Joseane Padilha da Silva

Evidence of field-evolved resistance of Spodoptera frugiperda to Bt corn expressing Cry1F in Brazil that is still sensitive to modified Bt toxins.

Brazil ranked second only to the United States in hectares planted to genetically modified crops in 2013. Recently corn producers in the Cerrado region reported that the control of Spodoptera frugiperda with Bt corn expressing Cry1Fa has decreased, forcing them to use chemicals to reduce the damage caused by this insect pest. A colony of S. frugiperda was established from individuals collected in 2013 from Cry1Fa corn plants (SfBt) in Brazil and shown to have at least more than ten-fold higher resistance levels compared with a susceptible colony (Sflab). Laboratory assays on corn leaves showed that in contrast to SfLab population, the SfBt larvae were able to survive by feeding on Cry1Fa corn leaves. The SfBt population was maintained without selection for eight generations and shown to maintain high levels of resistance to Cry1Fa toxin. SfBt showed higher cross-resistance to Cry1Aa than to Cry1Ab or Cry1Ac toxins. As previously reported, Cry1A toxins competed the binding of Cry1Fa to brush border membrane vesicles (BBMV) from SfLab insects, explaining cross-resistance to Cry1A toxins. In contrast Cry2A toxins did not compete Cry1Fa binding to SfLab-BBMV and no cross-resistance to Cry2A was observed, although Cry2A toxins show low toxicity to S. frugiperda. Bioassays with Cry1AbMod and Cry1AcMod show that they are highly active against both the SfLab and the SfBt populations. The bioassay data reported here show that insects collected from Cry1Fa corn in the Cerrado region were resistant to Cry1Fa suggesting that resistance contributed to field failures of Cry1Fa corn to control S. frugiperda.

Transcription profile of soybean-root-knot nematode interaction reveals a key role of phythormones in the resistance reaction

BackgroundRoot-knot nematodes (RKN– Meloidogyne genus) present extensive challenges to soybean crop. The soybean line (PI 595099) is known to be resistant against specific strains and races of nematode species, thus its differential gene expression analysis can lead to a comprehensive gene expression profiling in the incompatible soybean-RKN interaction. Even though many disease resistance genes have been studied, little has been reported about phytohormone crosstalk on modulation of ROS signaling during soybean-RKN interaction.ResultsUsing 454 technology to explore the common aspects of resistance reaction during both parasitism and resistance phases it was verified that hormone, carbohydrate metabolism and stress related genes were consistently expressed at high levels in infected roots as compared to mock control. Most noteworthy genes include those encoding glycosyltransferases, peroxidases, auxin-responsive proteins and gibberellin-regulated genes. Our data analysis suggests the key role of glycosyltransferases, auxins and components of gibberellin signal transduction, biosynthesis and deactivation pathways in the resistance reaction and their participation in jasmonate signaling and redox homeostasis in mediating aspects of plant growth and responses to biotic stress.ConclusionsBased on this study we suggest a reasonable model regarding to the complex mechanisms of crosstalk between plant hormones, mainly gibberellins and auxins, which can be crucial to modulate the levels of ROS in the resistance reaction to nematode invasion. The model also includes recent findings concerning to the participation of DELLA-like proteins and ROS signaling controlling plant immune or stress responses. Furthermore, this study provides a dataset of potential candidate genes involved in both nematode parasitism and resistance, which can be tested further for their role in this biological process using functional genomics approaches.

Compostos fenólicos totais e taninos condensados em frutas nativas do cerrado

The native fruits from Brazilian savanna are arousing increasing interest due to their nutritional and functional properties combined with the potential to add value and conserve the biodiversity of this biome. Many phenolic compounds have antioxidant capacity to neutralize free radicals generated in the body that are associated with many chronic degenerative diseases. The objective of this study was to evaluate the efficiency of three extraction solutions (acetone 70%, ethanol 95% and methanol 99.8 %) and determining the levels of phenolic compounds in 10 species of native fruits from Brazilian savanna, using the Folin-Ciocalteu method for total phenolic compounds and vanillin method for condensed tannins. Acetone 70 % proved to be the best solvent extractor for total phenolics and condensed tannins in fruits of Pouteria gardneriana, Eugenia dysenterica, E. klostzchiana, E. punicifolia, Plinia edulis, Campomanesia sp. and for total phenolics in Brosimium gaudichaudii and for condensed tannins in Jaracatia spinosa. The ethanol 95 % was more efficient in the extraction of total phenolics of Jaracatia spinosa and condensed tannins of Brosimium gaudichaudii. The methanol 99.8 % was more efficient in the extraction of condensed tannins of Eugenia dysenterica. The contents of phenolic compounds ranged between 90 and 327 mg of gallic acid equivalent per 100 g of pulp for the species of E. dysenterica and E. punicifolia respectively. The levels of condensed tannins ranged between 4 and 291 mg of catechin equivalent per 100 g of pulp for the species of E. dysenterica and E. calcyna respectively. Evaluated species of fruit from Brazilian savanna can be considered good sources of total phenolic compounds, although the specific nature of different types should be evaluated in future studies.

Morphology, distribution and abundance of antennal sensilla in three stink bug species (Hemiptera: Pentatomidae).

The neotropical stink bugs, Euschistus heros, Piezodorus guildinii and Edessa meditabunda, are important pests of soybean and other crops throughout Central America and in South America from Northern Argentina to Brazil. Mate finding and host plant location in these species depend largely on their chemical communication, and semiochemicals are important mediators of these behaviors. In this study scanning electron microscopy was used to examine the external morphology, distribution and abundance of antennal sensilla on males, females and 5th instar nymphs of these species. Nine morphologically different sensilla types were found: trichod sensilla, type 1 and 2 (ST1 and ST2), long and short basiconic sensilla (SB1, SB2, and SB3), slit-tipped and knob-shaped basiconic sensilla, long chaetic sensilla (Sch) and coeloconic sensilla (Sco). Differences were detected in the abundance and arrangement of the sensilla over the antennal segments in individuals of the same species and among the species studied. The Sch, Sco and the slit-tipped and knob-shaped basiconic sensilla accounted for the major difference in sensilla types among the species. The ST1 was the most abundant type and was restricted to the flagellum. The pedicel of E. heros differs from the pedicels of P. guildinii and E. meditabunda mainly by the absence of Sch. There was a sexual dimorphism in ST1, SB1 and SB2, and this may be an indicative of their roles in detection of male-produced sex pheromone and odors derived from the host plants. The SB2 was lacking in the antennal tip of both sexes and 5th instar nymphs, but was abundant on the second flagellar segment of females of the three species. The same types of sensilla were found on 5th instar nymphs, but always in significantly lower numbers. The morphology and putative functions of each sensilla were compared and discussed.

Vibrational communication and mating behaviour of Dichelops melacanthus (Hemiptera: Pentatomidae) recorded from loudspeaker membranes and plants

Vibrational communication is important for successful mating in various stink bugs species. The vibrational signals from males and females of Dichelops melacanthus Dallas (Hemiptera: Pentatomidae) are recorded from a nonresonant substrate (i.e. a loudspeaker membrane) to characterize the temporal and spectral properties of these vibrational signals, as well as on a resonant substrate (i.e. bean plants) to obtain information about how these signals are altered when they are transmitted through the plants. On the loudspeaker membrane, D. melacanthus males and females emit only one male or one female song, respectively. However, when the insects are placed on bean leaves, a more complex repertoire is recorded, with three different songs for each sex. The first female and male songs appear to have calling functions and the third male and female songs are emitted during courtship. The second female and male songs are emitted after the first song, although their functions in mating behaviour are not clear. The identified repertoire is similar to those of other Neotropical stink bugs, starting with songs 1 and 2 and developing into song 3. Frequency modulation is observed in the female songs recorded from the loudspeaker membrane and the plants. The signals recorded from plants present higher harmonic peaks compared with the signals recorded from the loudspeaker membrane. The presence of species and sex‐specific songs during mating confirms the important role of vibrational communication in mate location and recognition. The temporal and spectral characteristic signals are influenced by the substrate used to record the songs emitted by D. melacanthus.

Composição de ácidos graxos em polpa de frutas nativas do cerrado

Araticum (Annona Crassiflora Mart.), coquinho-azedo (Butia Capitata Mart.) and pequi (Caryocar Brasiliense Camb.) are native fruits from the Brazilian Savanna with strong potential for sustained exploration. The objective of this study was the characterization of lipid contents and fatty acid profiles of these species. Total lipids were hot extracted with petroleum ether in Soxhlet extractor. The oil used for methyl ester profiles was cold extracted by Bligh & Dyer and characterized by gas chromatograph, using flame ionization detector. The pequi pulp presented highest oil content (30.89 %); araticum and coquinho-azedo pulps presented respectively 2.14 and 2.73 %. The oleic and palmitic acids predominated in all three species and all of them presented prevalence of unsaturated fat acids. The highest value was found in araticum pulp (78.3 %), followed by coquinho-azedo pulp (63.3 %). The araticum and coquinho-azedo pulps presented highest content of linolenic acid (2.5 to 3.7 %). Methyl esters of caproic acid found in these pulps seems to be linked to perception of fruit aroma of these native fruits from Cerrado.

Chemical characterization of the essential oils from leaves of mandarins Sunki, Cleopatra and their hybrids

Sunki Mandarin [Citrus sunki (Hayata) hort. ex Tanaka] is a type of micromandarin from South China largely used as a rootstock in Brazil. Cleopatra mandarin (Citrus reshni hort. ex. Tanaka) is a well-formed tree, ornamental, with red-orange fruits. The main objective of this work was to analyze the chemical composition of the essential oil in leaves of accessions of Sunki and Cleopatra mandarins and hybrids. The essential oils were obtained from the leaves by hydrodistillation in a Clevenger-type apparatus and analyzed by gas chromatography/flame ionization detector (GC/FID) and GC/mass spectrometry (GC/MS). The oil yields ranged from 1.27% (BCG562 – Hybrid of ‘Sunki’ C12080) to 0.33% (BCG564 – Sunki from Florida) (dry weight basis), and fifty-five constituents were detected. The major constituents were β-pinene (2.5–49.9%), limonene (0–49.0%), sabinene (0.5–35.2%), linalool (0.7–27.3%), thymol methyl ether (0–22.3%), p-cymene (0.2–21.2%), γ-terpinene (0–15.1%), 1.8-cineole (0–11.0%), terpinen-4-ol (0.5–9.4%), (E)-β-ocimene (0.6–6.4%), α-pinene (1.1–4.4%), (E)-nerolidol (1.0–3.2%), α-terpineol (0.4–3.0%) and myrcene (0.7–2.4%).

Plant regeneration from embryogenic callus and cell suspensions of Brachiaria brizantha

In monocots, in vitro plant regeneration can be obtained through somatic embryogenesis (Vasil and Vasil 1980, 1982; Vasil 2005). Evidence of more than one morphogenic response in the same explant, under the control of different auxin/cytokinin ratios and concentrations, was observed in species of the Poaceae family such as sorghum (Sorghum bicolor, (L.) Moench.) , minor mil le t (Paspalum scrobiculatum L.), sugar cane (Saccharum officinarum L.), and baby bamboo (Pogonatherum paniceum Lam. Hack.) (reviewed by Wang et al. 2008). Brachiaria callus has been induced using seeds as explants from Brachiaria brizantha (Hochst. ex A. Rich.) Stapf, Brachiaria decumbens Stapf, Brachiaria ruziziensis Germain et Evrard and Brachiaria dictyoneura [Brachiaria humidicola (Rendle) Schweick vr. Lanero, ex B. dictyoneura] (Tohme et al. 1996). Subsequently, using the same methodology, calli were induced from 76% of isolated embryos from seeds of B. brizantha (Lenis-Manzano 1998). The formation of somatic embryos and multiple shoots from seedling apical meristems was demonstrated for B. ruziziensis (Ishigaki et al. 2009). In this system, mature seeds were used as source of explants and cultured on a medium containing 4 mg/L 2,4-dichlorophenoxyacetic acid (2,4-D). Ishigaki et al. (2012) reported that embryogenic calli were more suitable than shoot apex explants for genetic transformation of ruzigrass. B. brizantha cv. Marandu is the most important and cultivated forage grass in Brazil. It is tolerant to spittlebugs (Homoptera: Cercopidae complex, mainly genera Deois and Zulia), an economically important pest for Brachiaria (Felismino et al. 2012). This cultivar is considered as the key forage in beef cattle production in Brazil. Breeding is hindered by its apomictic reproductive mode (the production of clones of the mother plant), which reduces the possibility of being used for hybridizations (Valle and Savidan 1996). Alternatives to conventional breeding of B. brizantha would include genetic modification via transformation. However, methods of transformation and tissue culture of these plants are not yet established. The development of embryogenic calli has been reported from 73% of the B. brizantha cv. Marandu isolated seed embryos in culture and 67% of calli regenerated plants (Silveira et al. 2003). More recently, the histology of embryos isolated from the in vitro cultivated seed was shown (Lenis-Manzano et al. 2010). The formation of multiple shoots was reported in cultured basal segments from micropropagated plantlets of this cultivar (Pinheiro et al. 2000), a system that was efficient for in vitro chromosome duplication in Brachiaria. Somatic embryogenesis and organogenesis in B. brizantha is influenced by several factors such as genotype, explant type, and culture conditions (Cabral et al. 2011). A high rate of cell division in explants used for genetic transformation ofmonocots is a prerequisite for the integration of exogenous DNA (Hiei et al. 1997; Vasil 2005). Rapid cell proliferation is a characteristic of embryogenic tissue. Cell suspension cultures (CS) are well known for their cell division capacity and the possibility of developmental-stage Electronic supplementary material The online version of this article (doi:10.1007/s11627-015-9690-0) contains supplementary material, which is available to authorized users. G. B. Cabral (*) :V. T. C. Carneiro : J. P. da Silva :D. M. A. Dusi Embrapa Genetic Resources and Biotechnology, Brasília, Brazil e-mail: glaucia.cabral@embrapa.br

Ação antagônica e de metabólitos bioativos de Trichoderma spp. contra os patógenos Sclerotium rolfsii e Verticillium dahliae

The antagonistic effect shown by fungi of the genus Trichoderma was assessed against the plant pathogens Sclerotium rolfsii and Verticillium dahlia, isolated from ornamental aster and strawberry, respectively. Twenty isolates of the antagonist were employed; they belonged to the species T. asperellum, T. hamatum, T. harzianum, T. koningiopsis, T. spirale and Trichoderma sp. Antagonism was investigated by means of dual culture tests and bioactive metabolite production based on the overlapping plate method. Most selected isolates had an inhibitory effect on the mycelial growth of S. rolfsii and V. dahlia, compared to controls. Except for non-volatile metabolite production tests, inhibitory action against V. dahliae was highest. Even the thermostable non-volatile metabolites showed inhibitory effect against both tested pathogens, evidencing that the secondary metabolites of the antagonist have a significant effect on these pathogenic species.

New isolates of Trichoderma antagonistic to Sclerotinia sclerotiorum

Forty-nine isolates of Trichoderma from the Brazilian Midwest were evaluated for their antagonistic activity in vitro against Sclerotinia sclerotiorum (causal agent of white mold), which were then identified based on their nuclear ribosomal ITS sequences. Paired culture tests showed that all isolates exhibited some antagonism, with a maximum of 77% mycelial inhibition and complete inhibition of sclerotia production. Two isolates were found to be the most promising biocontrol agents, considering both antagonistic parameters (CEN1253 - T. koningiopsis and CEN1265 - T. brevicompactum). Five different species were identified: T. harzianum (23), T. spirale (9), T. koningiopsis (8), T. brevicompactum (7) and T. asperellum (2). These isolates are stored in the Embrapa Fungi Collection for Biological Control and the information obtained in the experiments will be incorporated into the database of biological assets within the genetic resources information system (Allele) and be made available for further studies.