Paternity identification in sugarcane polycrosses by using microsatellite markers

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Estudo da ação do inseticida Tiametoxam na sobrevivência e no comportamento de operárias de Apis mellifera L., 1758 (Hymenoptera: Apidae) em diferentes idades

The bee Apis mellifera has a great importance because it is the most economically valuable pollinator for crops worldwide, ensuring cross-pollination and increasing fruit yield. Moreover, agriculture increasingly develops chemicals to control weeds, fungi and insect pests to ensure productivity. Insecticides are used on a large scale in the state of São Paulo, in cultures of citrus for control of greening. Applications are usually made by aircraft and as a result of the effect derives a significant mortality is observed in apiaries near the plantations. Honey bees can get in contact with such chemical agent through their activities of water harvesting, plant resins, pollen and nectar. Intoxication resulting from this exposure can be lethal, which is easily detectable, or cause effects on the physiology and behavior of the insect. These, in turn are hardly detectable, such as paralysis, disorientation, behavioral changes, but can compromise the entire social structure of the colony, therefore aimed to study the effects of the insecticide Thiamethoxam behavior of honeybees A. mellifera. Newly emerged individuals and with 10 days of age were tested. Applications of 1 μL de Thiamethoxam, diluted in acetone, were made on the dorsal thorax with a microapplicator. Preliminarily, it was observed LD50 twenty four hours after topical treatment of Thiamethoxam. It was found that the LD50 for newly emerged honeybees is 8 ng/bee and for honeybees with 10 days of age is 18 ng/bee. The behaviors were analyzed 1 hour after application of insecticide at doses corresponding to LD50/100, LD50/50, LD50/10 and LD50, besides the control group. In the test reflex proboscis extension, there was impairment of behavior at doses of 8 and 18 ng/bee workers in newly emerged workers and 10 days of age, respectively. And in locomotor behavior was no change only at a dose of 18 ng/bee workers at 10 days of age... (Complete abstract click electronic access below)

Análise do efeito de inseticidas neonicotinóides sobre a orientação e coleta de alimento de operárias de Apis mellifera L., 1758 (Hymenoptera: Apidae)

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Xenia Effects on Quality of Maize Female Inbred in Production of Hybrid Seeds

In Maize (Zea maize L.), cost of hybrid seed production is directly related to the yield and quality of seed obtained per hectare of female parent. It is also important to consider the effects that a male parent can exert on the development of hybrid seed in the female parent. This effect is known as xenia. The objectives of this study were to evaluate xenia effects on 1) yield as 80K units, 2) germination of the hybrid seed and 3) susceptibility of the hybrid seed to mechanical damage. One female inbred and four male inbred lines were selected from a parent list of hybrids. The experiment was designed to allow individual cross pollination between each male inbred and the female inbred line. For use as a control, the female inbred was allowed to self pollinate. Experiments were conducted in Illinois and Iowa during 2008 and 2009 and in Nebraska during 2009. A significant inbred effect was detected on yield as 80k (α=0.001). The selfed female and pollination with male inbred B resulted in lower yields of hybrid seed. For germination, a significant inbred effect was detected (α=0.001), but was due to lower germination percentage of seed produced on the selfed female. All hybrid combinations resulted in higher germination percentages with no significant differences among hybrids. The inbred x mechanical damage interaction was significant (P=0.04) for effects on cold saturated soil germination tests. Use of inbred B resulted in a two-percentage-point reduction in cold germination when treated with the impact simulator. In a maize seed company, the production research group provides yield estimates for production of new hybrid combinations. Results from this study indicate that using only the female inbred yield may provide inaccurate estimates. Therefore to improve yield estimation, experiments should be designed to include male inbreds. Male inbreds can also impart a negative effect to the hybrid seed on tolerance to mechanical damage, thus lowering quality and increasing seed discard. When testing for hybrid seed germination, there is no need to consider distinct hybrid combinations. Female inbreds can be grown in open-pollinated fields to avoid loss of vigor observed with selfing. Advisor: George Graef

Studio molecolare dei meccanismi dell'autoincompatibilità gametofitica in pero europeo (Pyrus communis)

Self-incompatibility (SI) systems have evolved in many flowering plants to prevent self-fertilization and thus promote outbreeding. Pear and apple, as many of the species belonging to the Rosaceae, exhibit RNase-mediated gametophytic self-incompatibility, a widespread system carried also by the Solanaceae and Plantaginaceae. Pear orchards must for this reason contain at least two different cultivars that pollenize each other; to guarantee an efficient cross-pollination, they should have overlapping flowering periods and must be genetically compatible. This compatibility is determined by the S-locus, containing at least two genes encoding for a female (pistil) and a male (pollen) determinant. The female determinant in the Rosaceae, Solanaceae and Plantaginaceae system is a stylar glycoprotein with ribonuclease activity (S-RNase), that acts as a specific cytotoxin in incompatible pollen tubes degrading cellular RNAs. Since its identification, the S-RNase gene has been intensively studied and the sequences of a large number of alleles are available in online databases. On the contrary, the male determinant has been only recently identified as a pollen-expressed protein containing a F-box motif, called S-Locus F-box (abbreviated SLF or SFB). Since F-box proteins are best known for their participation to the SCF (Skp1 - Cullin - F-box) E3 ubiquitine ligase enzymatic complex, that is involved in protein degradation through the 26S proteasome pathway, the male determinant is supposed to act mediating the ubiquitination of the S-RNases, targeting them for the degradation in compatible pollen tubes. Attempts to clone SLF/SFB genes in the Pyrinae produced no results until very recently; in apple, the use of genomic libraries allowed the detection of two F-box genes linked to each S haplotype, called SFBB (S-locus F-Box Brothers). In Japanese pear, three SFBB genes linked to each haplotype were cloned from pollen cDNA. The SFBB genes exhibit S haplotype-specific sequence divergence and pollen-specific expression; their multiplicity is a feature whose interpretation is unclear: it has been hypothesized that all of them participate in the S-specific interaction with the RNase, but it is also possible that only one of them is involved in this function. Moreover, even if the S locus male and female determinants are the only responsible for the specificity of the pollen-pistil recognition, many other factors are supposed to play a role in GSI; these are not linked to the S locus and act in a S-haplotype independent manner. They can have a function in regulating the expression of S determinants (group 1 factors), modulating their activity (group 2) or acting downstream, in the accomplishment of the reaction of acceptance or rejection of the pollen tube (group 3). This study was aimed to the elucidation of the molecular mechanism of GSI in European pear (Pyrus communis) as well as in the other Pyrinae; it was divided in two parts, the first focusing on the characterization of male determinants, and the second on factors external to the S locus. The research of S locus F-box genes was primarily aimed to the identification of such genes in European pear, for which sequence data are still not available; moreover, it allowed also to investigate about the S locus structure in the Pyrinae. The analysis was carried out on a pool of varieties of the three species Pyrus communis (European pear), Pyrus pyrifolia (Japanese pear), and Malus × domestica (apple); varieties carrying S haplotypes whose RNases are highly similar were chosen, in order to check whether or not the same level of similarity is maintained also between the male determinants. A total of 82 sequences was obtained, 47 of which represent the first S-locus F-box genes sequenced from European pear. The sequence data strongly support the hypothesis that the S locus structure is conserved among the three species, and presumably among all the Pyrinae; at least five genes have homologs in the analysed S haplotypes, but the number of F-box genes surrounding the S-RNase could be even greater. The high level of sequence divergence and the similarity between alleles linked to highly conserved RNases, suggest a shared ancestral polymorphism also for the F-box genes. The F-box genes identified in European pear were mapped on a segregating population of 91 individuals from the cross 'Abbé Fétel' × 'Max Red Bartlett'. All the genes were placed on the linkage group 17, where the S locus has been placed both in pear and apple maps, and resulted strongly associated to the S-RNase gene. The linkage with the RNase was perfect for some of the F-box genes, while for others very rare single recombination events were identified. The second part of this study was focused on the research of other genes involved in the SI response in pear; it was aimed on one side to the identification of genes differentially expressed in compatible and incompatible crosses, and on the other to the cloning and characterization of the transglutaminase (TGase) gene, whose role may be crucial in pollen rejection. For the identification of differentially expressed genes, controlled pollinations were carried out in four combinations (self pollination, incompatible, half-compatible and fully compatible cross-pollination); expression profiles were compared through cDNA-AFLP. 28 fragments displaying an expression pattern related to compatibility or incompatibility were identified, cloned and sequenced; the sequence analysis allowed to assign a putative annotation to a part of them. The identified genes are involved in very different cellular processes or in defense mechanisms, suggesting a very complex change in gene expression following the pollen/pistil recognition. The pool of genes identified with this technique offers a good basis for further study toward a better understanding of how the SI response is carried out. Among the factors involved in SI response, moreover, an important role may be played by transglutaminase (TGase), an enzyme involved both in post-translational protein modification and in protein cross-linking. The TGase activity detected in pear styles was significantly higher when pollinated in incompatible combinations than in compatible ones, suggesting a role of this enzyme in the abnormal cytoskeletal reorganization observed during pollen rejection reaction. The aim of this part of the work was thus to identify and clone the pear TGase gene; the PCR amplification of fragments of this gene was achieved using primers realized on the alignment between the Arabidopsis TGase gene sequence and several apple EST fragments; the full-length coding sequence of the pear TGase gene was then cloned from cDNA, and provided a precious tool for further study of the in vitro and in vivo action of this enzyme.

Levantamento de ocorrência, alternativas de manejo, mecanismos de resistência e herança genética do capim-amargoso (Digitaria insularis) resistente ao herbicida glyphosate

O glyphosate é o principal herbicida utilizado no manejo de plantas daninhas na agricultura, aplicado em alguns sistemas de forma repetitiva ao longo de cada ano. Esta prática selecionou biótipos resistentes de espécies de plantas daninhas, sendo o capim-amargoso (Digitaria insularis) selecionado no Brasil. Portanto, se tornam necessários estudos para entender, manejar e reduzir a infestação do capim-amargoso resistente ao glyphosate. Dessa forma, esta pesquisa foi desenvolvida com os objetivos de: (i) mapear áreas do Brasil com possíveis infestações de capim-amargoso resistente ao glyphosate; (ii) avaliar alternativas químicas de seu manejo; (iii) elucidar os mecanismos de resistência ao glyphosate e; (iv) avaliar a herança genética dos genes que conferem resistência ao glyphosate. Para o desenvolvimento dos experimentos foram coletadas sementes de biótipos potencialmente resistentes de diversas regiões do Brasil onde ocorreram falhas de controle de D. insularis após a aplicação de glyphosate. Na primeira etapa da pesquisa foram realizados experimentos para determinação de uma dose discriminatória de triagementre as populações resistentes e suscetíveis ao glyphosate, através de curvas de dose-resposta, para identificar a resistência ao Glyphosate, sendo que estes dados foram utilizados para mapear a ocorrência de biótipos resistentes em algumas regiões do país. Na segunda etapa foi conduzido um experimento em casa-de-vegetação visando encontrar herbicidas alternativos ao Glyphosate para controle do capim-amargoso, utilizando herbicidas recomendados para as culturas do milho e algodão, tanto em condições de aplicação de pré como em pós-emergência da planta daninha. Na terceira etapa foram realizados ensaios para determinar a existência de absorção e translocação diferencial do glyphosate em biótipos suscetíveis e resistentes, juntamente com a análise molecular para comparar a região 106 do gene que codifica a EPSPs nestes biótipos. Por fim um estudo de polinização cruzada foi conduzido para avaliar se genes de resistência ao glyphosate são transferidos para a geração seguinte após inflorescências de biótipos suscetíveis serem acondicionadas com as de biótipos resistentes, submetendo a geração seguinte a experimentos de curva de dose-resposta com o glyphosate. Através do modelo de curva dose-resposta do programa estatístico R, determinou-se a dose de 960 g e.a ha-1, como a dose utilizada para triagem dos biótipos oriundos de diferentes regiões do Brasil. Com isto foram gerados mapas indicando a presença ou ausência de resistência ao herbicida, sendo que as região oeste do Paraná e sul do Mato Grosso do Sul apresentam maior número de localidades com a presença de biótipos resistentes. As alternativas de controle viáveis como pós-emergentes no estádio de um a dois perfilhos, foram os herbicidas Nicosulfuron, Imazapic + Imazapyr, Atrazine, Haloxifop-methyl e Tepraloxydim. Na pré-emergência do capim-amargoso os herbicidas Atrazine, Isoxaflutole, S-metolachlor, Clomazone, Diuron e Flumioxazin se apresentaram como eficazes para o controle desta espécie. Os resultados do experimento de absorção, translocação e comparação da região 106 não mostraram diferenças entre os biótipos resistente e suscetível. O experimento sobre cruzamento entre biótipos resistente e suscetível determinou a espécie D. insularis como autógama e sem transferência de genes que causam a resistência ao glyphosate.

Approaches and Strategy for Cancer Research and Surveillance Data: Integration, Information Pipeline, Data Models, and Informatics Opportunities

Thesis (Ph.D.)--University of Washington, 2016-04

The Effect of Legal Culture on the Development of International Evidentiary Practice: From the ‘Robing Room’ to the ‘Melting Pot’

This paper draws on some of the preliminary findings of a small pilot study which aimed to discover what evidentiary challenges a range of practitioners with experience of different international trials faced in the cases they were involved in, and what practices were developed to deal with these challenges. The findings in this study are based on the data collected from The Hague-based institutions, the ICC, the ICTY, the ICTY and ICTR Appeals Chamber, and the Special Tribunal for the Lebanon (STL). It is argued that professionals moving from institution to institution are engaged in a process of cross-pollination which itself influences the practices that develop, although a common understanding of certain evidentiary issues in international trials remains fragmented and at times elusive.

Identifying key knowledge needs for evidence-based conservation of wild insect pollinators: a collaborative cross-sectoral exercise

In response to evidence of insect pollinator declines, organisations in many sectors, including the food and farming industry, are investing in pollinator conservation. They are keen to ensure that their efforts use the best available science. We convened a group of 32 ‘conservation practitioners’ with an active interest in pollinators and 16 insect pollinator scientists. The conservation practitioners include representatives from UK industry (including retail), environmental non-government organisations and nature conservation agencies. We collaboratively developed a long list of 246 knowledge needs relating to conservation of wild insect pollinators in the UK. We refined and selected the most important knowledge needs, through a three-stage process of voting and scoring, including discussions of each need at a workshop. We present the top 35 knowledge needs as scored by conservation practitioners or scientists. We find general agreement in priorities identified by these two groups. The priority knowledge needs will structure ongoing work to make science accessible to practitioners, and help to guide future science policy and funding. Understanding the economic benefits of crop pollination, basic pollinator ecology and impacts of pesticides on wild pollinators emerge strongly as priorities, as well as a need to monitor floral resources in the landscape.

Depression by inbreeding after four sucessive self-pollination squash generations

The strategy for breeding F-1 hybrid squash is to develop parental lines through self-pollination. However, it increases plant mean homozygosis, which is not the natural genetic state of a cross-pollinated species, and can cause inbreeding depression. The objective of this work was to evaluate this depression with sucessive generations of self-pollination (without selection) in Cucurbita moschata, cv. Piramoita. Populations were obtained from lines with one to four generations of self-pollination (obtained by the SSD method), from the original cv. Piramoita (population SO). Randomized blocks were used with five treatments (different generations of self-pollination -S-0 to S-4), six replicates and five plants per plot. Regression analysis was made by the Wright inbreeding coefficient (F) to measure the homozygosis level effect on vigor loss. There was a linear reduction of mean weight and fruit length, seed production (number and weight) per fruit with the increase of the homozygosis level; however inbreeding did not affect seed quality (weight of 100 seeds and germination).

Biotic versus abiotic pollination in Oeceoclades maculata (Lindl.) Lindl. (Orchidaceae)

Geographical variation in the reproductive biology of widespread species often occurs at their distributional boundaries. We sought to determine whether such variation has occurred in an invasive orchid, Oeceoclades maculata, across its naturalized range. We compared its reproductive biology in a Brazilian population with that published for a population on the Caribbean island of Puerto Rico. In the state of Sao Paulo, O. maculata flowers between December and February, at the height of the rainy season. Similar fruit sets were observed in manual self (76%) and cross (70.4%) pollination treatments. The fruit set of plants protected from both pollinators and rainfall was 6.1%, whereas plants exposed only to rainfall had a fruit set of 41.4%, slightly less than the controls (48.3%). Like the Puerto Rico population, reproduction is primarily through rain-assisted autogamy, but unlike observations made on the island, outcrossing can eventually occur. We observed two butterfly species (Heliconius ethilla narcaea and Heliconius erato phyllis) pollinating O. maculata. Secretory epidermal cells and trichomes of the spur lumen produced 0.7 mu L of 25% (sucrose equivalents) nectar per flower each morning, which was stored in a dilated basal portion of the spur and reabsorbed by the afternoon. Thus, geographical variation in reproductive biology exists across the broad invasive range of O. maculata.

Linking herbivory and pollination: Costs and selection implications in Centrosema virginianum Bentham (Fabaceae: Papilionoideae)

This research first evaluated levels and type of herbivory experienced by Centrosema virginianum plants in their native habitat and how florivory affected the pollinator activity. I found that populations of C. virginianum in two pine rockland habitat fragments experienced higher herbivory levels (15% and 22%) compared with plants in the protected study site (8.6%). I found that bees (Hymenoptera) pollinated butterfly pea. Furthermore, I found that florivores had a negative effect in the pollinators visitation rates and therefore in the seed set of the population. ^ I then conducted a study using a greenhouse population of C. virginianum. I applied artificial herbivory treatments: control, mild herbivory and severe herbivory. Flower size, pollen produced, ovules produced and seeds produced were negatively affected by herbivory. I did not find difference in nectar volume and quality by flowers among treatments. Surprisingly, severely damaged plants produced flowers with larger pollen than those from mildly damaged and undamaged plants. Results showed that plants tolerated mild and severe herbivory with 6% and 17% reduction of total fitness components, respectively. However, the investment of resources was not equisexual. ^ A comparison in the ability of siring seeds between large and small pollen was necessary to establish the biological consequence of size in pollen performance. I found that fruits produced an average of 18.7 ± 1.52 and 17.7 ± 1.50 from large and small pollen fertilization respectively. These findings supported a pollen number-size trade-off in plants under severe herbivory treatments. As far as I know, this result has not previously been reported. ^ Lastly, I tested how herbivory influenced seed abortion patterns in plants, examining how resources are allocated on different regions within fruits under artificial herbivory treatments. I found that self-fertilized fruits had greater seed abortion rates than cross-fertilized fruits. The proportion of seeds aborted was lower in the middle regions of the fruits in cross-fertilized fruits, producing more vigorous progeny. Self-fertilized fruits did not show patterns of seedling vigor. I also found that early abortion was higher closer to the peduncular end of the fruits. Position of seeds within fruits could be important in the seed dispersion mechanism characteristic of this species. ^

Delays In Receiving Obstetric Care And Poor Maternal Outcomes: Results From A National Multicentre Cross-sectional Study.

The vast majority of maternal deaths in low-and middle-income countries are preventable. Delay in obtaining access to appropriate health care is a fairly common problem which can be improved. The objective of this study was to explore the association between delay in providing obstetric health care and severe maternal morbidity/death. This was a multicentre cross-sectional study, involving 27 referral obstetric facilities in all Brazilian regions between 2009 and 2010. All women admitted to the hospital with a pregnancy-related cause were screened, searching for potentially life-threatening conditions (PLTC), maternal death (MD) and maternal near-miss (MNM) cases, according to the WHO criteria. Data on delays were collected by medical chart review and interview with the medical staff. The prevalence of the three different types of delays was estimated according to the level of care and outcome of the complication. For factors associated with any delay, the PR and 95%CI controlled for cluster design were estimated. A total of 82,144 live births were screened, with 9,555 PLTC, MNM or MD cases prospectively identified. Overall, any type of delay was observed in 53.8% of cases; delay related to user factors was observed in 10.2%, 34.6% of delays were related to health service accessibility and 25.7% were related to quality of medical care. The occurrence of any delay was associated with increasing severity of maternal outcome: 52% in PLTC, 68.4% in MNM and 84.1% in MD. Although this was not a population-based study and the results could not be generalized, there was a very clear and significant association between frequency of delay and severity of outcome, suggesting that timely and proper management are related to survival.

Relationship Between Lower-limb Muscle Strength And Functional Independence Among Elderly People According To Frailty Criteria: A Cross-sectional Study.

Muscle strength and functional independence are considered to be determinants of frailty levels among elderly people. The aim here was to compare lower-limb muscle strength (LLMS) with functional independence in relation to sex, age and number of frailty criteria, and to ascertain the influence of these variables on elderly outpatients' independence. Quantitative cross-sectional study, in a tertiary hospital. The study was conducted on 150 elderly outpatients of both sexes who were in a cognitive condition allowing oral communication, between October 2005 and October 2007. The following instruments were used: five-times sit-to-stand test (FTSST), Functional Independence Measurement (FIM) and Lawton's Instrumental Activities of Daily Living Scale (IADL). Descriptive, comparative, multivariate, univariate and Cronbach alpha analyses were performed. The mean time taken in the FTSST was 21.7 seconds; the mean score for FIM was 82.2 and for IADL was 21.2; 44.7% of the subjects presented 1-2 frailty criteria and 55.3% > 3 criteria. There was a significant association between LLMS and functional independence in relation to the number of frailty criteria, without homogeneity regarding sex and age. Functional independence showed significant influence from sex and LLMS. Elderly individuals with 1 or 2 frailty criteria presented greater independence in all FTSST scores. The subjects with higher LLMS presented better functional independence.

Cross-sectional Study Comparing Different Therapeutic Modalities For Cystic Lymphangiomas In Children.

Here, we describe our experience with different therapeutic modalities used to treat cystic lymphangiomas in children in our hospital, including single therapy with OK-432, bleomycin and surgery, and a combination of the three modalities. We performed a retrospective, cross-sectional study including patients treated from 1998 to 2011. The effects on macrocystic lymphangiomas and adverse reactions were evaluated. Twenty-nine children with cystic lymphangiomas without any previous treatment were included. Under general anesthesia, patients given sclerosing agents underwent puncture of the lesion (guided by ultrasound when necessary) and complete aspiration of the intralesional liquid. The patients were evaluated with ultrasound and clinical examinations for a maximum follow-up time of 4 years. The proportions of patients considered cured after the first therapeutic approach were 44% in the surgery group, 29% in the bleomycin group and 31% in the OK-432 group. These proportions were not significantly different. Sequential treatment increased the rates of curative results to 71%, 74% and 44%, respectively, after the final treatment, which in our case was approximately 1.5 applications per patient. The results of this study indicate that most patients with cystic lymphangiomas do not show complete resolution after the initial therapy, regardless of whether the therapy is surgical or involves the use of sclerosing agents. To achieve complete resolution of the lesions, either multiple operations or a combination of surgery and sclerotherapy must be used and should be tailored to the characteristics of each patient.