2004. T citricidus did not present significative correlation with any of the studied climate factors, although higher populational densities in the months of May, June and August of 2004, and the second week of January, 2005, were observed. A. spiraecola predominated over the other species, with populational peaks from May to August of 2004, and in the third week of December of 2004. The populational peak of this specie occurred in August of 2004. It was obtained a positive statistical correlation among the populations of these aphids with thermic extent and the number of dry weather days. Negative correlation, in relation to relative humidity was observed only for A. gossypii. In the space between lines of the citrus culture, 22 infestating plants were identified, among which Annona coriacea M. (Anonaceae), host for T aurantii, Bidens pilosa L. (Asteraceae), host for A. spiraecola, and the infesting Emilia sonchifolia (L) DC (Asteraceae), Sonchus oleraceus L. (Asteraceae), Cassia sp (Leguminosae), Sida glaziovii K. Sch., Sida rhombfolia L., Sida urens L. (Malvaceae), Solanum americanum Mill and Solanum erianthum D. Don (Solanaceae) infested by A. gossypii, must be controlled because they are focuses of aphids that are detrimental to the culture. As for natural enemies, the parasitoid species found more often were the hymenopters of the Aphidiidae Family, Lysiphlebus testaceipes (Cresson, 1880) and Aphidius colemani (Viereck, 1912). In the predatory group, the Coccinellidae Family presented the largest number of species, being Hippodamia convergens (Guérrin-Meneville) the most frequent, followed by Cycloneda sanguinea (Linnaeus, 1763) and then by Scymnus sp, while Olla v-nigrun (Germ., 1824), Hyperaspis (Hyperaspis) festiva (Mulsant, 1850) and Hyperaspis sp. were found in smaller quantity during the sampling period. In the predatory group, the Chrysopidae were more abundante, being identified two species: Chrysoperla externa (Hagen, 1861), Ceraeochrysa cincta (Adams, 1982) and eight examples only identified as Leucochrysa (Nodita) sp. Among the Hemerobiidae, only the specie Nusalala tesselata (Gerstaecker, 1888) was observed in small quantities on the citrus leaves.
A flutuação populacional dos afídeos Aphis spiraecola (Patch, 1914), Aphis gossypii (Glover, 1887), Toxoptera aurantii (Boyer de Foscolombe, 1841) e Toxoptera citricidus (Kirkald, 1907) foi estudada, correlacionando-as com as variações climáticas no período de maio de 2004 a abril de 2005, através do uso da Análise de correlação de Pearson. As amostragens foram realizadas, quinzenalmente, através de armadilha Möericke. As maiores densidades populacionais de A. gossypii ocorreram em agosto e setembro de 2004, neste último, constatou-se o pico populacional. No período de julho a setembro de 2004 constatou-se as maiores incidências de T. aurantii, sendo o mês de agosto de 2004 o de maior pico populacional. T. citricidus não apresentou correlação significativa com nenhum dos fatores climáticos estudados, embora tenham sido observados maiores densidades populacionais nos meses de maio, junho e agosto de 2004, e segunda semana de janeiro de 2005. A. spiraecola predominou sobre as demais espécies com picos populacionais nos períodos de maio a agosto de 2004 e na terceira semana de dezembro de 2004. O período de pico populacional desta espécie foi agosto de 2004. Foi obtida correlação estatística positiva entre as populações desses afídeos com a amplitude térmica e o número de dias de estiagem. Correlação negativa, em relação à umidade relativa, foi observada somente para A. gossypii. Foram identificadas nas entrelinhas da cultura de citros, 22 plantas infestantes, das quais Annona coriacea M. (Anonaceae), hospedeira de T. aurantii, Bidens pilosa L. (Asteraceae), hospedeira de A. spiraecola, e as infestantes Emilia sonchifolia (L.) DC (Asteraceae), Sonchus oleraceus L. (Asteraceae), Cassia sp (Leguminosae), Sida glaziovii K. Sch., Sida rhombifolia L., Sida urens L. (Malvaceae), Solanum iv 5 americanum Mill e Solanum erianthum D. Don (Solanaceae) infestadas por A. gossypii, devem ser controladas, pois são focos de afídeos prejudiciais a cultura. Com relação aos inimigos naturais, as espécies de parasitóides mais encontradas, foram os hymenópteros da Família Aphidiidae, Lysiphlebus testaceipes (Cresson, 1880) e Aphidius colemani Viereck, 1912. Do grupo dos predadores a Família Coccinellidae apresentou o maior número de espécies, sendo que Hippodamia convergens (Guérrin-Meneville) foi à espécie mais freqüente, seguida por Cycloneda sanguinea (Linnaeus, 1763) e, posteriormente, Scymnus sp, enquanto que Olla v- nigrum (Germ.; 1824), Hyperaspis (Hyperaspis) festiva Mulsant, 1850 e Hyperaspis sp., foram encontradas em menor quantidade durante o período de amostragens. Entre o grupo dos predadores, os crisopídeos foram os mais abundantes, sendo identificadas duas espécies: Chrysoperla externa (Hagen, 1861), Ceraeochrysa cincta Adams, 1982 e oito exemplares identificados apenas como Leucochrysa (Nodita) sp. Entre os Hemerobiideos, apenas a espécie Nusalala tesselata (Gerstaecker, 1888) foi observada em pequenas quantidades nas folhas de citros.

O cancro cítrico, causado por Xanthomonas axonopodis pv. citri (Xac), é um dos mais graves problemas fitossanitário da citricultura brasileira. Com a introdução da lagarta minadora dos citros (Phyllocnistis citrella [LMC]), houve um aumento drástico do número de focos do cancro cítrico, além da mudança do padrão espacial de fortemente agregado para moderadamente agregado e ao acaso. Foram avaliados: (1) suscetibilidade de três condições foliares: folha intacta, ferimento mecânico e injúria de LMC nas fases ovo, 1º ínstar, 3º ínstar e pupa, inoculados em diferentes concentrações de Xac (101, 102, 104 e 106 ufc/ml); (2) período de suscetibilidade da folha intacta, do ferimento mecânico e injúria de LMC inoculados a 106 ufc/ml; e (3) efeito da temperatura (12, 15, 20, 25, 30, 35, 40 e 42°C) e duração do molhamento foliar (0, 4, 8, 12, 16, 20 e 24 h) em folhas sem ferimento inoculadas a 106 ufc/ml. A concentração mínima de inóculo para causar sintomas da doença em folha intacta, injúria de LMC fase ovo e 1º ínstar foi de 104 ufc/ml; em ferimento mecânico e injúria de LMC fase 3º ínstar e pupa, 102 ufc/ml. Injúria da fase pupa resultou em grande severidade do que nos demais tratamentos a 106 ufc/ml (2 vezes maior do que em folha intacta).O período de suscetibilidade da injúria de LMC foi seis vezes mais longo do que do ferimento mecânico e a máxima suscetibilidade foi três vezes maior que em folha intacta. A LMC está relacionada com o aumento do dano que expõem mesofilo foliar à infecção direta de Xac, aumentando a suscetibilidade da folha por longo período. A faixa de temperatura ótima para o desenvolvimento do cancro cítrico foi de 25-35ºC. Entretanto, a severidade foi mais alta entre 30-35ºC. Sintomas de cancro não desenvolveram a 42ºC e na ausência de molhamento foliar. O efeito da temperatura foi maior do que a duração do molhamento foliar. A mínima duração de molhamento para infecção de Xac foi menor que 4 horas.
Asiatic citrus canker, caused by X. axonopodispv.citri (Xac), is one of the most serious phytosanitary problems in Brazilian citrus crops.The introduction of the citrus leafminer (Phyllocnistis citrella [CLM]), has resulted in an increase in the number of disease foci and has changed the spatial pattern of citrus canker symptomatic trees from strong aggregation to intermediate aggregation and random patterns. We evaluated: (1) susceptibility of three distinct leaf condition: intact leaves, mechanical wounding, and injury caused by CLM at the egg stage, 1st instar, 3rd instar, and pupal stage, inoculated at different concentration of Xac (101, 102, 104, and 106 cfu/ml); (2) susceptibility period of intact leaves, mechanical wounding, and CLM injury inoculated at 106 ufc/ml; and (3) effect of temperature (12, 15, 20, 25, 30, 35, 40 and 42°C) and leaf wetness durations (0, 4, 8, 12, 16, 20, and 24 h) on unwounded leaves inoculated at 106 ufc/ml. The minimum inoculum concentration to cause symptom development in intact leaves, CLM injury at the egg stage and 1st instar was 104 cfu/ml; in mechanically wounded leaves and CLM injuries at the 3rd instar and pupa stage, 102 cfu/ml. The injuries from pupa stage resulted in greater disease severity than other treatments at 106 cfu/ml (two times higher than in the intact leaf). Susceptibility period of CLM injury was six time longer than mechanical wounding and maximum observed susceptibility was three times higher than intact leaf. CLM is related to the amount of damage that exposes mesophyll cells to direct Xac infection, increasing the susceptibility of leaf for long period. Optimal temperature for development of citrus canker was 25–35°C. However, disease severity was highest in the range 30–35°C. Symptoms did not develop at 42°C and zero hour of leaf wetness duration. Effect of temperature was .greater than leaf wetness duration. The minimum wetness duration for Xac infection was as short as 4 h.

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Seis cultivares de laranjeiras doces (Sanguínea, Olivelands, Vaccaro Blood, Folha Murcha, São Miguel e Finike) foram avaliadas no campo em quatro experimentos e cada experimento sobre um porta-enxerto (limoeiro ‘Cravo’, citrumeleiro ‘Swingle’, tangerineiras ‘Sunki’ e ‘Cleópatra’). Durante a safra 2005/2006, foram avaliadas quanto às características que convencionalmente são atribuídas à qualidade dos frutos (massa, altura, diâmetro, rendimento de suco, teor de sólidos solúveis, acidez titulável e ratio). Na safra 2006/2007, foram quantificados os danos provocados pela Clorose Variegada dos Citros e, em ambas, quanto à produção de frutos e de sólidos solúveis por árvore. Na ocasião da colheita de cada cultivar, seus frutos mostraram-se semelhantes quanto às características químicas. As cultivares Olivelands e Finike destacaram-se em relação à produção de frutos, porém a ‘Olivelands’ se destacou também pela maior produção de sólidos solúveis por planta. Quanto a reação à CVC, houve diferenças entre as cultivares quanto à intensidade de doença e na produção de frutos de plantas sadias em comparação com as doentes, nos dois anos de avaliação. ‘Olivelands’ e ‘Finike’ apresentaram baixos índices de doença e pequena diferença na produção de frutos e sólidos entre suas plantas sadias e doentes, mostrando maior tolerância à infecção por Xilella fastidiosa.
Six sweet orange cultivars (Sanguínea, Olivelands, Vaccaro Blood, Folha Murcha, San Miguel and Finike) were evaluated in four experiments. Each budded on a rootstock (Rangpur lime, ‘Cleopatra’ and ‘Sunki’ mandarins and ‘Swingle’ citrumelo). Over 2005/2006 season, the characteristics that are conventionally attributed to the fruits quality (weight, height, diameter, yield of juice, soluble solids, acidity and ratio) were evaluated. In the 2006/2007, the damage caused by Citrus Variegated Chlorosis, and in both, as the fruit and soluble solids yield per tree were quantified. At the each cultivar harvest, its fruits have shown similar chemical characteristics. 'Olivelands' and 'Finike' highlighted up to the fruit yield, but the 'Olivelands' stood out also by the increased production of soluble solids per tree. In reaction to Xilella fastidiosa infection, there were differences to the cultivars on the disease intensity and the fruit yield by healthy trees compared with patients in two years of evaluation. 'Olivelands' and 'Finike' showed low rates of disease and small difference in the fruit and soluble solids yield to their plants healthy and sick, showing greater tolerance to Xilella fastidiosa infection.

Includes bibliographical references (leaves 152-164) Microscopy investigations into the oil glands, localisation of the rind oils and the development of oleocellosis have been carried out in Washington navel orange (Citrus sinensis [L.] Osbeck). Aims to develop an improved understanding of the physiological basis of the rind disorder.

A citricultura paulista está estabelecida, em sua maioria, em solos com baixos teores de matéria orgânica e, em decorrência deste fato, diversos aspectos associados à carência de nutrientes têm sido atribuídos à baixa disponibilidade da matéria orgânica. Para otimização de sistemas agrícolas que priorizem a sustentabilidade recomenda-se, por exemplo, o uso de cobertura morta (palhada), o incremento de matéria orgânica e a melhoria da fertilidade química física e biológica do solo, resultando em aumento de produtividade, maior estabilidade das produções e melhor disponibilidade de água para as plantas. O presente trabalho estudou em condições de campo, durante quatro safras, os efeitos da adição de cobertura morta ("mulch") - 0, 15 e 30 t.ha-1.ano-1 usando a palhada de gramínea Tifton como fonte de cobertura morta e ácido bórico aplicado via solo, nas doses de 0, 6 e 12 kg.ha-1 como fonte de boro. Foram avaliados, além da produtividade (t.ha-1 e número de frutos.ha-1), os aspectos associados à fertilidade de solo, aspectos físicos e nutricionais associados às variações dos tratamentos nos atributos hídricos do solo, a detecção da presença de carbono da palha no solo por metodologia isotópica do δ13c e δ15N, a atividade da biomassa microbiana, a influência dos tratamentos na qualidade do suco e aspectos econômicos quanto ao uso de palhada e boro no sistema produtivo. O experimento foi conduzido no município de Araras-SP, em pomar de laranja 'Pêra', com 9 tratamentos – 3 doses de palhada e 3 doses de boro, 4 repetições e 4 plantas por parcela, totalizando 144 plantas em um pomar comercial de 8 anos. A aplicação de 15 t.ha-1 e 30 t.ha-1 de palhada proporcionaram um aumento de 3,38 t.ha-1 (15,6%) e 8,89 t.ha-1 (41,1%) respectivamente. Em ambos os casos, a aplicação foi inviável economicamente, pois o custo da aplicação da cobertura morta foi maior que a receita do incremento de produtividade. Para o número de frutos.ha-1 a aplicação de palhada incrementou 9,3% e 24,4% para os tratamentos de 15 t.ha-1 e 30 t.ha-1 respectivamente. Na camada de 0 a 20 cm de profundidade, a biomassa microbiana foi ativada na dose de 30 t.ha-1 de palhada, sendo 64,7% superior à testemunha e 46,9% superior à dose de 15 t.ha-1. A aplicação de palhada de gramínea aumentou a participação de carbono de planta C4 na matéria orgânica do solo, sendo que a participação foi de 44%, 54,8% e 60,9%, para as doses de 0, 15 e 30 t.ha-1 respectivamente, mas a aplicação de palhada não afetou as características hídricas do solo nas profundidades de 20 e 40 cm. O efeito da aplicação de boro foi observado somente em ano de alta produtividade, acima de 40 t.ha-1, com a dose de 12 kg.ha-1 produzindo 5,8 t.ha-1 (16,7%) a mais que a testemunha. A relação foliar K/Ca e o teor de foliar de enxofre afetaram negativamente a produtividade, pois para cada unidade que aumenta na relação K/Ca houve diminuição de 30 t. 30 t-1 e para cada unidade de aumento do teor foliar de enxofre ocorreu redução de 4 t.ha-1. Recomenda-se que a relação K/Ca deve estar em torno de 0,4 e o teor foliar de enxofre em torno de 3 g.kg-1, já a relação P/S afetou positivamente a produtividade, para cada unidade que aumenta nesta relação, ocorreu um aumento de 17 t.ha-1. O aspecto ano é um fator importante, onde o efeito da alternância bienal de alta/baixa safra ficou evidente.
The citrus production in the state of São Paulo has been established in its majority, on soils with low organic matter and due to this fact, several aspects related to the lack of nutrients have been attributed to the low availability of organic matter. To introduce a sustainable management for the citrus production system, the producers have been adopted the use of mulch to cover the plants. This practice has proved to increase the soil organic matter content and has been recommended for mid- and long-term improvement of the physical and chemical and biological parameters of the soil, resulting in higher productivity, a better stability in crop production and higher water availability to the citrus plants. This work studied for 4 citrus growing seasons at field conditions the effects of the addition of mulch at rates of 0, 15 and 30 t.ha-1 added every year, using Tifton as source of mulch and boric acidic added to the soil at rates of 0, 6 and 12 kg.ha-1. Measures included yield (t.ha-1 and fruit number.ha-1), the soil nutrient composition, nutritional parameters associated to the physical and chemical variations in the soil due to the treatments, the isotopic δ13c carbon and δ15N nitrogen variation in the soil due to straw addition, the soil microbial biomass, the fruit (juice) quality and the economic aspects involving the use of these two practices – mulch and boron, in the citrus production system. The experiment was set up in Araras, São Paulo State, in a 8-year old commercial orchard of sweet orange var. Pêra, with 9 treatments – 3 rates of mulch and 3 rates of boron, 4 replicates with 4 plants for plot, in a total of 144 plants. The application of 15 t.ha-1 and 30 t.ha-1 of mulch increased 3,38 t.ha-1 (15,6%) and 8,89 t.ha-1 (41.1%) respectively, in both cases the use of mulch showed not to be economically viable, due to its high cost in the market. The fruit productivity from the mulch treatments increased 9.3% and 24.4%, in treatments with mulch added at the rates 15 t.ha-1 and 30 t.ha-1, respectively. At 0-20 cm soil depth, the microbial biomass increased when 30 t.ha-1 mulch was added, with 64.7% and 46.9%, corresponding to 0 t/ha and 15 t.ha-1 mulch, respectively. The use of Tifton grass as mulch was positively related to increases of carbon from C4-plants in the soil organic matter content, with increases after 4 years of growth as high as 44.4%, 54.8% and 60.9%, corresponding to the rates of 0, 15 and 30 t.ha-1 of mulch. Nevertheless, the addition of mulch has not affected the water content of the soil at 20 and 40 cm depth. The effect of boron was observed only at the high productivity season (more 40 t.ha-1), with the treatment of 12 kg t.ha-1 yielding 5.8 t.ha-1 (16.7%), when compared to the control (0 kg/ha B) treatment. The K/Ca ratio and the S-leaf concentration were negatively correlated with yield, when increases of one unit in K/Ca ratio corresponded to a decrease of 30 t.ha-1 and one unit of S-leaf concentration to minus 4 t.ha-1. It is recommended that K/Ca ratio should be around 0.4 and the S-leaf concentration around 3 g/kg. The leaf ratio P/S affected positively the yield, when increases of one unit in this ratio, corresponded to increases of 17 t.ha-1 in yield increases. The season (year) was an important factor, showing an alternate variation of high/low yield in every two years.

A transformação genética possibilita a introdução de genes de interesse agronômico no genoma das plantas e pode ser empregada na tentativa de obter plantas resistentes a doenças. No entanto, para se obter uma planta transgênica é necessário primeiramente estabelecer um procotolo eficiente de regeneração de plantas in vitro. Assim, o objetivo desse trabalho foi estudar a organogênese in vitro e a transformação genética de limão Volkameriano e laranja azeda com um fragmento do gene da capa protéica do CTV. Para a organogênese in vitro utilizou-se, como explante, segmento internodal, obtido de planta cultivada em casa-de-vegetação, segmento de epicótilo, coletado de plântula cultivada in vitro e segmento de cotilédone associado ao hipocótilo obtido de semente introduzida in vitro. Esses explantes foram mantidos em meio de cultura EME suplementado com 6-benzilaminopurina (BAP 0,0; 0,5; 1,0; 1,5; 2,0 mg L- 1), sendo incubados sob fotoperíodo de 16 h de luz ou em condições de escuro por 30 dias e então transferidos para fotoperíodo de 16 h de luz. A avaliação foi realizada após 45 dias de cultivo, determinando-se o número de explantes responsivos e o número de gemas por explante. A caracterização anatômica do processo de regeneração foi realizada por meio de cortes histológicos. Pela análise dos dados foi possível verificar que a organogênese in vitro ocorreu a partir dos três tipos de explantes testados, sendo que, nas duas espécies em estudo, os melhores resultados foram obtidos com o cultivo de segmento de cotilédone associado ao hipocótilo. As concentrações de BAP que estimularam as melhores taxas de regeneração foram de 1,0 e 1,5 mg L-1, para limão Volkameriano, e 0,5 e 1,0 mg L-1 para laranja azeda. A incubação dos explantes em ausência de luz favoreceu a regeneração in vitro. Pela análise histológica foi possível observar que o processo de regeneração, a partir dos três tipos de explantes testados, ocorreu por meio de organogênese indireta. O protocolo de desenvolvimento estabelecido durante os experimentos de organogênese in vitro foi utilizado para a transformação genética dessas espécies via Agrobacterium, contendo o plasmídeo pCAMBIA 2201, com um fragmento do gene da capa protéida do CTV, em uma construção gênica do tipo hairpin. As gemas de limão Volkameriano e laranja azeda identificadas como transgênicas pelo teste histoquímico GUS foram enxertadas in vitro em citrange Carrizo. A confirmação da transformação genética foi realizada pela análise de PCR, a qual mostrou a amplificação de um fragmento de 671 pb, correspondente a parte do gene amplificada.
Genetic transformation permits the introduction of agronomically important genes in plant genome and can be utilized in order to produce disease resistant plants. However for the recovery of transgenic plants is required to establish an efficient in vitro plant regeneration protocol. In this work the aim was to study an in vitro organogenesis and the genetic transformation of Volkamer lemon and sour orange with a sequence of the CTV coat protein gene. For in vitro organogenesis explant internodal segments collected from plants cultivated in greenhouse, epicotyl segments obtained from in vitro cultivated seedlings and cotyledon fragment with hypocotyl attached obtained from in vitro germinated seed were used as explant. These explants were cultured in EME medium supplemented with benzilaminopurine (BAP 0,0; 0,5; 1,0; 1,5; 2,0 mg L-1). Cultures were maintained under a 16 h photoperiod or in the dark for 10 d and then transferred to a 16 h photoperiod. The evaluation was performed 45 d after the incubation determining the number of responsive explant and the number of buds per explant. The anatomical characterization of in vitro regeneration process was carried out through histological analyses. The in vitro organogenesis occurred in the three types of explant tested, however cotyledon fragment with hypocotyl attached showed higher morphogenetic potential in both species. The best responses of regeneration were obtained when the medium was supplementation with 1,0 e 1,5 mg L-1 BAP for the Volkamer lemon and 0,5 e 1,0 mg L-1 BAP for the sour orange. The incubation in darkness favored the in vitro regeneration. The histological analyses showed that the regeneration process occurred through indirect organogenesis in the three types of explants tested. The developed protocol was use for genetic transformation of Volkamer lemon and sour orange with Agrobacterium, containing pCAMBIA 2201 plasmid with a sequence of CTV coat protein gene, in a hairpin construction. Volkamer lemon and sour orange shoots identified as transgenic by histochemical test GUS were micrografted into Carrizo citrange. PCR analysis were performed after micrografted showing the presence of the 671 pb fragment of the transgene.

A embriogênese somática é uma técnica alternativa com potencial aplicação na propagação clonal de plantas, além de ser uma excelente ferramenta para estudos básicos e análise dos eventos moleculares e bioquímicos que ocorrem durante a embriogênese vegetal. Contudo, apesar de várias pesquisas relativas a embriogênese somática, a falta de conhecimento sobre os fatores que controlam o fenômeno, comprova que existem ainda muitos pontos a serem entendidos sobre o processo. Deste modo, o presente estudo foi concebido com o intuito de estudar a embriogênese somática de duas espécies de citros, as quais apresentam diferentes graus de eficiência na produção de embriões somáticos. Inicialmente, buscou-se avaliar comparativamente o processo de embriogênese somática de duas espécies de citros, observando-se as diferenças estruturais através de análises histológicas e histoquímicas, e as diferenças na expressão do gene AtSERK1 nos calos, através de hibridização in situ. Para instalação dos experimentos, foram utilizados calos provenientes de nucelos de duas espécies, Citrus sinensis L. Osbeck, variedade Valência, e Citrus limonia Osbeck, variedade limão ‘Cravo’. Amostras de calos em meio de multiplicação, em meio de obtenção de embriões, e embriões somáticos nas diferentes fases de desenvolvimento foram coletados para observações anatômicas e histoquímicas através de microscopia óptica e realização de estudo da expressão gênica através de hibridização in situ. Com relação à morfologia, os principais resultados obtidos demonstraram que existem diferenças anatômicas e histoquímicas entre calos de limão ‘Cravo’ e ‘Valência’. Em relação à expressão gênica, os resultados evidenciaram a expressão do gene AtSERK1 em calos das duas espécies de citros, tanto em meio de multiplicação, com sacarose, como em meio de indução, com maltose, indicando que o potencial embriogênico já está instalado no calo em meio de multiplicação. Pode-se concluir também que este gene é conservado entre Arabidopsis e Citrus. Desenvolveram-se, também, experimentos para estudar os efeitos de alguns tratamentos (frio, dessecação, agrupamentos celulares) na otimização da indução e sincronização da embriogênese somática de C. sinensis, var. Valência. Neste sentido, calos nucelares de laranja ‘Valência’, procedentes do meio de multiplicação foram mantidos no mesmo meio líquido em agitador orbital a 200 rpm durante 24h e posteriormente peneirados de acordo com os tratamentos. As peneiras utilizadas apresentaram malhas de 150 µm (P1), 300 µm (P2) e 600 µm (P3). Além do efeito das peneiras também foram observados os efeitos de exposição ao frio (4 o C por quatro semanas, ausência de luz) e dessecação (27 o C, em placas de Petri na ausência de meio de cultura, seis dias, ausência de luz), sendo posteriormente mantidos em B.O.D., a 26 ± 1 o C e intensidade luminosa de 300 lux. Os resultados obtidos permitiram concluir que em relação ao desenvolvimento e produção de embriões somáticos o fator peneira foi superior ao fator estresse na freqüência embriogênica. O desenvolvimento de embriões somáticos em Citrus sinensis ocorre mais eficientemente a partir de agrupamentos celulares de tamanhos específicos.
Somatic embryogenesis is an alternative technique with potential application for clonal propagation of plants, and an excellent tool for basic studies and analysis of the molecular and biochemical events that occur during embryogenesis. However, although many studies have been done, the factors that control somatic embryogenesis are not completely understood. Thus, the present study proposes to evaluate aspects of somatic embryogenesis of two citrus species, which differ in efficiency for the production of somatic embryos. Initially, a comparison of the embryogenic process was done in terms of structural and histochemical analysis and evaluation of the expression of the gene AtSERK1 in callus cultures through in situ hybridization. For the experiments, callus cultures obtained from nucelli of two species, Citrus sinensis L. Osbeck, cv. Valencia, and Citrus limonia Osbeck, cv. Rangpur lime were used for anatomical and histochemical observations, callus samples from cultures in multiplication medium, in embryo induction medium, and somatic embryos in different stages of development were collected. Callus and embryo samples were also collected for analysis of gene expression through in situ hybridization. The anatomical and histochemical analysis showed differences between Rangpur lime and Valencia callus cultures. The in situ hybridization results evidenced the expression of the gene AtSERK1 in cultures of both citrus species, and also in both culture conditions: multiplication medium, with sucrose, and embryo induction medium, with maltose, indicating that the embriogenic potential is already installed in the callus cultures in multiplication medium. The results also lead to the conclusion that the gene is conserved between Arabidopsis thaliana and Citrus. Experiments aiming to synchronize the somatic embryogenesis formation in C. sinensis, cv. Valencia, were installed testing the effect of cold, desiccation and cell cluster size on somatic embryo formation. For that, callus cultures of ‘Valência’ sweet orange were cultivated in liquid medium in an orbital shaker, at 200 rpm, for 24h, and sieved through the following screens: 150 µm (P1), 300 µm (P2) and 600 µm (P3). The cell aggregates were then exposed to cold (4 o C, for four weeks, in the dark) and desiccation (27 o C, in Petri plates without culture medium, six days, in the dark) treatments, and cultured in incubators at 26 ± 1 o C and 300 lux of light intensity. The results lead to the conclusion that the separation of cultures in clusters of different sizes and the stress treatments were effective for synchronization and embryogenesis frequency. The size of cell clusters was the most important factor.

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Coordenação de Aperfeiçoamento de Pessoal de Nível Superior - CAPES
The citrus industry is one of the most important agricultural activities to Brazil, considering economic and social aspects. For production of fresh market fruit, there is little diversity of studies in Brazil, and investments are insufficient to change that. The northeastern region is fortunate to have the best conditions for development of the culture, and the micro-region around Garanhuns County, in Pernambuco Estate, has a great potential for the production of citrus fruits, for table consuming, because of the favourable soil and climatic conditions. The objective of this study was to evaluate the varieties of citrus, for table consuming, at the conditions of Southern Agreste region of Pernambuco, on agronomic parameters and physical characteristics, chemical and physico-chemical characteristics of fruits, in order to support the selection of genotypes for the region and contribute to the diversification of fruit plantation. The Citrus collection where the experiment took place is located at the Experimental Station of Brejão, which belongs to the Agricultural Institute of Pernambuco (IPA). For the study of the collection were used morphological descriptors proposed for citrus. Was measured the physical characteristics of the fruits: shape, epicarp color, height and diameter, fruit mass, thickness of the epicarp, mesocarp thickness, number of seeds per fruit and juice yield. Regarding physicalchemical and chemical parameters, were evaluated total soluble solids, titratable acidity, content of ascorbic acid (vitamin C) and hydrogenic potential. To evaluate plant growth, the variables chosen were stem diameter above and below the graft, plant height and canopy volume. The experimental design was randomized blocks with four replications and one plant per plot, being spaced 6x4 m. The treatments consisted in 15 varieties of oranges, two of lime and one of lemon. It was revealed that six varieties studied are within the requirements of the Brazilian Program for the Improvement Standards and Commercial Packaging of horticultural and five varieties of oranges had different maturation than is usually reported in the literature.
A citricultura é uma das atividades agrícolas de maior importância econômica e social para o Brasil. Para produção de frutas de consumo “in natura”, há pouca diversidade de estudos no Brasil, e os investimentos são insuficientes para mudar este quadro. A região nordeste é privilegiada por dispor das melhores condições para desenvolvimento da cultura, e a microrregião de Garanhuns, em Pernambuco, apresenta potencial para a produção de frutos cítricos de mesa, devido suas condições edafoclimáticas. Objetivou-se com este trabalho avaliar as variedades de citros de mesa nas condições edafoclimáticas da região Agreste Meridional de Pernambuco, mediante parâmetros agronômicos e características físicas, físicoquímicas e químicas dos frutos, visando dar suporte à seleção de genótipos para a região e contribuir com a diversificação da fruticultura. A coleção de Citros onde foi realizado o experimento está implantada na Estação Experimental de Brejão, pertencente ao Instituto Agronômico de Pernambuco (IPA). Para o estudo da coleção foram utilizados descritores morfológicos propostos para os citros. Foram aferidas as características físicas dos frutos como: formato; coloração do epicarpo; altura e diâmetro; massa do fruto; espessura do epicarpo mais mesocarpo; espessura do endocarpo; número de sementes por fruto e rendimento em suco. Para as análises físico-químicas e químicas foram avaliados: sólidos solúveis, acidez titulável, teor de ácido ascórbico (vitamina C) e potencial hidrogênionico. Nas plantas as variáveis de crescimento avaliadas foram: diâmetro do caule acima e abaixo da linha de enxertia, altura da planta e volume da copa. O delineamento experimental adotado foi em blocos ao acaso, com quatro repetições e uma planta por parcela, sendo espaçadas 6x4 m. Os tratamentos foram compostos por 15 variedades de laranjas, duas de limas e uma de limão. Foi revelado que das variedades estudadas, apenas seis laranjas, as limas e o limão estão dentro das exigências do Programa Brasileiro para a Modernização da Horticultura e que cinco variedades de laranjas apresentaram maturação diferente das relatadas na literatura.