Reconhecendo a Microbiologia no nosso dia a dia pelo método PBL por estudantes do ensino médio

A microbiologia é a ciência que estuda os microrganismos e a relação destes com a nossa vida e meio ambiente. Entretanto, a microbiologia é um tema transmitido ao ensino médio, quase que exclusivamente de forma conceitual e teórica pelo professor, exigindo para sua compreensão, um alto ní­vel de abstração por parte do aluno, dificultando o processo de aprendizagem. O presente trabalho teve como objetivo aplicar a metodologia do PBL no ensino de microbiologia para o ensino médio, relacionando-a com o cotidiano dos estudantes. O PBL coloca o aluno como papel central na produção do conhecimento, visto que o mesmo deve procurar soluções para um problema que lhe é apresentado. O trabalho realizado fez parte do projeto de extensão "Reconhecendo a microbiologia no nosso dia a dia" , financiado pela Secretaria da Ciência, Tecnologia e Ensino Superior do Estado do Paraná (SETI-PR). Neste projeto atuaram docentes e discentes de pós-graduação (mestrado e doutorado) e graduação, abrangendo cursos de diferentes áreas (Artes visuais, Biologia, Biomedicina, Farmácia, entre outros). Foram alvo desta ação de extensão 13 turmas do ensino médio público, totalizando 272 estudantes, em três cidades do Paraná (Londrina, Tamarana e São Jerônimo da Serra). Primeiramente 10 diferentes tutoriais foram preparados e alguns destes foram aplicados nas escolas públicas. A metodologia PBL teve uma boa aceitação, tanto pelos estudantes quanto pelos professores. Sendo assim, a aplicação do método PBL para o ensino de Microbiologia para o Ensino Médio se mostrou possí­vel e eficiente. Os alunos de ensino básico adotaram atitudes independentes, reflexivas e crí­ticas, participando como protagonista na construção do próprio conhecimento. Deste modo, a metodologia dialética associada com aulas práticas estimulou o trabalho em equipe, contribuindo para formação de cidadãos mais conscientes em relação aos microrganismos, pois compreenderam o papel destes em diversos aspectos de sua vida, saúde e meio ambiente

Reconhecendo a Microbiologia no nosso dia a dia pelo método PBL por estudantes do ensino médio

A microbiologia é a ciência que estuda os microrganismos e a relação destes com a nossa vida e meio ambiente. Entretanto, a microbiologia é um tema transmitido ao ensino médio, quase que exclusivamente de forma conceitual e teórica pelo professor, exigindo para sua compreensão, um alto nível de abstração por parte do aluno, dificultando o processo de aprendizagem. O presente trabalho teve como objetivo aplicar a metodologia do PBL no ensino de microbiologia para o ensino médio, relacionando-a com o cotidiano dos estudantes. O PBL coloca o aluno como papel central na produção do conhecimento, visto que o mesmo deve procurar soluções para um problema que lhe é apresentado. O trabalho realizado fez parte do projeto de extensão “Reconhecendo a microbiologia no nosso dia a dia”, financiado pela Secretaria da Ciência, Tecnologia e Ensino Superior do Estado do Paraná (SETI-PR). Neste projeto atuaram docentes e discentes de pós-graduação (mestrado e doutorado) e graduação, abrangendo cursos de diferentes áreas (Artes visuais, Biologia, Biomedicina, Farmácia, entre outros). Foram alvo desta ação de extensão 13 turmas do ensino médio público, totalizando 272 estudantes, em três cidades do Paraná (Londrina, Tamarana e São Jerônimo da Serra). Primeiramente 10 diferentes tutoriais foram preparados e alguns destes foram aplicados nas escolas públicas. A metodologia PBL teve uma boa aceitação, tanto pelos estudantes quanto pelos professores. Sendo assim, a aplicação do método PBL para o ensino de Microbiologia para o Ensino Médio se mostrou possível e eficiente. Os alunos de ensino básico adotaram atitudes independentes, reflexivas e críticas, participando como protagonista na construção do próprio conhecimento. Deste modo, a metodologia dialética associada com aulas práticas estimulou o trabalho em equipe, contribuindo para formação de cidadãos mais conscientes em relação aos microrganismos, pois compreenderam o papel destes em diversos aspectos de sua vida, saúde e meio ambiente

Cloning, Expression, and Purification of Histidine-Tagged Escherichia coli Dihydrodipicolinate Reductase

This work was supported in part by funds from an Oklahoma State Regent Grant for Higher Education (021606), P20RR016478 grant from the National Center for Research Resources (NCRR) a component of National Institute of Health (NIH), and a grant from the University of Central Oklahoma office of Research and Grants to L.C.The enzyme dihydrodipicolinate reductase (DHDPR) is a component of the lysine biosynthetic pathway in bacteria and higher plants. DHDPR catalyzes the NAD(P)H dependent reduction of 2,3-dihydrodipicolinate to the cyclic imine L-2,3,4,5,-tetrahydropicolinic acid. The dapB gene that encodes dihydrodipicolinate reductase has previously been cloned, but the expression of the enzyme is low and the purification is time consuming. Therefore the E. coli dapB gene was cloned into the pET16b vector to improve the protein expression and simplify the purification. The dapB gene sequence was utilized to design forward and reverse oligonucleotide primers that were used to PCR the gene from Escherichia coli genomic DNA. The primers were designed with NdeI or BamHI restriction sites on the 5’and 3’ terminus respectively. The PCR product was sequenced to confirm the identity of dapB. The gene was cloned into the expression vector pET16b through NdeI and BamHI restriction endonuclease sites. The resulting plasmid containing dapB was transformed into the bacterial strain BL21 (DE3). The transformed cells were utilized to grow and express the histidine-tagged reductase and the protein was purified using Ni-NTA affinity chromatography. SDS/PAGE gel analysis has shown that the protein was 95% pure and has approximate subunit molecular weight of 28 kDa. The protein purification is completed in one day and 3 liters of culture produced approximately 40–50 mgs of protein, an improvement on the previous protein expression and multistep purification.Yeshttp://www.plosone.org/static/editorial#pee

Multivariate meta-analysis of the association of G-protein beta 3 gene (GNB3) haplotypes with cardiovascular phenotypes

The objective of the present study was to review previous investigations on the association of haplotypes in the G-protein β3 subunit (GNB3) gene with representative cardiovascular risk factors/phenotypes: hypertension, overweight, and variation in the systolic and diastolic blood pressures (SBP and DBP, respectively) and as well as body mass index (BMI). A comprehensive literature search was undertaken in Pubmed, Web of Science, EMBASE, Biological Abstracts, LILACS and Google Scholar to identify potentially relevant articles published up to April 2011. Six genetic association studies encompassing 16,068 participants were identified. Individual participant data were obtained for all studies. The three most investigated GNB3 polymorphisms (G-350A, C825T and C1429T) were considered. Expectation–maximization and generalized linear models were employed to estimate haplotypic effects from data with uncertain phase while adjusting for covariates. Study-specific results were combined through a random-effects multivariate meta-analysis. After carefully adjustments for relevant confounding factors, our analysis failed to support a role for GNB3 haplotypes in any of the investigated phenotypes. Sensitivity analyses excluding studies violating Hardy–Weinberg expectations, considering gender-specific effects or more extreme phenotypes (e.g. obesity only) as well as a fixed-effects “pooled” analysis also did not disclose a significant influence of GNB3 haplotypes on cardiovascular phenotypes. We conclude that the previous cumulative evidence does not support the proposal that haplotypes formed by common GNB3 polymorphisms might contribute either to the development of hypertension and obesity, or to the variation in the SBP, DBP and BMI.This work was supported by the Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP, Brazil, to T.V.P.). This work was also supported in part by the Global Center of Excellence Program (No. F03, to M.D.) founded by the Japan Society for the Promotion of Science, Japan (to Y.S.) and Grants-in-Aid for Scientific Research from the Ministry of Education, Culture, Sports, Science, and Technology of Japan (Numbers 18209023, 18018021, and 19659149 to Y.Y.)

Population Genetics of GYPB and Association Study between GYPB*S/s Polymorphism and Susceptibility to P. falciparum Infection in the Brazilian Amazon

Merozoites of Plasmodium falciparum invade through several pathways using different RBC receptors. Field isolates appear to use a greater variability of these receptors than laboratory isolates. Brazilian field isolates were shown to mostly utilize glycophorin A-independent invasion pathways via glycophorin B (GPB) and/or other receptors. The Brazilian population exhibits extensive polymorphism in blood group antigens, however, no studies have been done to relate the prevalence of the antigens that function as receptors for P. falciparum and the ability of the parasite to invade. Our study aimed to establish whether variation in the GYPB*S/s alleles influences susceptibility to infection with P. falciparum in the admixed population of Brazil.Two groups of Brazilian Amazonians from Porto Velho were studied: P. falciparum infected individuals (cases); and uninfected individuals who were born and/or have lived in the same endemic region for over ten years, were exposed to infection but have not had malaria over the study period (controls). The GPB Ss phenotype and GYPB*S/s alleles were determined by standard methods. Sixty two Ancestry Informative Markers were genotyped on each individual to estimate admixture and control its potential effect on the association between frequency of GYPB*S and malaria infection.GYPB*S is associated with host susceptibility to infection with P. falciparum; GYPB*S/GYPB*S and GYPB*S/GYPB*s were significantly more prevalent in the in the P. falciparum infected individuals than in the controls (69.87% vs. 49.75%; P<0.02). Moreover, population genetics tests applied on the GYPB exon sequencing data suggest that natural selection shaped the observed pattern of nucleotide diversity.Epidemiological and evolutionary approaches suggest an important role for the GPB receptor in RBC invasion by P. falciparum in Brazilian Amazons. Moreover, an increased susceptibility to infection by this parasite is associated with the GPB S+ variant in this population

Identification of Close Relatives in the HUGO Pan-Asian SNP Database

The HUGO Pan-Asian SNP Consortium has recently released a genome-wide dataset, which consists of 1,719 DNA samples collected from 71 Asian populations. For studies of human population genetics such as genetic structure and migration history, this provided the most comprehensive large-scale survey of genetic variation to date in East and Southeast Asia. However, although considered in the analysis, close relatives were not clearly reported in the original paper. Here we performed a systematic analysis of genetic relationships among individuals from the Pan-Asian SNP (PASNP) database and identified 3 pairs of monozygotic twins or duplicate samples, 100 pairs of first-degree and 161 second-degree of relationships. Three standardized subsets with different levels of unrelated individuals were suggested here for future applications of the samples in most types of population-genetics studies (denoted by PASNP1716, PASNP1640 and PASNP1583 respectively) based on the relationships inferred in this study. In addition, we provided gender information for PASNP samples, which were not included in the original dataset, based on analysis of X chromosome data

Population Genetic Structure of Peninsular Malaysia Malay Sub-Ethnic Groups

Patterns of modern human population structure are helpful in understanding the history of human migration and admixture. We conducted a study on genetic structure of the Malay population in Malaysia, using 54,794 genome-wide single nucleotide polymorphism genotype data generated in four Malay sub-ethnic groups in peninsular Malaysia (Melayu Kelantan, Melayu Minang, Melayu Jawa and Melayu Bugis). To the best of our knowledge this is the first study conducted on these four Malay sub-ethnic groups and the analysis of genotype data of these four groups were compiled together with 11 other populations' genotype data from Indonesia, China, India, Africa and indigenous populations in Peninsular Malaysia obtained from the Pan-Asian SNP database. The phylogeny of populations showed that all of the four Malay sub-ethnic groups are separated into at least three different clusters. The Melayu Jawa, Melayu Bugis and Melayu Minang have a very close genetic relationship with Indonesian populations indicating a common ancestral history, while the Melayu Kelantan formed a distinct group on the tree indicating that they are genetically different from the other Malay sub-ethnic groups. We have detected genetic structuring among the Malay populations and this could possibly be accounted for by their different historical origins. Our results provide information of the genetic differentiation between these populations and a valuable insight into the origins of the Malay sub-ethnic groups in Peninsular Malaysia

Genetic risk factors associated with essential hypertension in the quilombo populations from Ribeira River Valley - São Paulo, Brazil

A hipertensão essencial, um problema de saúde pública mundial, é uma doença multifatorial, cujo componente genético varia entre 25 a 60%. A despeito da alta prevalência e incidência da hipertensão essencial em populações de ancestralidade africana, os estudos sobre o componente genético da hipertensão essencial têm sido primariamente realizados em populações de ancestralidade européia ou asiática. Dessa maneira, o objetivo geral deste trabalho foi investigar e quantificar potenciais fatores de risco genéticos associados à hipertensão essencial em populações dos remanescentes de quilombos do Vale do Ribeira SP, indivíduos afro-descendentes, oriundos de populações parcialmente isoladas, previamente bem caracterizadas do ponto de vista clínico, genealógico e genético-populacional. A amostra foi constituída de 759 indivíduos adultos com estudo clínico e antropométrico oriundos das seguintes populações quilombolas: Abobral, Galvão, São Pedro, Pedro Cubas, André Lopes, Nhunguara, Sapatu, Pilões, Ivaporunduva, Maria Rosa, Poça e Reginaldo, localizadas no Vale do Ribeira, no estado de São Paulo. Estimativas de ancestralidade (baseadas em 48 marcadores INDEL autossômicos) sugerem que a contribuição africana, européia e ameríndia na amostra estudada é de aproximadamente 39,7%, 39,0% e 21,3%, respectivamente (N=307), sendo igualmente distribuídas em normotensos e hipertensos (37,6%, 39,0% e 19,9% e 41,1%, 37,8% e 20,4%, P = 0,845). O estudo dos fatores de risco genéticos associados à hipertensão foi realizado por duas abordagens: (i) um estudo transversal com indivíduos não aparentados (N = 383) e (ii) um estudo de associação baseado em famílias (N total = 759, 79 famílias). Foram selecionados para estes estudos sete polimorfismos (I/D, Glu298Asp, C825T, G-350A, M235T, A/C, A486V) em seis genes candidatos com forte base funcional: ACE, NOS3, GNB3, AGT, ADD2 e GRK4. As análises do estudo transversal revelaram uma associação significativa entre o polimorfismo C825T do GNB3 e a hipertensão essencial (P=0,036), quando consideramos o fenótipo mais extremo (PAS maior igaul a 160mmHg e/ou PAD maior igual a 95mmHg, ou uso de medicamentos anti-hipertensivos). Análises haplotípicas do GNB3 (variantes G-350A e C825T) revelaram que cada cópia do haplótipo C825/G-350 aumenta em 1,73 a chance de hipertensão em relação ao haplótipo basal T825/G-350 (OR = 1,73; IC 95% = 1,05 2,84, P = 0,029). O haplótipo C825/G-350 estaria relacionado com um aumento na média esperada da PAS e da PAD (Delta = 6,07 mmHg; IC de 95%: 1,64 10,50 mmHg; P = 0,007 e Delta = 3.75 mmHg; IC de 95%: 1,16 6,35 mmHg; P = 0,005, respectivamente). Estudos de interação gene-gene também foram conduzidos pela técnica de redução de dimensionalidade de múltiplos fatores generalizada (GMDR). Os modelos de predição por GMDR revelaram interações de alta ordem entre as variantes avaliadas e o risco de hipertensão essencial. Os melhores modelos de interação foram: NOS3/GRK4, ACE/GNB3/AGT e ACE/GNB3/AGT/ADD2/GRK4, com uma acurácia média balanceada de teste de 60% (P=0,006), 59% (P =0,019), e 59% (P = 0,025), respectivamente. As análises de segregação em famílias indicaram uma associação significativa entre o polimorfismo G-350A do gene GNB3 e a hipertensão essencial, quando consideramos o fenótipo classificado de acordo com o critério da OMS (PAS maior igual a 140mmHg e/ou PAD maior igual a 90mmHg, ou uso de medicamentos anti-hipertensivos, P = 0,018), e também quando consideramos o fenótipo mais extremo (PAS maior igual a 160mmHg e/ou PAD maior igual a 95mmHg, ou uso de medicamentos anti-hipertensivos, P = 0,001). Finalmente, as estimativas da herdabilidade corroboram a noção de que o componente genético da pressão arterial é elevado nessas populações: herdabilidades (ajustadas para idade, gênero e IMC) de 36,1 e 42.9% para PAS e PAD, respectivamente. Apresentamos nesse trabalho uma investigação extensiva e original sobre o componente genético da hipertensão essencial em uma população brasileira de afrodescendentes (com características históricas, genéticas e ambientais peculiares), reforçando a hipótese de um mecanismo poligênico de regulação da pressão arterial nas populações remanescentes de quilombos do Vale do Ribeira - SP.Essential hypertension is a worldwide public health problem. Although this pathology has a multi-factorial etiology, the heritability of blood pressure is high, ranging from 25 to 60%. Despite the high prevalence and incidence of essential hypertension among subjects of African ancestry, most genetic association studies has been carried out either in subjects with European descent or in Asian-derived individuals. Thus, the aim of this study was to investigate genetic risk factors putatively associated with essential hypertension in the quilombo populations, which are partially isolated African-derived populations, well characterized in terms of clinical, genealogical and population data, living in the Ribeira River Valley São Paulo, Brazil. A total of 759 adults with clinical and anthropometric data sampled from the 12 populations (Abobral, Galvão, São Pedro, Pedro Cubas, André Lopes, Nhunguara, Sapatu, Pilões, Ivaporunduva, Maria Rosa, Poça and Reginaldo) were studied. Ancestry estimates (based on 48 ancestry informative INDEL markers) suggested that the African, European and Amerindian contributions were 39.7%, 39.0% and 21.3% respectively (N = 307) to the studied sample. These proportions did not differ significantly between hypertensive and normotensive subjects(41.1%, 37.8% , 20.4% and 37.6%, 39.0% e 19.9%, respectively; P = 0.845). Two approaches were employed to investigate potential genetic risk factors for essential hypertension: (i) a cross-sectional study with unrelated individuals (N = 383) and (ii) a family-based association study (N = 759, 79 families). Seven polymorphisms (I/D, Glu298Asp, C825T, G-350A, M235T, A/C, A486V) in six candidate genes with strong functional basis (ACE, NOS3, GNB3, AGT, ADD2 and GRK4) were selected. The cross-sectional study revealed a significant association between the C825T GNB3 polymorphism and essential hypertension (P = 0.036) defined as a SBP > or = 160 mmHg and/or DBP > or = 95 mmHg, or use antihypertensive drugs). Haplotypic analyses of both G-350A and C825T GNB3 variants indicated that each copy of the haplotype C825/G-350 increases in 1.73 the odds of hypertension compared to the reference T825/G-350 haplotype (OR = 1.73, CI 95% = 1.05 - 2.84, P = 0.029). The haplotype C825/G-350 was also associated with augmented levels of both SBP and DBP (expected mean difference, Delta = 6.7, 95% CI: 1.64 10.50, P = 0.007 and Delta = 3.75, CI 95%: 1.16 6.35, P = 0.005, respectively) compared to the reference haplotype. Gene-gene interactions were also investigated by the use of the generalized multifactor dimensionality reduction (GMDR). GMDR prediction models revealed high-order interactions for hypertension between and among the studied variants. The best interaction models were NOS3/GRK4, ACE/GNB3/AGT, and ACE/GNB3/AGT/ADD2/GRK4, with an average testing balanced accuracy of 60% (P = 0.006) 59% (P = 0.019), and 59% (P = 0.025), respectively. The family-based association study showed a significant association of the GNB3 G-350 allele with essential hypertension (P = 0.018) when the diagnosis of essential hypertension was based on the WHO criteria (SBP > or = 140mmHg and / or DBP > or = 90 mmHg, or use of antihypertensive drugs, P = 0.018). Qualitatively analogous results were observed when a more extreme phenotype was considered (i.e. hypertension defined as a SBP > or = 160mmHg and / or DBP > or = 95 mmHg, or use of antihypertensive drugs, P = 0.001) Finally, heritability estimates corroborated the notion that the genetic component of blood pressure is high in these populations. Heritability (adjusted for age, gender and BMI) estimates of 36.1 and 42.9% for SBP and DBP, respectively, were obtained. We presented in this work an extensive and original investigation about the genetic component of essential hypertension in an African-Brazilian population (with peculiar historical, genetical and environmental characteristics). This study strengthens the hypothesis of a polygenic mechanism of regulation of blood pressure in the quilombos from Ribeira River Valley São Paulo