Use of reverse transcriptase polymerase chain reaction (RT-PCR) in molecular screening of Newcastle disease virus in poultry and free-living bird populations

The aim of this study was to evaluate a simple molecular method of reverse transcriptase polymerase chain reaction (RT-PCR) to differentiate Newcastle disease virus strains according to their pathogenicity, in order to use it in molecular screening of Newcastle disease virus in poultry and free-living bird populations. Specific primers were developed to differentiate LaSota-LS-(vaccine strain) and Sao Joao do Meriti-SJM-strain (highly pathogenic strain). Chickens and pigeons were experimentally vaccinated/infected for an in vivo study to determine virus shedding in feces. Validation of sensitivity and specificity of the primers (SJM and LS) by experimental models used in the present study and results obtained in the molecular analysis of the primers by BLAST made it possible to generalize results. The development of primers that differentiate the level of pathogenicity of NDV stains is very important, mainly in countries where real-time RT-PCR is still not used as a routine test. These primers were able to determine the presence of the agent and to differentiate it according to its pathogenicity. © 2012 Springer Science+Business Media B.V.

Exequibility of differential gene expression analysis by DDRT-PCR in murine bone marrow cells

Model of study: Experimental study. Introduction: Recently, stem cell research has generated great interest due to its applicability in regenerative medicine. Bone marrow is considered the most important source of adult stem cells and the establishment of new methods towards gene expression analysis regarding stem cells has become necessary. Thus Differential Display Reverse Transcription Polymerase Chain Reaction (DDRT-PCR) may be an accessible tool to investigate small differences in the gene expression of different stem cells in distinct situations. Aim: In the present study, we investigated the exequibility of DDRT-PCR to identify differences in global gene expression of mice bone marrow cells under two conditions. Methods: First, bone marrow cells were isolated fresh and a part was cultivated during one week without medium replacement. Afterwards, both bone marrow cells (fresh and cultivated) were submitted to gene expression analyses by DDRT-PCR. Results: Initially, it was possible to observe in one week-cultured bone marrow cells, changes in morphology (oval cells to fibroblastic-like cells) and protein profile, which was seen through differences in band distribution in SDS-Page gels. Finally through gene expression analysis, we detected three bands (1300, 1000 and 225 bp) exclusively expressed in the fresh bone marrow group and two bands (400 and 300 bp) expressed specifically in the cultivated bone marrow cell group. Conclusions: In summary, the DDRT-PCR method was proved efficient towards the identification of small differences in gene expression of bone marrow cells in two defined conditions. Thus, we expect that DDRT-PCR can be fast and efficiently designed to analyze differential gene expression in several stem cell types under distinct conditions.

An Integrative Genomic and Transcriptomic Analysis Reveals Potential Targets Associated with Cell Proliferation in Uterine Leiomyomas

Background: Uterine Leiomyomas (ULs) are the most common benign tumours affecting women of reproductive age. ULs represent a major problem in public health, as they are the main indication for hysterectomy. Approximately 40-50% of ULs have non-random cytogenetic abnormalities, and half of ULs may have copy number alterations (CNAs). Gene expression microarrays studies have demonstrated that cell proliferation genes act in response to growth factors and steroids. However, only a few genes mapping to CNAs regions were found to be associated with ULs. Methodology: We applied an integrative analysis using genomic and transcriptomic data to identify the pathways and molecular markers associated with ULs. Fifty-one fresh frozen specimens were evaluated by array CGH (JISTIC) and gene expression microarrays (SAM). The CONEXIC algorithm was applied to integrate the data. Principal Findings: The integrated analysis identified the top 30 significant genes (P<0.01), which comprised genes associated with cancer, whereas the protein-protein interaction analysis indicated a strong association between FANCA and BRCA1. Functional in silico analysis revealed target molecules for drugs involved in cell proliferation, including FGFR1 and IGFBP5. Transcriptional and protein analyses showed that FGFR1 (P = 0.006 and P<0.01, respectively) and IGFBP5 (P = 0.0002 and P = 0.006, respectively) were up-regulated in the tumours when compared with the adjacent normal myometrium. Conclusions: The integrative genomic and transcriptomic approach indicated that FGFR1 and IGFBP5 amplification, as well as the consequent up-regulation of the protein products, plays an important role in the aetiology of ULs and thus provides data for potential drug therapies development to target genes associated with cellular proliferation in ULs. © 2013 Cirilo et al.

Factors affecting the adoption of green supply chain management practices in Brazil: Empirical evidence

The aim of this study is to identify and analyse the factors that affect the adoption of Green Supply Chain Management practices based on empirical evidence from the Brazilian electronics sector. Data are collected in a survey of 100 electronics companies and analysed using statistical analysis of variance and regression methods. The study finds that the size of the company, previous experience with Environmental Management Systems, and the use of hazardous inputs are positively correlated with GSCM practices adoption. Surprisingly, formal pressure from the stronger tier/player in the supply chain is not correlated with the adoption of GSCM practices. Finally, we present some explanations for these findings and suggestions for future research. © 2013 Copyright Taylor and Francis Group, LLC.

Extensive impact of saturated fatty acids on metabolic and cardiovascular profile in rats with diet-induced obesity: A canonical analysis

Background: Although hypercaloric interventions are associated with nutritional, endocrine, metabolic, and cardiovascular disorders in obesity experiments, a rational distinction between the effects of excess adiposity and the individual roles of dietary macronutrients in relation to these disturbances has not previously been studied. This investigation analyzed the correlation between ingested macronutrients (including sucrose and saturated and unsaturated fatty acids) plus body adiposity and metabolic, hormonal, and cardiovascular effects in rats with diet-induced obesity. Methods: Normotensive Wistar-Kyoto rats were submitted to Control (CD; 3.2 Kcal/g) and Hypercaloric (HD; 4.6 Kcal/g) diets for 20 weeks followed by nutritional evaluation involving body weight and adiposity measurement. Metabolic and hormonal parameters included glycemia, insulin, insulin resistance, and leptin. Cardiovascular analysis included systolic blood pressure profile, echocardiography, morphometric study of myocardial morphology, and myosin heavy chain (MHC) protein expression. Canonical correlation analysis was used to evaluate the relationships between dietary macronutrients plus adiposity and metabolic, hormonal, and cardiovascular parameters. Results: Although final group body weights did not differ, HD presented higher adiposity than CD. Diet induced hyperglycemia while insulin and leptin levels remained unchanged. In a cardiovascular context, systolic blood pressure increased with time only in HD. Additionally, in vivo echocardiography revealed cardiac hypertrophy and improved systolic performance in HD compared to CD; and while cardiomyocyte size was unchanged by diet, nuclear volume and collagen interstitial fraction both increased in HD. Also HD exhibited higher relative β-MHC content and β/α-MHC ratio than their Control counterparts. Importantly, body adiposity was weakly associated with cardiovascular effects, as saturated fatty acid intake was directly associated with most cardiac remodeling measurements while unsaturated lipid consumption was inversely correlated with these effects. Conclusion: Hypercaloric diet was associated with glycemic metabolism and systolic blood pressure disorders and cardiac remodeling. These effects directly and inversely correlated with saturated and unsaturated lipid consumption, respectively. © 2013 Oliveira Junior et al.; licensee BioMed Central Ltd.

Nucleotide and phylogenetic analysis of human papillomavirus types 6 and 11 isolated from recurrent respiratory papillomatosis in Brazil

There are few studies about the distribution of natural molecular variants of low-risk HPVs. Our aim was to evaluate the E6 early gene variability among HPV-6 and HPV-11 isolates detected in recurrent respiratory papillomatosis (RRP) samples obtained in a cohort of Brazilian patients. We also performed a phylogenetic analysis in order to compare nucleotide sequences identified in our study with previously reported isolates from different anatomic sites (laryngeal papillomas, genital warts, cervical cancer and anal swabs) obtained from other parts of the world to determine the phylogenetic relationships of variants detected in Brazil. The complete coding region of the E6 gene of 25 samples was cloned and sequenced: 18 isolates of HPV-6 (72%) and 7 isolates of HPV-11 (28%). A total of four different HPV-6 genomic variants and two HPV-11 genomic variants was identified. It was not possible to correlate specific variants with disease severity. Phylogenetic trees for both HPV types were constructed enclosing both E6 sequences detected in our study and formerly published sequences. In both phylogenetic trees, the sequences from Brazil did not group together. We could not establish a geographical association between HPV-6 or HPV-11 variants, unlike HPV-16 and HPV-18. © 2013 Elsevier B.V.

A bayesian analysis for the parameters of the exponential-logarithmic distribution

The exponential-logarithmic is a new lifetime distribution with decreasing failure rate and interesting applications in the biological and engineering sciences. Thus, a Bayesian analysis of the parameters would be desirable. Bayesian estimation requires the selection of prior distributions for all parameters of the model. In this case, researchers usually seek to choose a prior that has little information on the parameters, allowing the data to be very informative relative to the prior information. Assuming some noninformative prior distributions, we present a Bayesian analysis using Markov Chain Monte Carlo (MCMC) methods. Jeffreys prior is derived for the parameters of exponential-logarithmic distribution and compared with other common priors such as beta, gamma, and uniform distributions. In this article, we show through a simulation study that the maximum likelihood estimate may not exist except under restrictive conditions. In addition, the posterior density is sometimes bimodal when an improper prior density is used. © 2013 Copyright Taylor and Francis Group, LLC.

Analysis of inteins in the Candida parapsilosis complex for simple and accurate species identification

Inteins are coding sequences that are transcribed and translated with flanking sequences and then are excised by an autocatalytic process. There are two types of inteins in fungi, mini-inteins and full-length inteins, both of which present a splicing domain containing well-conserved amino acid sequences. Full-length inteins also present a homing endonuclease domain that makes the intein a mobile genetic element. These parasitic genetic elements are located in highly conserved genes and may allow for the differentiation of closely related species of the Candida parapsilosis (psilosis) complex. The correct identification of the three psilosis complex species C. parapsilosis, Candida metapsilosis, and Candida orthopsilosis is very important in the clinical setting for improving antifungal therapy and patient care. In this work, we analyzed inteins that are present in the vacuolar ATPase gene VMA and in the threonyl-tRNA synthetase gene ThrRS in 85 strains of the Candida psilosis complex (46 C. parapsilosis, 17 C. metapsilosis, and 22 C. orthopsilosis). Here, we describe an accessible and accurate technique based on a single PCR that is able to differentiate the psilosis complex based on the VMA intein. Although the ThrRS intein does not distinguish the three species of the psilosis complex by PCR product size, it can differentiate them by sequencing and phylogenetic analysis. Furthermore, this intein is unusually present as both mini- and full-length forms in C. orthopsilosis. Additional population studies should be performed to address whether this represents a common intraspecific variability or the presence of subspecies within C. orthopsilosis. Copyright © 2013, American Society for Microbiology. All Rights Reserved.

Reação de 16 porta-enxertos sob laranjeira 'Valência' ao agente causal, Candidatus Liberibacter asiaticus

Pós-graduação em Agronomia (Genética e Melhoramento de Plantas) - FCAV

Caracterização morfométrica, patogênica e genética de isolados de Colletrotrichum gloeosporioides, agente causal da antracnose em manga (Mangifera indica L.)

Pós-graduação em Agronomia (Produção Vegetal) - FCAV

Podridão floral dos citros: variabilidade, sobrevivência e controle do agente causal, Colletotrichum acutatum

Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)

Fenômenos emocionais no contexto explicativo do modo causal de seleção por conseqüências

B. F. Skinner deu início ao behaviorismo radical, enquanto filosofia de uma ciência do comportamento, e reuniu os argumentos experimentais e teóricos que fundamentaram tal ciência. A proposta skinneriana distinguiu-se da psicologia da primeira metade do século XX por instituir o monismo físico como visão de homem e recomendar a abordagem de respostas abertas e encobertas no contexto de relações indivíduoambiente. No entanto, a adoção do modo causal de seleção por conseqüências para a explicação do comportamento pode ser tida como controversa no âmbito de análise de fenômenos emocionais, por estes apresentarem componentes operantes, mas também respondentes. Tendo em vista que a seleção ao nível filogenético permite a compreensão do estabelecimento de relações respondentes incondicionadas, objetivou-se analisar até que ponto um modelo selecionista permite a explicação de relações respondentes condicionadas, nos casos referentes a fenômenos emocionais. A investigação de elaborações da análise do comportamento resultou na proposição de um modelo interpretativo de fenômenos emocionais por meio de inter-relações entre processos respondentes e operantes. A coerência interna do sistema explicativo skinneriano é preservada por manter-se o pressuposto básico de que, em se tratando da ontogênese, relações historicamente estabelecidas com o ambiente explicam a ocorrência de respostas abertas, encobertas, respondentes, operantes ou com ambos os componentes. Considerando que clareza conceitual tende a ser um pré-requisito para o desenvolvimento de alternativas de intervenção, contrastou-se o modelo interpretativo proposto ao contexto de realização de análises funcionais na literatura da análise do comportamento. Preliminarmente, tal modelo demonstrou ser uma ferramenta útil, na medida em que favorece a compreensão de diferentes funções das variáveis que participam das relações comportamentais em foco, de uma perspectiva contextualizada e histórica.

Molecular analysis of three FUT3 gene single nucleotide polymorphisms and their relationship with the lewis erythrocytary phenotype in a human population of japanese-ancestry living in Tomé Açu, a town in the Brazilian Amazon

The Lewis blood group system involves two major antigens, Le^ª and Le^b. Their antigenic determinants are not primary gene products but are synthesized by the transfer of sugar subunits to a precursory chain by a specific enzyme which is the product of the FUT3 gene (Lewis gene). The presence of three FUT3 gene single nucleotide polymorphisms (SNPs) (59T > G; 508G > A and 1067T > A) was related to the Lewis phenotype of erythrocytes from 185 individuals of Japanese ancestry living in the town of Tomé-Açu in the Brazilian Amazon region. This relationship was detected using a serological hemagglutination test and the Dot-ELISA assay along with the molecular technique PCR-RFLP. We found that the three SNPs investigated in this study only accounted for a proportion of the Lewis-negative phenotype of the erythrocytes.

Molecular analysis of three FUT3 gene single nucleotide polymorphisms and their relationship with the lewis erythrocytary phenotype in a human population of japanese-ancestry living in Tomé Açu, a town in the Brazilian Amazon

The Lewis blood group system involves two major antigens, Le^a and Le^b. Their antigenic determinants are not primary gene products but are synthesized by the transfer of sugar subunits to a precursory chain by a specific enzyme which is the product of the FUT3 gene (Lewis gene). The presence of three FUT3 gene single nucleotide polymorphisms (SNPs) (59T > G; 508G > A and 1067T > A) was related to the Lewis phenotype of erythrocytes from 185 individuals of Japanese ancestry living in the town of Tomé-Açu in the Brazilian Amazon region. This relationship was detected using a serological hemagglutination test and the Dot-ELISA assay along with the molecular technique PCR-RFLP. We found that the three SNPs investigated in this study only accounted for a proportion of the Lewis-negative phenotype of the erythrocytes.

Bacterial Pathogens Related to Chronic Suppurative Otitis Media in Individuals With Cleft Palate: Bacteriological Culture and Polymerase Chain Reaction

Objective: To characterize the microbial etiology of chronic suppurative otitis media comparing the methods of classical bacteriological culture and polymerase chain reaction.Design/Setting/Patients: Bacteriological analysis by classical culture and by molecular polymerase chain reaction of 35 effusion otitis samples from patients with cleft lip and palate attending the Hospital for Rehabilitation of Craniofacial Anomalies of the University of Sao Paulo, Bauru, Brazil.Interventions: Collection of clinical samples of otitis by effusion through the external auditory tube.Main Outcome Measure: Otolaryngologic diagnosis of chronic suppurative otitis media.Results: Positive cultures were obtained from 83% of patients. Among the 31 bacterial lineages the following were isolated. In order of decreasing frequency: Pseudomonas aeruginosa (54.9%), Staphylococcus aureus (25.9%), and Enterococcus faecalis (19.2%). No anaerobes were isolated by culture. The polymerase chain reaction was positive for one or more bacteria investigated in 97.1% of samples. Anaerobe lineages were detected by the polymerase chain reaction method, such as Fusobacterium nucleatum, Bacteroides fragilis, and Peptostreptococcus anaerobius.Conclusions: Patients with cleft lip and palate with chronic suppurative otitis media presented high frequency of bacterial infection in the middle ear. The classical bacteriological culture did not detect strict anaerobes, whose presence was identified by the polymerase chain reaction method.