Uso do coeficiente de repetibilidade na seleção de clones de pessegueiro para o litoral sul de Santa Catarina

O presente trabalho teve como objetivo estimar os coeficientes de repetibilidade de caracteres relacionados à produtividade do pessegueiro, além de identificar e selecionar clones promissores para o Litoral Sul de Santa Catarina. Foram selecionadas plantas híbridas que apresentavam boa adaptação ao clima, produção, tamanho, sabor e firmeza dos frutos. Essas seleções foram enxertadas sobre o porta -enxerto Okinawa e plantadas no ano de 2005, em três diferentes locais: 1- Estação Experimental de Urussanga, situada a 40 metros de altitude; 2- Propriedade de fruticultor, situada a 200 metros de altitude; 3 - Propriedade de fruticultor, situada a 350 metros de altitude. As mudas foram plantadas no espaçamento de 6 x 1 metros e conduzidas no sistema de V . As avaliações foram realizadas durante os anos de 2007 e 2008. Os coeficientes de repetibilidade foram estimados pelo método da máxima verossimilhança restrita (REML), e a predição dos valores fenotípicos e genotípicos, pela melhor predição linear não viciada (BLUP), por meio do software Selegen-REML/BLUP. Todos os caracteres apresentaram considerável variabilidade genética, com estimativas de coeficientes de repetibilidade (r) variando de média a alta magnitude (0,54 a 0,74) para o caráter massa média dos frutos (MMF) e de baixa a média magnitude (0,22 a 0,39) para o caráter produção de frutos/planta (PTF). Os clones 1770 e 1443 apresentaram bom desempenho na média de todos os locais, enquanto os clones 0470 e 1307 se destacaram no local 1, o clone 1444 no local 2, os clones 0740 e 0926 no local 3 e o clone 1770 nos locais 1 e 2 de avaliação.

Relação entre características morfológicas de frutos e incidência de podridão carpelar em clones de macieira 'Gala' e 'Fuji' sobre diferentes porta-enxertos

A podridão carpelar tem-se tornado uma importante doença no Brasil, deixando de ser considerada uma doença secundária dentro do grupo das doenças de verão. O desenvolvimento e o formato dos frutos podem ser influenciados por eventos climáticos que ocorrem durante o período de polinização e frutificação da macieira. Além de outros fatores como nutrição das plantas, manejo da condução, tipo de porta-enxerto e cultivar copa. As alterações no formato dos frutos advindas destes fatores podem influenciar no aumento da intensidade da doença. O objetivo deste trabalho foi relacionar características morfológicas de frutos com a incidência de podridão carpelar, em clones de macieira, utilizando diferentes porta-enxertos, durante os ciclos de produção de 2009/2010 e 2010/2011, no município de Vacaria, no Estado do Rio Grande do Sul. O delineamento experimental foi em blocos casualizados, em arranjo fatorial 9 x 2, com nove clones das cultivares Gala e Fuji e dois porta-enxertos EM-9 e Marubakaido com interenxerto de EM-9. As características morfológicas dos frutos avaliados foram: relação entre comprimento e diâmetro de frutos (C/D), distância entre lóbulos, abertura calicinar, comprimento do tubo calicinar, número de sementes e classe de sintomas da doença nos carpelos. A doença não foi identificada nos clones da cultivar Gala, em ambos os ciclos de produção. Houve interação entre os fatores clones e porta-enxertos para as características morfológicas avaliadas, em ambos os ciclos de produção. A maior incidência da doença nos clones de 'Fuji' apresentou uma relação positiva com as características morfológicas dos frutos com maior abertura calicinar, menor relação C/D e maior distância entre lóbulos dos frutos, quando comparados aos clones de 'Gala'. O clone 'Fuji Suprema', enxertado sobre porta-enxerto EM-9, apresentou 19,33 % de incidência de podridão carpelar, sendo significativamente superior quando comparado aos 6,67% de incidência do porta-enxerto Marubakaido com interenxerto de EM-9. Dentre os gêneros de fungos isolados de carpelos de maçãs, Alternaria é o gênero que apresentou maior frequência na proporção de 14 e 20% nos ciclos de 2009/2010 e 2010/2011, respectivamente.

Antibiotic susceptibility and molecular epidemiology of Panton-Valentine leukocidin-positive meticillin-resistant Staphylococcus aureus: An international survey

The antibiotic susceptibility and molecular epidemiology of Panton-Valentine leukocidin (PVL)-positive meticillin-resistant Staphylococcus aureus (MRSA) isolates reported from 17 countries in the Americas, Europe and, Australia-Asia were analysed. Among a total of 3236 non-duplicate isolates, the lowest susceptibility was observed to erythromycin in all regions. Susceptibility to ciprofloxacin showed large variation (25%, 75% and 84% in the Americas, Europe and Australia-Asia, respectively). Two vancomycin-intermediate PVL-positive MRSA isolates were reported, one from Hong Kong and the other from The Netherlands. Resistance to trimethoprim/sulfamethoxazole and linezolid was <1%. Among 1798 MRSA isolates from 13 countries that were tested for the requested 10 non-β-lactam antibiotics, 49.4% were multisusceptible. However, multiresistant isolates (resistant to at least three classes of non-β-lactam antibiotics) were reported from all regions. Sequence type 30 (ST30) was reported worldwide, whereas ST80 and ST93 were exclusive to Europe and Australia, respectively. USA300 and related clones (ST8) are progressively replacing the ST80 clone in several European countries. Eight major clusters were discriminated by multilocus variable-number tandem repeat assay (MLVA), showing a certain geographic specificity. PVL-positive MRSA isolates frequently remain multisusceptible to non-β-lactam agents, but multiresistance is already prevalent in all regions. Surveillance of MRSA susceptibility patterns should be monitored to provide clinicians with the most current information regarding changes in resistance patterns.

Caracterização biométrica e respostas fisiológicas de diásporos de murucizeiro a tratamentos para superação da dormência

Diásporos (pirênios) de Byrsonima crassifolia (L.) H.B.K., dos clones Cristo, Santarém 2 e Tocantins 1, do Banco de Germoplasma da Embrapa Amazônia Oriental, foram caracterizados e submetidos a tratamentos pré-germinativos com o objetivo de aumentar a porcentagem e acelerar a germinação das sementes. Na caracterização, foram considerados os seguintes aspectos: massa, comprimento, largura, número de sementes e de lóculos por pirênio e espessura do endocarpo. Os tratamentos para superação da dormência consistiram em: a) pré-embebição dos pirênios em solução de ácido giberélico (500 mg L-1); b) pré-embebição em água seguida de fratura no endocarpo, e c) pré-embebição em solução de ácido giberélico com posterior fratura do endocarpo. Um tratamento-testemunha, representado por pirênios não submetidos a tratamento pré-germinativo, foi acrescentado para fins de comparação. Os pirênios dos três clones, com grande frequência, contêm mais de uma semente. A presença de pirênios desprovidos de sementes foi de apenas 1% nos clones Cristo e Santarém 2 e de 4% no clone Tocantins 1. Os diásporos do clone Cristo apresentaram maior massa e maiores dimensões. Em termos de número médio de sementes por pirênio, observaram-se valores semelhante nos clones Cristo (1,7) e Santarém 2 (1,8), enquanto esse valor foi menor no clone Tocantins 1 (1,3). Os três clones apresentaram endocarpo com espessura semelhante. Os tratamentos pré-germinativos proporcionaram aumentos na porcentagem de germinação e reduções no tempo médio de germinação, com respostas mais expressivas quando os pirênios foram previamente embebidos em solução de ácido giberélico ou em água e, posteriormente, submetidos à fratura do endocarpo.

Diversidade genética de clones de bananeira 'Prata-Anã' (AAB) por meio de marcadores SSR

Diversidade genética de 20 clones de 'Prata-Anã Gorutuba', quatro clones de 'Prata-Rio', quatro clones de 'Prata-Catarina' e as cultivares Caipira, Thap Maeo, Tropical, Maçã e Prata-Anã Comum foi avaliada por meio de marcadores moleculares Simple Sequence Repeats. De um total de 19 pares de primers SSRs utilizados, 57,8% deles amplificaram bandas polimórficas e distintas, 26,3% não produziram produtos específicos e 15,7% apresentaram falhas na amplificação de alguns indivíduos. O dendrograma indicou a formação de dois grupos. O primeiro grupo com a cultivar triploide Caipira, genoma exclusivamente A; enquanto o segundo (formado por sete subgrupos) agrupou todas as cultivares resultantes da hibridação natural ou artificial entre Musa acuminata e M. balbisiana, o subgrupo II, Tropical (AAAB) e o subgrupo III, Maçã (AAB). Os subgrupos IV, V, VI e VII foram formados, respectivamente, por: 'Prata-Catarina' clones 1 e 2; 'Prata-Rio' clones 1 e 2; 'Prata-Catarina' clone 3; 'Prata-Gorutuba' clones 12 e 17, 'Prata-Catarina' clone 4, 'Prata-Rio' clone 4, 'Thap Maeo' e 'Prata-Anã'. O subgrupo VIII foi formado exclusivamente pelos clones de' Prata-Anã Gorutuba'. Os resultados indicam a eficiência dos marcadores microssatélites na discriminação e na caracterização dos clones da 'Prata-Anã Gorutuba' da cultivar Prata-Anã.

Repetibilidade da produção, número e peso de fruto em cirigueleira (Spondias purpurea L.)

A repetibilidade permite estimar o número de avaliações ou ciclos produtivos para selecionar genótipos superiores com maior eficiência e menor custo operacional. Este trabalho estimou coeficientes de repetibilidade da produção, número e peso de fruto em 11 clones de cirigueleira, visando a avaliar a possibilidade de praticar seleção fenotípica individual. Foram analisados dados médios de 5 safras, oriundos de 3 plantas por clone, propagadas por estaquia. Os coeficientes foram estimados pelos métodos da análise de variância, componentes principais e análise estrutural. O método de componentes principais, baseados nas matrizes de variâncias e covariâncias, estima os maiores coeficientes em todas as características avaliadas. Com exceção do coeficiente estimado para número de frutos, pelo método de componentes principais, todos os outros atingiram valores iguais ou menores que 0,38. De acordo com os resultados obtidos, é possível realizar seleção fenotípica individual entre os clones de cirigueleira avaliados, com base na produção e número de frutos por planta. As avaliações de 6; 4 e 8 safras para produção, número e peso de frutos permitem selecionar clones promissores com cerca de 80 % de acurácia. O clone IPA-6 produziu 17,62 kg de frutos/planta, superando os demais materiais avaliados.

Allelic diversity and population structure in Vibrio cholerae O139 Bengal based on nucleotide sequence analysis

Comparative analysis of gene fragments of six housekeeping loci, distributed around the two chromosomes of Vibrio cholerae, has been carried out for a collection of 29 V. cholerae O139 Bengal strains isolated from India during the first epidemic period (1992 to 1993). A toxigenic O1 ElTor strain from the seventh pandemic and an environmental non-O1/non-O139 strain were also included in this study. All loci studied were polymorphic, with a small number of polymorphic sites in the sequenced fragments. The genetic diversity determined for our O139 population is concordant with a previous multilocus enzyme electrophoresis study in which we analyzed the same V. cholerae O139 strains. In both studies we have found a higher genetic diversity than reported previously in other molecular studies. The results of the present work showed that O139 strains clustered in several lineages of the dendrogram generated from the matrix of allelic mismatches between the different genotypes, a finding which does not support the hypothesis previously reported that the O139 serogroup is a unique clone. The statistical analysis performed in the V. cholerae O139 isolates suggested a clonal population structure. Moreover, the application of the Sawyer's test and split decomposition to detect intragenic recombination in the sequenced gene fragments did not indicate the existence of recombination in our O139 population.

SUSCETIBILIDADE DE CLONES DE LIMA ÁCIDA ‘TAHITI’ À PODRIDÃO FLORAL DOS CITROS

RESUMO A podridão floral dos citros (PFC), causada por Colletotrichum acutatum Simmons e C. gloeosporioides, é a doença fúngica mais importante em limeira-ácida ‘Tahiti’, pois, leva à queda prematura de flores e frutos, acarretando a redução da produção. Avaliou-se a suscetibilidade à PFC dos clones de lima-ácida ‘Tahiti’ “IAC 5”, “IAC 5-1”, “CNPMF/EECB”, “CNPMF 2000” e “CNPMF 2001”, em Bebedouro-SP. Todos os clones são suscetíveis à doença. A maior incidência de PFC em plantas do clone “IAC 5-1” indica que este clone apresenta maior suscetibilidade. Mesmo com sintomas, plantas do clone “CNPMF/EECB” apresentam maior fixação de frutos.

Microscopic, chemical, and molecular-biological investigation of the decayed medieval stained window glasses of two Catalonian churches

We investigated the decayed historical church window glasses of two Catalonian churches, both under Mediterranean climate. Glass surfaces were studied by scanning electron microscopy (SEM), energy dispersive spectrometry (EDS), and X-ray diffraction (XRD). Their chemical composition was determined by avelength-dispersive spectrometry (WDS) microprobe analysis. The biodiversity was investigated by molecular methods: DNA extraction from glass, amplification by PCR targeting the16S rRNA and ITS regions, and fingerprint analyses by denaturing gradient gel electrophoresis (DGGE). Clone libraries containing either PCR fragments of the bacterial 16S rDNA or the fungal ITS regions were screened by DGGE. Clone inserts were sequenced and compared with the EMBL database.

Glyoxysomal malate-dehydrogenase and malate synthase from Soybean cotyledons (Glycine max. L.): Enzyme association, antibody-production and cDNA cloning

In order to investigate a possible association between soybean malate synthase (MS; L-malate glyoxylate-lyase, CoA-acetylating, EC 4.1.3.2) and glyoxysomal malate dehydrogenase (gMDH; (S)-malate: NAD(+) oxidoreductase, EC 1.1.1.37), two consecutive enzymes in the glyoxylate cycle, their elution profiles were analyzed on Superdex 200 HR fast protein liquid chromatography columns equilibrated in low- and high-ionic-strength buffers. Starting with soluble proteins extracted from the cotyledons of 5-d-old soybean seedlings and a 45% ammonium sulfate precipitation, MS and gMDH coeluted on Superdex 200 HR (low-ionic-strength buffer) as a complex with an approximate relative molecular mass (M(r)) of 670000. Dissociation was achieved in the presence of 50 mM KCl and 5 mM MgCl2, with the elution of MS as an octamer of M, 510 000 and of gMDH as a dimer of M, 73 000. Polyclonal antibodies raised to the native copurified enzymes recognized both denatured MS and gMDH on immunoblots, and their native forms after gel filtration. When these antibodies were used to screen a lambda ZAP II expression library containing cDNA from 3-d-old soybean cotyledons, they identified seven clones encoding gMDH, whereas ten clones encoding MS were identified using an antibody to SDS-PAGE-purified MS. Of these cDNA clones a 1.8 kb clone for MS and a 1.3-kb clone for gMDH were fully sequenced. While 88% identity was found between mature soybean gMDH and watermelon gMDH, the N-terminal transit peptides showed only 37% identity. Despite this low identity, the soybean gMDH transit peptide conserves the consensus R(X(6))HL motif also found in plant and mammalian thiolases.

Epigenetic regulatory elements: Recent advances in understanding their mode of action and use for recombinant protein production in mammalian cells.

Successful generation of high producing cell lines requires the generation of cell clones expressing the recombinant protein at high levels and the characterization of the clones' ability to maintain stable expression levels. The use of cis-acting epigenetic regulatory elements that improve this otherwise long and uncertain process has revolutionized recombinant protein production. Here we review and discuss new insights into the molecular mode of action of the matrix attachment regions (MARs) and ubiquitously-acting chromatin opening elements (UCOEs), i.e. cis-acting elements, and how these elements are being used to improve recombinant protein production. These elements can help maintain the chromatin environment of the transgene genomic integration locus in a transcriptionally favorable state, which increases the numbers of positive clones and the transgene expression levels. Moreover, the high producing clones tend to be more stable in long-term cultures even in the absence of selection pressure. Therefore, by increasing the probability of isolating a high producing clone, as well as by increasing transcription efficiency and stability, these elements can significantly reduce the time and cost required for producing large quantities of recombinant proteins.

Sofosbuvir Inhibits Hepatitis E Virus Replication In Vitro and Results in an Additive Effect When Combined With Ribavirin.

Infection with hepatitis E virus genotype 3 may result in chronic hepatitis in immunocompromised patients. Reduction of immunosuppression or treatment with ribavirin or pegylated interferon-α can result in viral clearance. However, safer and more effective treatment options are needed. Here, we show that sofosbuvir inhibits the replication of hepatitis E virus genotype 3 both in subgenomic replicon systems as well as a full-length infectious clone. Moreover, the combination of sofosbuvir and ribavirin results in an additive antiviral effect. Sofosbuvir may be considered as an add-on therapy to ribavirin for the treatment of chronic hepatitis E in immunocompromised patients.

Involvement of a transforming-growth-factor-beta-like molecule in tumor-cell-derived inhibition of nitric-oxide synthesis in cerebral endothelial cells.

Nitric oxide (NO) has been shown to exert cytotoxic effects on tumor cells. We have reported that EC219 cells, a rat-brain-microvessel-derived endothelial cell line, produced NO through cytokine-inducible NO synthase (iNOS), the induction of which was significantly decreased by (a) soluble factor(s) secreted by DHD/PROb, an invasive sub-clone of a rat colon-carcinoma cell line. In this study, the DHD/PROb cell-derived NO-inhibitory factor was characterized. Northern-blot analysis demonstrated that the induction of iNOS mRNA in cytokine-activated EC219 cells was decreased by PROb-cell-conditioned medium. When DHD/PROb cell supernatant was fractionated by affinity chromatography using Con A-Sepharose or heparin-Sepharose, the NO-inhibitory activity was found only in Con A-unbound or heparin-unbound fractions, respectively, indicating that the PROb-derived inhibitory factor was likely to be a non-glycosylated and non-heparin-binding molecule. Pre-incubation of DHD/PROb-cell supernatant with anti-TGF-beta neutralizing antibody completely blocked the DHD/PROb-derived inhibition of NO production by EC219 cells. Addition of exogenous TGF-beta 1 dose-dependently inhibited NO release by EC219 cells. The presence of active TGF-beta in the DHD/PROb cell supernatant was demonstrated using a growth-inhibition assay. Moreover, heat treatment of medium conditioned by the less invasive DHD/REGb cells, which constitutively secreted very low levels of active TGF-beta, increased both TGF-beta activity and the ability to inhibit NO production in EC219 cells. Thus, DHD/PROb colon-carcinoma cells inhibited NO production in EC219 cells by secreting a factor identical or very similar to TGF-beta.

Prevalence and clinical outcomes for patients with MET protein expression in patients with non-small cell lung cancer in Europe: Results from the European Thoracic Oncology Platform Lungscape Project.

Aim The reported prevalence of MET overexpression varies from 25-55% in non-small cell lung cancer (NSCLC) and clinical correlations are emerging slowly. In a well-defined NSCLC cohort of the Lungscape program, we explore the epidemiology, the natural history of IHC MET positivity and its association to OS, RFS and TTR. Methods Resected stage I-III NSCLC identified based on the quality of clinical data and FFPE tissue availability were assessed for MET expression using immunohistochemistry (IHC) on TMAs (CONFIRM anti total c-MET assay, clone SP44, Ventana BenchMark platform). All cases were analysed at participating pathology laboratories using the same protocol, after passing an external quality assurance program. MET positive status is defined as ≥ 50% of tumor cells staining with 2+ or 3+ intensity. Results A total of 2709 cases are included in the iBiobank and will be analysed. IHC MET expression is currently available for 1552 patients, with positive MET IHC staining in 380 cases [24.5%; IHC 3+ in 157 cases (41.3%) and 2+ in 223 cases (58.7%)]. The cohort of 1552 patients includes 48.2%, 44.7% and 4.4% cases of adenocarcinoma, squamous and large cell histologies, respectively. IHC MET status was independent of stage, age and smoking history. Significant differences in MET positivity were associated with gender (32% vs. 21% for female vs. male, p < 0.001), with performance status (25% vs. 18% for 0 vs. 1-3, p = 0.006), and histology (34%, 14% and 24% for adenocarcinoma, squamous and large cell carcinoma, p < 0.001). IHC MET positivity was independent of the IHC ALK status (p = 0.08). At last FU, 52% of patients were still alive, with a median FU of 4.8 yrs. No association of IHC MET was found with OS, RFS or TTR. Conclusions The preliminary results for this large multicentre European cohort describe a prevalence of MET overexpression that seems lower than previous observations in NSCLC, such as reported for the OAM4971g trial, suggesting potential biological differences between surgically resected and metastatic disease. Analysis for the full cohort is ongoing and results will be presented. Disclosure L. Bubendorf: Disclosures: Stock ownership: Roche Advisory boards: Roche, Pfizer Research support: Roche; K. Schulze: Full time employee of Roche; A. Das-Gupta: I am a full time employee of Roche. All other authors have declared no conflicts of interest.

Enhancement of viability of radiosensitive (PBMC) and resistant (MDA-MB-231) clones in low-dose-rate cobalt-60 radiation therapy

AbstractObjective:In the present study, the authors investigated the in vitrobehavior of radio-resistant breast adenocarcinoma (MDA-MB-231) cells line and radiosensitive peripheral blood mononuclear cells (PBMC), as a function of different radiation doses, dose rates and postirradiation time kinetics, with a view to the interest of clinical radiotherapy.Materials and Methods:The cells were irradiated with Co-60, at 2 and 10 Gy and two different exposure rates, 339.56 cGy.min–1 and the other corresponding to one fourth of the standard dose rates, present over a 10-year period of cobalt therapy. Post-irradiation sampling was performed at pre-established kinetics of 24, 48 and 72 hours. The optical density response in viability assay was evaluated and a morphological analysis was performed.Results:Radiosensitive PBMC showed decrease in viability at 2 Gy, and a more significant decrease at 10 Gy for both dose rates. MDAMB- 231 cells presented viability decrease only at higher dose and dose rate. The results showed MDA-MB-231 clone expansion at low dose rate after 48–72 hours post-radiation.Conclusion:Low dose rate shows a possible potential clinical impact involving decrease in management of radio-resistant and radiosensitive tumor cell lines in cobalt therapy for breast cancer.