EVALUATION OF PHOTODEGRADABLE TAPE ON GRAFTING IN CITRUS NURSERY TREES

The use of a photodegradable tape was evaluated on 'Valencia' sweet orange nursery trees budded both on Rangpur lime and Swingle citrumelo in a greenhouse in Bebedouro-SP, Brazil, from September to November 2009. On both rootstocks three wrapping procedures were evaluated: i) conventional polyethylene tape wrapped around the bud eye; ii) photodegradable tape wrapped around the bud eye, and iii) photodegradable tape wrapped around the graft junction without covering the bud eye. The following variables were measured: time spent for wrapping, percentage of bud sprouting, length and stem diameter of the scion shoot, and percentage of commercially valuable nursery trees. The trial was conducted following a randomized complete block design, with six treatments, four replications and 12 trees per plot. The use of photodegradable tape, with or without covering the bud eye, anticipated bud sprouting; despite of the longer time spent with wrapping when the photodegradable tape was used. Plants grafted onto the less vigorous Swingle citrumelo rootstock showed lower bud sprout percentages when the bud eye was covered with the photodegradable tape.

Xylella fastidiosa: An in vivo system to study possible survival strategies within citrus xylem vessels based on global gene expression analysis

Xylella fastidiosa inhabits the plant xylem, a nutrient-poor environment, so that mechanisms to sense and respond to adverse environmental conditions are extremely important for bacterial survival in the plant host. Although the complete genome sequences of different Xylella strains have been determined, little is known about stress responses and gene regulation in these organisms. In this work, a DNA microarray was constructed containing 2,600 ORFs identified in the genome sequencing project of Xylella fastidiosa 9a5c strain, and used to check global gene expression differences in the bacteria when it is infecting a symptomatic and a tolerant citrus tree. Different patterns of expression were found in each variety, suggesting that bacteria are responding differentially according to each plant xylem environment. The global gene expression profile was determined and several genes related to bacterial survival in stressed conditions were found to be differentially expressed between varieties, suggesting the involvement of different strategies for adaptation to the environment. The expression pattern of some genes related to the heat shock response, toxin and detoxification processes, adaptation to atypical conditions, repair systems as well as some regulatory genes are discussed in this paper. DNA microarray proved to be a powerful technique for global transcriptome analyses. This is one of the first studies of Xylella fastidiosa gene expression in vivo which helped to increase insight into stress responses and possible bacterial survival mechanisms in the nutrient-poor environment of xylem vessels.

Mating system parameters at hierarchical levels of fruits, individuals and populations in the Brazilian insect-pollinated tropical tree, Tabebuia roseo-alba (Bignoniaceae)

For many tree species, mating system analyses have indicated potential variations in the selfing rate and paternity correlation among fruits within individuals, among individuals within populations, among populations, and from one flowering event to another. In this study, we used eight microsatellite markers to investigate mating systems at two hierarchical levels (fruits within individuals and individuals within populations) for the insect pollinated Neotropical tree Tabebuia roseo-alba. We found that T. roseo-alba has a mixed mating system with predominantly outcrossed mating. The outcrossing rates at the population level were similar across two T. roseo-alba populations; however, the rates varied considerably among individuals within populations. The correlated paternity results at different hierarchical levels showed that there is a high probability of shared paternal parentage when comparing seeds within fruits and among fruits within plants and full-sibs occur in much higher proportion within fruits than among fruits. Significant levels of fixation index were found in both populations and biparental inbreeding is believed to be the main cause of the observed inbreeding. The number of pollen donors contributing to mating was low. Furthermore, open-pollinated seeds varied according to relatedness, including half-sibs, full-sibs, self-sibs and self- half-sibs. In both populations, the effective population size within a family (seed-tree and its offspring) was lower than expected for panmictic populations. Thus, seeds for ex situ conservation genetics, progeny tests and reforestation must be collected from a large number of seed-trees to guarantee an adequate effective population in the sample.

Regeneration and characterization of somatic hybrids combining sweet orange and mandarin/mandarin hybrid cultivars for citrus scion improvement

Protoplast fusion between sweet orange and mandarin/mandarin hybrids scion cultivars was performed following the model "diploid embryogenic callus protoplast + diploid mesophyll-derived protoplast". Protoplasts were isolated from embryogenic calli of 'Pera' and 'Westin' sweet orange cultivars (Citrus sinensis) and from young leaves of 'Fremont', Nules', and 'Thomas' mandarins (C. reticulata), and 'Nova' tangelo [C. reticulata x (C. paradisi x C. reticulata)]. The regenerated plants were characterized based on their leaf morphology (thickness), ploidy level, and simple sequence repeat (SSR) molecular markers. Plants were successfully generated only when 'Pera' sweet orange was used as the embryogenic parent. Fifteen plants were regenerated being 7 tetraploid and 8 diploid. Based on SSR molecular markers analyses all 7 tetraploid regenerated plants revealed to be allotetraploids (somatic hybrids), including 2 from the combination of 'Pera' sweet orange + 'Fremont' mandarin, 3 'Pera' sweet orange + 'Nules' mandarin, and 2 'Pera' sweet orange + 'Nova' tangelo, and all the diploid regenerated plants showed the 'Pera' sweet orange marker profile. Somatic hybrids were inoculated with Alternaria alternata and no disease symptoms were detected 96 h post-inoculation. This hybrid material has the potential to be used as a tetraploid parent in interploid crosses for citrus scion breeding.

Genetic transformation of Citrus sinensis with Citrus tristeza virus (CTV) derived sequences and reaction of transgenic lines to CTV infection

Transgenic Citrus sinensis (L.) Osb. plants, cvs. Valencia and Hamlin, expressing Citrus tristeza virus (CTV) derived sequences were obtained by genetic transformation. The gene constructs were pCTV-CP containing the 25 kDa major capsid protein gene (CTV-CP), pCTV-dsCP containing the same CTV-CP gene in an intron-spliced hairpin construct, and pCTV-CS containing a 559 nt conserved region of the CTV genome. The transgenic lines were identified by PCR and the transgene integration was confirmed by Southern blot. Transgene mRNA could be detected in most transgenic lines containing pCTV-CP or pCTV-CS transgene. The mRNA of pCTV-dsCP transgene was almost undetectable, with very light bands in most analyzed plants. The transgene transcription appears to be closely linked to the type of gene construct. The virus challenge assays reveals that all transgenic lines were infected. However, it was possible to identify propagated clones of transgenic plants of both cultivars studied with a low virus titer, with values similar to the non-inoculated plants (negative control). These results suggested that the transgenic plants present some level of resistance to virus replication. The higher number of clones with low virus titer and where mRNA could not be detected or was presented in a very light band was found for pCTV-dsCP-derived transgenic lines.

Morphological description of the mouthparts of the Asian citrus psyllid, Diaphorina citri Kuwayama (Hemiptera: Psyllidae)

Scanning (SEM) and transmission (TEM) electron microscopy were used to elucidate the morphology of the rostrum, as well as the mandibular and maxillary stylets of the psyllid Diaphorina cirri, vector of phloem-inhabiting bacteria associated with citrus huanglongbing (HLB) disease. D. cirri has a cone-shaped rostrum that extends behind the pair of prothoracic coxae. The stylet bundle comprises a pair of mandibular (Md) and maxillary (Mx) stylets with a mean length of 513.3 mu m; when retracted, their proximal portions form a loop and are stored in the crumena (Cr). Serial cross-sections of the rostrum revealed that the mandibles are always projected in front of the maxillary stylets. The two maxillary stylets form the food and salivary canals, with diameters of 0.9 mu m and 0.4 mu m respectively. These two canals merge at the end of the stylets forming a common duct with a length of 4.3 mu m and a mean diameter of 0.9 mu m. The acrostyle, a distinct anatomical structure present in the common duct of aphid maxillary stylets, was not observed by TEM in the ultrathin cross-sections of the common duct (CD) of D. citri. This study provides new information on D. citri mouthparts that may help to understand the behaviour of this important vector of HLB-associated bacteria. (C) 2011 Elsevier Ltd. All rights reserved.

Glyphosate has low toxicity to citrus plants growing in the field

Gravena, R., Filho, R. V., Alves, P. L. C. A., Mazzafera, P. and Gravena, A. R. 2012. Glyphosate has low toxicity to citrus plants growing in the field. Can. J. Plant Sci. 92: 119-127. There has been controversy over whether glyphosate used for weed management in citrus fields causes significant toxicity to citrus plants. Glyphosate may be toxic to non-target plants exposed to accidental application or drift. This work evaluated glyphosate toxicity in plants of Valencia citrus (Citrus sinensis. L. Osbeck) grafted onto 'Rangpur lime' (Citrus limonia L. Osbeck) and citrumelo 'Swingle' (Poncirus trifoliata (L.) Raf x Citrus paradisi Mad) by trunk- or foliar-directed herbicide applications under field conditions. In the first experiment, glyphosate was sprayed at rates of 0, 90, 180, 260, 540, 1080 and 2160 g a.e. ha(-1) directly on the trunk to a height of 5 cm above the grafting region. In the second experiment, glyphosate was sprayed on the plant canopies at rates of 0, 0.036, 0.36, 3.6, 36, 360 and 720 g a.e. ha(-1). There was no visual damage caused by glyphosate applied directly to the trunk, but the plants were affected by glyphosate sprayed directly on the canopies at rates over 360 g a.e. ha(-1). The main symptom was observed in the new shoots formed after the application, indicating an effect on meristems. Little or no effect was observed in mature leaves. Eight days after application the levels of shikimate, total free amino acids and total phenolic compounds were unaffected. All plants affected by glyphosate recovered between 6 and 12 mo after the treatments. Therefore, despite some transient symptoms Valencia citrus grafted onto 'Rangpur lime' and citrumelo 'Swingle' were tolerant to glyphosate.

IN VITRO TRYPANOCIDAL EVALUATION OF PINANE DERIVATIVES FROM ESSENTIAL OILS OF RIPE FRUITS FROM Schinus terebinthifolius RADDI (ANACARDIACEAE)

Essential oils of ripe fruits from Schinus terebinthifolius (Anacardiaceae), obtained using a pilot extractor and a Clevenger apparatus were chemically characterized. Due the high amount of (-)-alpha-pinene in both oils, this monoterpene was tested against the protozoan parasite Trypanosoma cruzi, showing a moderate potential (IC50 63.56 mu g/mL) when compared to benznidazole (IC50 43.14 mu g/mL). Otherwise, (-)-alpha-pinene oxide did not showed anti-trypanosomal activity (IC50 > 400 mu g/mL) while (-)-pinane showed an IC50 of 56.50 mu g/mL. The obtained results indicated that the epoxydation of a-pinene results to the loss of the anti-parasitic activity while its hydrogenation product, contributed slightly to the increased activity.

Potential dietary sources of ellagic acid and other antioxidants among fruits consumed in Brazil: Jabuticaba (Myrciaria jaboticaba (Vell.) Berg)

BACKGROUND: The objectives of this study were (1) to evaluate the content of ellagic acid in fruits consumed by the Brazilian population, including native ones; (2) to further characterize rich sources in relation to ascorbic acid, phenolics contents and in vitro antioxidant capacity; and (3) to study the distribution and effect of ripening stage on ellagitannins content of jabuticaba (Myrciaria jaboticaba). The content of free ellagic acid and ellagic acid derivatives such as ellagitannins was analyzed using high-performance liquid chromatography (HPLC). RESULTS: Ellagic acid was detected in 10 out of a total of 35 fruits analyzed. The content of free ellagic acid in fruits varied from 0.0028 to 0.085 g kg(-1) (FW) and total ellagic acid varied from 0.215 to 3.11 g kg(-1) (FW). All the seven fruits belonging to the Myrtaceae family evaluated in this study presented high contents of ellagitannins in their composition, with jabuticaba, grumixama and cambuci (all native from Brazil) showing the highest total ellagic acid contents. Jabuticaba, the most consumed in Brazilamongthose and already adapted to commercial plantations, contained concentrated phenolics compounds, including ellagitannins, in the peel. Anthocyanins (cyanidin derivatives) increased significantly through ripening of jabuticaba and were not present in the pulp or seeds. Samples collected from three different locations during summer, winter and spring had total ellagic contents varying from 1.88 to 3.31 g kg(-1) (FW). The decrease in ellagic acid content with ripening was more accentuated for pulp (eight times) compared to seeds (2.3 times) and peel (2.0 times). CONCLUSION: These results showed the potential of jabuticaba as dietary source of ellagic acid and reinforced consumption of the whole fruit by the population. (C) 2011 Society of Chemical Industry

Dispersal strategies of Aceria guerreronis (Acari: Eriophyidae), a coconut pest

The dispersal of plant-feeding mites can occur involuntarily, through transportation of infested plant parts, or voluntarily, by walking to new plant parts or to suitable spots where biotic (phoresis) or abiotic (wind, agricultural tools, etc.) factors carry them over long distances. Elucidating the dispersal mechanisms of the coconut mite, Aceria guerreronis Keifer, is important for understanding the process of colonization of new fruits of a same or different plants, essential for the improvement of control strategies of this serious coconut pest. Thus, the objective of this work was to investigate the voluntary dispersal mechanisms of this mite. The hypothesis that the coconut mite disperses by walking, phoresis or wind were tested. The coconut mite was shown to be able to walk short distances between fruits of the same bunch or between bunches of the same plant. Phoresis on insects of the orders Hymenoptera (Apidae), Coleoptera (Curculionidae) and Lepidoptera (Phycitidae) was evaluated in the laboratory and in the field. Although in the laboratory mites were shown to be able to climb onto honeybees, field investigations failed to show these insects as important carriers of the pest, corroborating findings of previous works; however, both laboratory and field investigations suggested the curculionid Parisoschoenus obesulus Casey to be able to transport the coconut mite between plants. Similarly, laboratory and field investigations suggested wind to be important in the dispersal of the coconut mite between plants.

Antioxidant capacity and mineral content of pulp and peel from commercial cultivars of citrus from Brazil

Four Citrus species (C. sinensis, cvs. Pera and Lima; C. latifolia Tanaka cv. Tahiti; C limettioides Tanaka cv. Sweet lime and C. reticulate, cv. Ponkan) grown in Brazil were characterised in relation to contents of minerals, ascorbic acid, total polyphenols and antioxidant capacity of pulps and peels. In general, the peels demonstrated significantly higher contents of all compounds than the pulps (p < 0.05), with the exception of the Pera orange pulp that presented the highest acid ascorbic content (68 mg/100 ml), while the Tahiti lime peel presented the lowest (8 mg/100 g). Citrus showed high levels of potassium, calcium and magnesium, and the peels were considered sources of these minerals. The Ponkan mandarin peel presented the highest antioxidant capacity. The antioxidant capacity of citrus was correlated both to vitamin C and phenolics. Aside from citrus pulps, the peels are also good sources of bioactive compounds and minerals, and can be explored for their health promoting values in food products. (C) 2012 Elsevier Ltd. All rights reserved.

Ectopic expression of a fruit phytoene synthase from Citrus paradisi Macf. promotes abiotic stress tolerance in transgenic tobacco

Abscisic acid (ABA) is an important regulator of plant responses to environmental stresses and an absolute requirement for stress tolerance. Recently, a third phytoene synthase (PSY3) gene paralog was identified in monocots and demonstrated to play a specialized role in stress-induced ABA formation, thus suggesting that the first committed step in carotenogenesis is a key limiting step in ABA biosynthesis. To examine whether the ectopic expression of PSY, other than PSY3, would similarly affect ABA level and stress tolerance, we have produced transgenic tobacco containing a fruit-specific PSY (CpPSY) of grapefruit (Citrus paradisi Macf.). The transgenic plants contained a single- or double-locus insertion and expressed CpPSY at varying transcript levels. In comparison with the wild-type plants, the CpPSY expressing transgenic plants showed a significant increase on root length and shoot biomass under PEG-, NaCl- and mannitol-induced osmotic stress. The enhanced stress tolerance of transgenic plants was correlated with the increased endogenous ABA level and expression of stress-responsive genes, which in turn was correlated with the CpPSY copy number and expression level in different transgenic lines. Collectively, these results provide further evidence that PSY is a key enzyme regulating ABA biosynthesis and that the altered expression of other PSYs in transgenic plants may provide a similar function to that of the monocot's PSY3 in ABA biosynthesis and stress tolerance. The results also pave the way for further use of CpPSY, as well as other PSYs, as potential candidate genes for engineering tolerance to drought and salt stress in crop plants.

Rootstocks for high fruit yield and quality of 'Tahiti' lime under rain-fed conditions

Despite considerable research conducted on 'Tahiti' lime [Citrus latifolia (Yu Tanaka) Tanaka] in several countries, few long-term studies have focused on rootstock effects on fruit production and quality under non-irrigated conditions. As for many other fruit crops, rootstock studies for 'Tahiti' lime are often based on the evaluation of several horticultural responses simultaneously, instead of considering multivariate statistical approaches which may provide with more comprehensive information. Consequently, a trial was installed to evaluate the horticultural performance of non-irrigated 'Tahiti' lime trees budded onto the following 12 rootstocks: 'HRS 801' and 'HRS 827' hybrids; 'Rubidoux', 'FCAV' and 'Flying Dragon' trifoliates; 'Sun Chu Sha Kat' and 'Sunki' mandarins; 'Cravo Limeira' and 'Cravo FCAV' 'Rangpur' limes; 'Carrizo' citrange, 'Swingle' citrumelo, and 'Orlando' tangelo. The trial was installed in 2001, in an 8 m x 5 m spacing with no supplementary irrigation. Measurements of yield, fruit quality oriented to different consuming markets, canopy volume and tree tolerance to drought, were performed. A multivariate cluster analysis identified both 'Rangpur' lime rootstocks as those inducing larger cumulative yield and higher percentage of fruits for the domestic market, with highest drought tolerance to the trees. Despite of their high susceptibility to drought stress under non-irrigated conditions, the 'Flying Dragon' and 'FCAV' trifoliate rootstocks performed outstandingly for 'Tahiti' lime, inducing higher yield efficiency, early bearing and larger percentage of high-quality fruits for foreign markets, with smaller trees more suitable for high-density plantings. (c) 2012 Elsevier B.V. All rights reserved.

Evaluation of the schistosomicidal activity of the steroidal alkaloids from Solanum lycocarpum fruits

Solanum lycocarpum (Solanaceae), a Brazilian medicinal plant known as "wolf fruit," contains about 1.5% of glycoalkaloids in its dried fruits, consisting mainly of solamargine and solasonine. The present work reports the obtainment of the alkaloidic extract of the S. lycocarpum fruit by acid-base extraction and the isolation of the major alkaloid heterosides by chromatographic means, as well as the evaluation of their in vitro schistosomicidal activities. The in vitro schistosomicidal activities of the alkaloidic extract of S. lycocarpum fruits and its isolated steroidal alkaloids were undertaken against adult worms of Schistosoma mansoni. The alkaloidic extract (20, 32, and 50 mu g mL(-1)), solasonine (50 mu M), solamargine (32 and 50 mu M), and equimolar mixture of glycoalkaloids (20, 32, and 50 mu M) lead to the separation of all couple worms and extensive disruption on their teguments, such as sloughing, as well as their deaths within 24 h of incubation. In addition, the alkaloidic extract (10 and 15 mu g mL(-1)), solasonine (50 mu M), solamargine (10, 15, and 20 mu M), and equimolar mixtures of glycoalkaloids (10 and 15 mu M) reduced the development of eggs produced by the adult worms. Solamargine, containing the sugar chain moiety chacotriose, was more active than the solasonine, which contains solatriose sugar chain moiety. A synergistic effect was also observed for a mixture of solamargine and solasonine. Therefore, the alkaloidic extract of S. lycocarpum, and its major components, solamargine and solasonine, showed promising schistosomicidal activity.

Effect of contact and systemic insecticides on the sharpshooter Bucephalogonia xanthophis (Hemiptera: cicadellidae), a vector of Xylella fastidiosa in citrus

The knockdown and toxic effects of insecticides of different chemical groups and modes of action registered for citrus in Brazil were investigated for effective control of Bucephalogonia xanthophis, a sharpshooter vector of Xylella fastidiosa in citrus. The active ingredients dimethoate (1.2 mL/1.2L), imidacloprid (0.24 mL/1.2L) and lambda-cyhalothrin (0.24 mL/1.2L), as well as a control (water), were sprayed onto branches of potted-citrus nursery trees to evaluate the effect of residual contact. The insects were confined on sprayed branches by using sleeve cages, in groups of 10 per branch (5 branches/treatment). Lambdacyhalothrin showed a knockdown effect on B. xanthophis (>70% mortality within 2 h of exposure), and the residues were effective for approximately one wk. Imidacloprid, lambdacyhalothrin and dimethoate suppressed the vector populations for up to 3 wk after application, when the insects were exposed to sprayed plants for at least 24 h. In another experiment, 2 neonicotinoid insecticides (thiamethoxam and imidacloprid) were applied by soil drench to potted nursery trees, in order to study their systemic effect, i.e., mortality by ingestion on sharpshooter adults. Thiamethoxam and imidacloprid effectively controlled the vectors at all concentrations tested, when the insects were exposed to treated plants for 24 h (>80% mortality) or 48 h (near 100% mortality). The knockdown effect of thiamethoxam and lambda-cyhalothrin might be particularly important to prevent vector transmission of X. fastidiosa in citrus groves.