Estudos sobre a germinação de sementes de marupã (Simaruba amara Aubl.). I. Composição química e curva de embebição das sementes germinação em diferentes temperaturas.

This work investigates the imbibition of Marupá (Simaruba amara Aubl.) seeds, their chemical composition and germination capacity at 20°, 25°, 30° e 35°C. The imbibition, expressed as percentage of the original weight of the seeds, was approximately 150,0% after 24 hours and 179,3% after 144 hours. The greatest percentage of germination and emergence velocity index (66% and 1.02 respectively) were obtained at. 30°C. Total carbohydrate content was 78.1% -in the coat and 46.7% in the whole seed, white the lipid content of the cotyledons was 45,7% and in the whole seed 23.8%. These results should aid futures studies on seed germination physiology of this species.Neste trabalho estuda-se a embebição das sementes de Marupã (Simaruba amara Aubl.), sua composição química e germinação à 20°, 25°, 30° e 35°C. A embebição, expressa como percentagem do peso inicial das sementes, foi aproximadamente 150,0% depois de 24 hs e 179,3% depois de 144 hs. A maior percentagem de germinação e I. V. E., respectivamente 66% e 1,02, foram obtidos à 30°C. 0 teor de carboidrato total foi de 78,1% no tegumento e 46,7% na semente inteira, enquanto que o de lipídios foi 45,7% nos cotilédones e 23,8% na semente inteira. Os resultados poderão auxiliar estudos posteriores na fisiologia da germinação de sementes desta espécie

Expressed sequence tag analysis of the human pathogen Paracoccidioides brasiliensis yeast phase: Identification of putative homologues of Candida albicans virulence and pathogenicity genes

Paracoccidioides brasiliensis, a thermodimorphic fungus, is the causative agent of the prevalent systemic mycosis in Latin America, paracoccidioidomycosis. We present here a survey of expressed genes in the yeast pathogenic phase of P. brasiliensis. We obtained 13,490 expressed sequence tags from both 5' and 3' ends. Clustering analysis yielded the partial sequences of 4,692 expressed genes that were functionally classified by similarity to known genes. We have identified several Candida albicans virulence and pathogenicity homologues in P. brasiliensis. Furthermore, we have analyzed the expression of some of these genes during the dimorphic yeast-mycelium-yeast transition by real-time quantitative reverse transcription-PCR. Clustering analysis of the mycelium-yeast transition revealed three groups: (i) RBT, hydrophobin, and isocitrate lyase; (ii) malate dehydrogenase, contigs Pb1067 and Pb1145, GPI, and alternative oxidase; and (iii) ubiquitin, delta-9-desaturase, HSP70, HSP82, and HSP104. the first two groups displayed high mRNA expression in the mycelial phase, whereas the third group showed higher mRNA expression in the yeast phase. Our results suggest the possible conservation of pathogenicity and virulence mechanisms among fungi, expand considerably gene identification in P. brasiliensis, and provide a broader basis for further progress in understanding its biological peculiarities.Univ São Paulo, Dept Ciencias Farmaceut, Fac Ciencias Farmaceut Ribeirao Preto, BR-14040903 Ribeirao Preto, SP, BrazilUniv São Paulo, Fac Filosofia Ciencias & Letras Ribeirao Pret, BR-14040903 Ribeirao Preto, SP, BrazilInst Pasteur, Unite Genet Mol Levures, Paris, FranceUniv Vale do Paraiba, UNIVAP, Vale Do Paraiba, BrazilUniv Mogi das Cruzes, Nucleo Integrado Biotecnol, Mogi Das Cruzes, BrazilUniversidade Federal de São Paulo, Dept Microbiol Imunol & Parasitol, São Paulo, BrazilUniversidade Federal de São Paulo, Dept Microbiol Imunol & Parasitol, São Paulo, BrazilWeb of Scienc

Predicting the Proteins of Angomonas deanei, Strigomonas culicis and Their Respective Endosymbionts Reveals New Aspects of the Trypanosomatidae Family

Endosymbiont-bearing trypanosomatids have been considered excellent models for the study of cell evolution because the host protozoan co-evolves with an intracellular bacterium in a mutualistic relationship. Such protozoa inhabit a single invertebrate host during their entire life cycle and exhibit special characteristics that group them in a particular phylogenetic cluster of the Trypanosomatidae family, thus classified as monoxenics. in an effort to better understand such symbiotic association, we used DNA pyrosequencing and a reference-guided assembly to generate reads that predicted 16,960 and 12,162 open reading frames (ORFs) in two symbiont-bearing trypanosomatids, Angomonas deanei (previously named as Crithidia deanei) and Strigomonas culicis (first known as Blastocrithidia culicis), respectively. Identification of each ORF was based primarily on TriTrypDB using tblastn, and each ORF was confirmed by employing getorf from EMBOSS and Newbler 2.6 when necessary. the monoxenic organisms revealed conserved housekeeping functions when compared to other trypanosomatids, especially compared with Leishmania major. However, major differences were found in ORFs corresponding to the cytoskeleton, the kinetoplast, and the paraflagellar structure. the monoxenic organisms also contain a large number of genes for cytosolic calpain-like and surface gp63 metalloproteases and a reduced number of compartmentalized cysteine proteases in comparison to other TriTryp organisms, reflecting adaptations to the presence of the symbiont. the assembled bacterial endosymbiont sequences exhibit a high A+T content with a total of 787 and 769 ORFs for the Angomonas deanei and Strigomonas culicis endosymbionts, respectively, and indicate that these organisms hold a common ancestor related to the Alcaligenaceae family. Importantly, both symbionts contain enzymes that complement essential host cell biosynthetic pathways, such as those for amino acid, lipid and purine/pyrimidine metabolism. These findings increase our understanding of the intricate symbiotic relationship between the bacterium and the trypanosomatid host and provide clues to better understand eukaryotic cell evolution.Fundação de Amparo à Pesquisa do Estado do Rio de Janeiro (FAPERJ)Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)ERC AdG SISYPHEUniv Fed Rio de Janeiro, Inst Biofis Carlos Chagas Filho, Lab Ultraestrutura Celular Hertha Meyer, BR-21941 Rio de Janeiro, BrazilUniv Fed Rio de Janeiro, Inst Biofis Carlos Chagas Filho, Lab Metab Macromol Firmino Torres de Castro, BR-21941 Rio de Janeiro, BrazilLab Bioinformat, Lab Nacl Computacao Cient, Rio de Janeiro, BrazilINRIA Grenoble Rhone Alpes, BAMBOO Team, Villeurbanne, FranceUniv Lyon 1, CNRS, UMR5558, Lab Biometrie & Biol Evolut, F-69622 Villeurbanne, FranceUniv Estadual Campinas, Inst Biol, Dept Genet Evolucao & Bioagentes, São Paulo, BrazilUniv São Paulo, Fac Ciencias Farmaceut Ribeirao Preto, Dept Ciencias Farmaceut, São Paulo, BrazilLab Nacl Ciencia & Tecnol Bioetano, São Paulo, BrazilUniv Fed Minas Gerais, Inst Ciencias Biol, Dept Bioquim & Imunol, Belo Horizonte, MG, BrazilUniv Fed Goias, Inst Ciencias Biol, Mol Biol Lab, Goiania, Go, BrazilFundacao Oswaldo Cruz, Inst Carlos Chagas, Lab Biol Mol Tripanossomatideos, Curitiba, Parana, BrazilFundacao Oswaldo Cruz, Inst Carlos Chagas, Lab Genom Func, Curitiba, Parana, BrazilUniv Estadual Campinas, Ctr Pluridisciplinar Pesquisas Quim Biol & Agr, São Paulo, BrazilUniv Fed Minas Gerais, Inst Ciencias Biol, Dept Parasitol, Belo Horizonte, MG, BrazilUniv Fed Santa Catarina, Dept Microbiol Imunol & Parasitol, Ctr Ciencias Biol, Lab Protozool & Bioinformat, Florianopolis, SC, BrazilUniv Fed Vicosa, Dept Bioquim & Biol Mol, Ctr Ciencias Biol & Saude, Vicosa, MG, BrazilInst Butantan, Lab Especial Ciclo Celular, São Paulo, BrazilUniv São Paulo, Dept Biol, Fac Filosofia Ciencias & Letras Ribeirao Preto, São Paulo, BrazilUniversidade Federal de São Paulo, Escola Paulista Med, Dept Microbiol Imunol & Parasitol, São Paulo, BrazilUniversidade Federal de São Paulo, Escola Paulista Med, Dept Microbiol Imunol & Parasitol, São Paulo, BrazilWeb of Scienc

Identification of metabolic pathways influenced by the G-protein coupled receptors GprB and GprD in Aspergillus nidulans

Heterotrimeric G-protein-mediated signaling pathways play a pivotal role in transmembrane signaling in eukaryotes. Our main aim was to identify signaling pathways regulated by A. nidulans GprB and GprD G-protein coupled receptors (GPCRs). When these two null mutant strains were compared to the wild-type strain, the DeltagprB mutant showed an increased protein kinase A (PKA) activity while growing in glucose 1% and during starvation. In contrast, the DeltagprD has a much lower PKA activity upon starvation. Transcriptomics and (1)H NMR-based metabolomics were performed on two single null mutants grown on glucose. We noted modulation in the expression of 11 secondary metabolism gene clusters when the DeltagprB and DeltagprD mutant strains were grown in 1% glucose. Several members of the sterigmatocystin-aflatoxin gene cluster presented down-regulation in both mutant strains. The genes of the NR-PKS monodictyphenone biosynthesis cluster had overall increased mRNA accumulation in DeltagprB, while in the DeltagprD mutant strain the genes had decreased mRNA accumulation. Principal component analysis of the metabolomic data demonstrated that there was a significant metabolite shift in the DeltagprD strain. The (1)H NMR analysis revealed significant expression of essential amino acids with elevated levels in the DeltagprD strain, compared to the wild-type and DeltagprB strains. With the results, we demonstrated the differential expression of a variety of genes related mainly to secondary metabolism, sexual development, stress signaling, and amino acid metabolism. We propose that the absence of GPCRs triggered stress responses at the genetic level. The data suggested an intimate relationship among different G-protein coupled receptors, fine-tune regulation of secondary and amino acid metabolisms, and fungal development

The Genome of Anopheles darlingi, the main neotropical malaria vector

Anopheles darlingi is the principal neotropical malaria vector, responsible for more than a million cases of malaria per year on the American continent. Anopheles darlingi diverged from the African and Asian malaria vectors ∼100 million years ago (mya) and successfully adapted to the New World environment. Here we present an annotated reference A. darlingi genome, sequenced from a wild population of males and females collected in the Brazilian Amazon. A total of 10 481 predicted protein-coding genes were annotated, 72% of which have their closest counterpart in Anopheles gambiae and 21% have highest similarity with other mosquito species. In spite of a long period of divergent evolution, conserved gene synteny was observed between A. darlingi and A. gambiae. More than 10 million single nucleotide polymorphisms and short indels with potential use as genetic markers were identified. Transposable elements correspond to 2.3% of the A. darlingi genome. Genes associated with hematophagy, immunity and insecticide resistance, directly involved in vectorhuman and vectorparasite interactions, were identified and discussed. This study represents the first effort to sequence the genome of a neotropical malaria vector, and opens a new window through which we can contemplate the evolutionary history of anopheline mosquitoes. It also provides valuable information that may lead to novel strategies to reduce malaria transmission on the South American continent. The A. darlingi genome is accessible at www.labinfo.lncc.br/index.php/anopheles- darlingi. © 2013 The Author(s)

Nucellar tissue culture, isolation and radiosensitivity of protoplasts in Citrus sinensis (L.) Osbeck cv. Pera

O longo período de juvenilidade, a alta heterozigosidade, a escassez de conhecimentos sobre o modo de herança de muitas características de interesse, a auto-incompatibilidade, a esterilidade e a embrionia nucelar são sérios obstáculos para os programas de melhoramento das espécies cítricas. Uma abordagem recente para o melhoramento genético é baseada na cultura de tecidos, incluindo o uso de protoplastos, a mutagênese in vitro e a seleção in vitro de características desejadas. No presente trabalho, foram estudados os fatores envolvidos na obtenção de calos embriogênicos de Citrus sinensis cv. Pera. Foram cultivados nucelos e óvulos extraídos de frutos de diferentes idades após a antese, em quatro diferentes meios de cultura e nas condições de escuro e de luminosidade. Os explantes desenvolveram embrióides e plântulas, os quais originaram calos embriogênicos a partir da região do hipocótilo. Os nucelos apresentaram as maiores percentagens de formação de calos embriogênicos nas diversas condições de cultivo testadas. O aparecimento e a proliferação, o dos calos embriogênicos ocorreram em meios de cultura sem horm6nios vegetais. O uso do 2,4-D nos meios de cultura resultou na inibição da embriogênese nucelar in vitro. Houve o surgimento de calos, sem características embriogênicas, a partir dos tegumentos dos óvulos. Esta observação foi confirmada através de análises histológicas. Visando determinar as condições ideais para o isolamento de protoplastos viáveis, a partir dos calos embriogênicos otimização. Este método foi bastante eficaz, tendo aumentado cerca de 15 vezes a produção de protoplastos viáveis. As melhores condições experimentais para o isolamento foram: digestão enzimática por 5 horas, em meio com as enzimas cellulase 307,6 mg/10 ml e macerozyme 30,3 mg/10 ml, e os estabilizadores osmóticos manitol 328,0 mM e sacarose 336,2 mM. A eficiência de isolamento (1,2 x 10 6 protoplastos viáveis/g) alcançada com as condições experimentais otimizadas, possibilita que os protoplastos venham a ser empregados em programas de melhoramento do cultivar Pera. Os protoplastos foram submetidos a várias doses de radiação gama 42 horas após o isolamento, para uma verificação da sensitividade dos mesmos, tendo sido o valor de LD 50 estimado em torno de 37,5 Gy.There are serious obstacles to the breeding programmes in citrus: prolonged juvenile period high heterozigosity, scarcity of information about the inheritance of many important characters, self-incompatibility, sterility, and nucellar embriony. A recent approach to the genetic improvement is based on tissue culture, including the use of protoplasts, "in vitro" mutagenesis and "in vitro" selection of desired characters. ln the present work, the factors involved in obtaining embryogenic callus in Citrus sinensis cv.Pera were studied. Ovules and nucelli extracted from fruitlets of different ages after anthesis were cultured on four different nutrient media including dark and light conditions. The explants developed embryoids and plantlets, which originated embryogenic callus from the hypocotyl region. Nucelli showed the highest percentages of embryogenic callus formation in all culture conditions tested. Emergence and proliferation of embryogenic callus occurred on nutrient media without hormones. The use of 2,4-D on nutrient media resulted in the inhibition of "in vitro" nucellar embryogenesis. There was non-embryogenic callus formation from the ovules integuments. This, observation was confirmed through histological analysis . Aiming to determine the ideal conditions to isolate viable protoplasts from embryogenic callus, the Simplex optimization method was used. This method was very efficient and increased the production of viable protoplasts about 15 times. The best experimental conditions for the isolation were: enzymatic digestion during 5 hours, on medium containing 307.6 mg/10 ml cellulase and 30.3 mg/10 ml macerozyme, and the osmotic stabilizers 328.0 mM mannitol and 336.2 mM sucrose. The isolation efficiency (1.2 x 106 viable protoplasts/g) reached with the optimized experimental conditions, enables protoplasts cultivar Pera. ln order to verify the radiosensitivity, protoplasts were submitted to several gamma radiation doses 42 hours after isolation and the LD50 value was estimated around 37.5 Gy