Use of the checkerboard DNA-DNA hybridization technique for bacteria detection in Aedes aegypti (Diptera:Culicidae) (L.)

<p>Abstract</p> <p>Background</p> <p>Bacteria associated with insects can have a substantial impact on the biology and life cycle of their host. The checkerboard DNA-DNA hybridization technique is a semi-quantitative technique that has been previously employed in odontology to detect and quantify a variety of bacterial species in dental samples. Here we tested the applicability of the checkerboard DNA-DNA hybridization technique to detect the presence of <it>Aedes aegypti</it>-associated bacterial species in larvae, pupae and adults of <it>A. aegypti</it>.</p> <p>Findings</p> <p>Using the checkerboard DNA-DNA hybridization technique we could detect and estimate the number of four bacterial species in total DNA samples extracted from <it>A. aegypti </it>single whole individuals and midguts. <it>A. aegypti </it>associated bacterial species were also detected in the midgut of four other insect species, <it>Lutzomyia longipalpis, Drosophila melanogaster</it>, <it>Bradysia hygida </it>and <it>Apis mellifera</it>.</p> <p>Conclusions</p> <p>Our results demonstrate that the checkerboard DNA-DNA hybridization technique can be employed to study the microbiota composition of mosquitoes. The method has the sensitivity to detect bacteria in single individuals, as well as in a single organ, and therefore can be employed to evaluate the differences in bacterial counts amongst individuals in a given mosquito population. We suggest that the checkerboard DNA-DNA hybridization technique is a straightforward technique that can be widely used for the characterization of the microbiota in mosquito populations.</p

El recien nacido con dolor: el papel del equipo de enfermeria

It's a quantitative, descriptive and cross study conducted with the nurse team working in the field of neonatology at the three hospitals, Alfenas-MG. Aimed to describe the forms of pain assessment of the newborn used by nurse team and analyze the practice of nursing as pain management of neonates. Data were collected through a semi-structured formulary, from August to September 2008, of 42 nurses working. The analyze was performed using SPSS software using descriptive statistics and correlation test. It was observed that the respondents recognize that the newborn is capable of feeling pain and evaluated by physiological and behavioral changes and that there isn't pain assessment scales standardized in the institutions. For the management, carry out pharmacological and nonpharmacological interventions. There is need to train professionals contributing to the assessment and management of pain, promoting of holistic care of the neonate.Estudio cuantitativo, descriptivo y transversal realizado con profesionales de Enfermería que actúan en el área de Neonatología en los tres hospitales de Alfenas, Minas Gerais. Se objetivó describir las formas de evaluación del dolor del recién nacido utilizados por el personal de enfermería y analizar la práctica de la enfermería como el manejo del dolor del neonato. La recolección de datos fue hecha por medio de un formulario semiestructurado, entre agosto y septiembre de 2008, con 42 profesionales. El análisis fue hecho con el software SPSS, utilizando la estadística descriptiva y la prueba de correlación. Los entrevistados reconocen que el neonato es capaz de sentir dolor y esto se evalúa a través de alteraciones fisiológicas y de comportamiento, no hay utilización de escalas de evaluación del dolor estandarizadas en las instituciones. Para el manejo, realizan intervenciones farmacológicas y no farmacológicas. Hay necesidad de capacitar profesionales contribuyendo para la evaluación y manejo del dolor, para la promoción del cuidado integral al neonato.Estudo quantitativo, descritivo e transversal realizado com profissionais de enfermagem que atuam em neonatologia nos três hospitais de Alfenas-MG. Objetivou descrever as formas de avaliação de dor do recém-nascido utilizadas pela equipe de enfermagem e analisar a prática da enfermagem quanto ao manejo da dor do neonato. A coleta de dados foi feita por meio de formulário semiestruturado, de agosto a setembro de 2008, com 42 profissionais. A análise foi feita pelo software SPSS utilizando estatística descritiva e teste de correlação. Os entrevistados acreditam que o recém-nascido é capaz de sentir dor e a avaliam por meio de alterações fisiológicas e comportamentais, e que não há utilização de escalas de avaliação álgica padronizadas nas instituições. Para o manejo, realizam intervenções farmacológicas e não farmacológicas. Há necessidade de capacitar os profissionais, contribuindo para a avaliação e o manejo da dor, e promovendo o cuidado integral ao neonato.Universidade de São Paulo Escola de Enfermagem de Ribeirao Preto Departamento de Enfermagem Materno-infantil e Saude PublicaUniversidade Federal de São Paulo (UNIFESP) Departamento de Enfermagem Pediatrica Escola Paulista de EnfermagemUniversidade Federal de Alfenas Escola de Enfermagem Programa de Pos-graduacao em EnfermagemUniversidade de São Paulo Escola de Enfermagem de Ribeirao Preto Departamento de Enfermagem Geral e EspecializadaUniversidade Federal de Alfenas Escola de EnfermagemPrograma de Pos-graduacao em Enfermagem Programa de Pos-graduacao em EnfermagemEscola de Enfermagem Escola de EnfermagemUniversidade de São PauloUNIFESP, Depto. de Enfermagem Pediatrica Escola Paulista de Enfermagem (EPE)SciEL

Rapid preparation of nuclei-depleted detergent-resistant membrane fractions suitable for proteomics analysis

<p>Abstract</p> <p>Background</p> <p>Cholesterol-rich membrane microdomains known as lipid rafts have been implicated in diverse physiologic processes including lipid transport and signal transduction. Lipid rafts were originally defined as detergent-resistant membranes (DRMs) due to their relative insolubility in cold non-ionic detergents. Recent findings suggest that, although DRMs are not equivalent to lipid rafts, the presence of a given protein within DRMs strongly suggests its potential for raft association in vivo. Therefore, isolation of DRMs represents a useful starting point for biochemical analysis of lipid rafts. The physicochemical properties of DRMs present unique challenges to analysis of their protein composition. Existing methods of isolating DRM-enriched fractions involve flotation of cell extracts in a sucrose density gradient, which, although successful, can be labor intensive, time consuming and results in dilute sucrose-containing fractions with limited utility for direct proteomic analysis. In addition, several studies describing the proteomic characterization of DRMs using this and other approaches have reported the presence of nuclear proteins in such fractions. It is unclear whether these results reflect trafficking of nuclear proteins to DRMs or whether they arise from nuclear contamination during isolation. To address these issues, we have modified a published differential detergent extraction method to enable rapid DRM isolation that minimizes nuclear contamination and yields fractions compatible with mass spectrometry.</p> <p>Results</p> <p>DRM-enriched fractions isolated using the conventional or modified extraction methods displayed comparable profiles of known DRM-associated proteins, including flotillins, GPI-anchored proteins and heterotrimeric G-protein subunits. Thus, the modified procedure yielded fractions consistent with those isolated by existing methods. However, we observed a marked reduction in the percentage of nuclear proteins identified in DRM fractions isolated with the modified method (15%) compared to DRMs isolated by conventional means (36%). Furthermore, of the 21 nuclear proteins identified exclusively in modified DRM fractions, 16 have been reported to exist in other subcellular sites, with evidence to suggest shuttling of these species between the nucleus and other organelles.</p> <p>Conclusion</p> <p>We describe a modified DRM isolation procedure that generates DRMs that are largely free of nuclear contamination and that is compatible with downstream proteomic analyses with minimal additional processing. Our findings also imply that identification of nuclear proteins in DRMs is likely to reflect legitimate movement of proteins between compartments, and is not a result of contamination during extraction.</p

A Selection Index for Gene Expression Evolution and Its Application to the Divergence between Humans and Chimpanzees

The importance of gene regulation in animal evolution is a matter of long-standing interest, but measuring the impact of selection on gene expression has proven a challenge. Here, we propose a selection index of gene expression as a straightforward method for assessing the mode and strength of selection operating on gene expression levels. The index is based on the widely used McDonald-Kreitman test and requires the estimation of four quantities: the within-species and between-species expression variances as well as the sequence heterozygosity and divergence of neutrally evolving sequences. We apply the method to data from human and chimpanzee lymphoblastoid cell lines and show that gene expression is in general under strong stabilizing selection. We also demonstrate how the same framework can be used to estimate the proportion of adaptive gene expression evolution