RESUMO
Plasmodium vivax is the most common human malaria parasite in Asian countries including Pakistan. Present study was designed to explore the genetic diversity of plasmodium vivax genotypes based on Pvmsp-3α and Pvmsp-3βgenes using allelic specific nested PCR and RFLP assays markers from field isolates in district Mardan, Pakistan. Blood samples of 200 P. vivax malarial patients were collected after taking their written informed consent. Genetic diversity in nested PCR products was determined by Restriction Fragment Length Polymorphism (RFLP) utilizing Alu1 and PstI restriction enzymes for alpha and beta gene products digestion, respectively. For analysis the genetic diversity of the sub allelic variants of Pvmsp3α and Pvmsp3β genes, Chi-Square test was performed by utilizing Minitab programming software 18. The P value 0.05 was considered as statistically significant. For Pvmsp 3α genes after gel electrophoresis of digested products, four distinct genotypes were obtained from total of 50 samples; type A: 35 (70%) (1.5-2.0 kb), 12 of type B (24%) (1.5-1.7 kb), 2 of type C (4%) (0.5-1.5) and one for type D (2%) (0.5-0.65 kb) which could be characterized into 9 allelic pattern (A1-A4, B1-B3, C1, D), in which A3 remained the most predominant. For Pvmsp-3βgenes, three distinct genotypes were obtained from 50 samples; 40(80%) of type A (1.5-2.5 kb), 9 (18%) of type B (1.0-1.5kb) and 1(2%) of type C (0.65 kb) which could be characterized into 6 allelic patterns (A1-A3, B1-B2, and C1). Most dominant one in Type A was A1 alleles which were noted (46%), while in Type B, the most dominant were B1 (10%).This study is the first ever report of molecular epidemiology and genetic variation in Pvmsp-3α and Pvmsp-3β genes of P. vivax isolates by using PCR/RFLP from District Mardan and [...].
O Plasmodium vivax é o parasita da malária humana mais comum nos países asiáticos, incluindo o Paquistão. O presente estudo foi desenhado para explorar a diversidade genética de genótipos de Plasmodium vivax baseados nos genes Pvmsp-3α e Pvmsp-3β, usando marcadores de ensaios alélicos nested PCR e RFLP de isolados de campo no distrito de Mardan, Paquistão. Amostras de sangue de 200 pacientes com malária por P. vivax foram coletadas após assinatura do termo de consentimento livre e esclarecido. A diversidade genética em produtos de PCR nested foi determinada por polimorfismo de fragmento de restrição (RFLP) utilizando as enzimas de restrição Alu1 e PstI para a digestão dos produtos dos genes alfa e beta, respectivamente. Para análise da diversidade genética das variantes subalélicas dos genes Pvmsp3α e Pvmsp3β, o teste Qui-quadrado foi realizado utilizando o software de programação Minitab 18. O valor P = 0,05 foi considerado estatisticamente significativo. Para os genes Pvmsp 3α, após eletroforese em gel de produtos digeridos, quatro genótipos distintos foram obtidos de um total de 50 amostras; tipo A: 35 (70%) (1,5-2,0 kb), 12 do tipo B (24%) (1,5-1,7 kb), 2 do tipo C (4%) (0,5-1,5) e um para o tipo D (2%) (0,5-0,65 kb), que podem ser caracterizados em nove padrões alélicos (A1-A4, B1-B3, C1, D), em que A3 permaneceu como o mais predominante. Para Pvmsp-3βgenes, três genótipos distintos foram obtidos a partir de 50 amostras; 40 (80%) do tipo A (1,5-2,5 kb), 9 (18%) do tipo B (1,0-1,5 kb) e 1 (2%) do tipo C (0,65 kb), que podem ser caracterizados em seis padrões alélicos (A1-A3, B1-B2 e C1). Os mais dominantes no tipo A foram o alelo A1, observados em 46%, enquanto, no tipo B, os mais dominantes foram B1 (10%). Este estudo é o primeiro relato de epidemiologia molecular e variação genética em Pvmsp-3α. Os genes Pvmsp-3β de isolados de P. vivax utilizando PCR/RFLP do Distrito Mardan mostraram um nível notável de diversidade genética nos genes estudados [...].
Assuntos
Humanos , Merozoítos , Plasmodium vivax/genética , Plasmodium vivax/parasitologia , Polimorfismo de Fragmento de Restrição/genética , Proteínas de Membrana/análise , Proteínas de Membrana/genéticaRESUMO
Molecular-typing can help in unraveling epidemiological scenarios and improvement for disease control strategies. A literature review of Mycobacterium tuberculosis transmission in Brazil through genotyping on 56 studies published from 1996-2019 was performed. The clustering rate for mycobacterial interspersed repetitive units - variable tandem repeats (MIRU-VNTR) of 1,613 isolates were: 73%, 33% and 28% based on 12, 15 and 24-loci, respectively; while for RFLP-IS6110 were: 84% among prison population in Rio de Janeiro, 69% among multidrug-resistant isolates in Rio Grande do Sul, and 56.2% in general population in São Paulo. These findings could improve tuberculosis (TB) surveillance and set up a solid basis to build a database of Mycobacterium genomes.
Assuntos
Humanos , Polimorfismo de Fragmento de Restrição/genética , Repetições Minissatélites/genética , Mycobacterium tuberculosis/genética , Brasil/epidemiologia , Técnicas de Tipagem Bacteriana , Epidemiologia Molecular , Sequenciamento Completo do Genoma , Genótipo , Mycobacterium tuberculosis/isolamento & purificaçãoRESUMO
ABSTRACT Objective Methylenetetrahydrofolate reductase (MTHFR) is involved in DNA methylation that is associated with autoimmune pathology. We investigated the association between MTHFR genetic polymorphisms at g.677C>T and g.1298A>C and their haplotypes, and the risk of thyroid dysfunction among Jordanian females. Subjects and methods A case-control study involving 98 hypothyroidism cases, 66 hyperthyroidism cases and 100 controls was conducted. Polymerase chain reaction/restriction fragment length polymorphism technique was performed to determine genotypes. Statistical analysis using SPSS software was performed. Results Genetic analysis showed a significant difference in genotype frequency of g.1298A>C between cases, and controls [hypothyroidism: AA (45.9%), AC (37.8%), CC (16.3%); hyperthyroidism: AA (9.1%), AC (69.7%), CC (21.2%); controls: AA (37.8%), AC (29.6%), CC (32.7%); CChypo vs. AAhypo: 2.55, 95% CI: (1.18-5.52); OR at least on Chypo: 1.79, 95% CI: (1.07-2.99)]; CChyper vs. AAhyper: 4.01, 95% CI: (1.79-9.01); OR at least on Chyper: 0.18, 95% CI: (0.07-0.48)]. There was no significant difference in genotype frequency of g.677C>T between cases and controls [hypothyroidism: CC (50.0%), CT (32.7%), TT (17.3%); hyperthyroidism: CC (77.3%), CT (15.2%), TT (7.6%); controls: CC (55.6%), CT (32.3%), TT (12.1%)]. There was a significant difference of MTHFR haplotypes among hypothyroidism cases and controls. TA and CC had a lower hypothyroidism risk whereas; TC showed a higher risk. Conclusions g.1298A>C genetic polymorphism of MTHFR may modulate the risk of thyroid disease. CC, TA, and TC haplotypes affect the risk of hypothyroidism. Larger samples should be included in the future to verify the role of MTHFR polymorphisms in thyroid diseases.
Assuntos
Humanos , Feminino , Adulto , Pessoa de Meia-Idade , Adulto Jovem , Polimorfismo de Fragmento de Restrição/genética , Polimorfismo de Nucleotídeo Único/genética , Metilenotetra-Hidrofolato Redutase (NADPH2)/genética , Hipertireoidismo/genética , Hipotireoidismo/genética , Haplótipos , Estudos de Casos e Controles , Reação em Cadeia da Polimerase , Fatores de Risco , Metilação de DNA , Predisposição Genética para Doença , Alelos , Genótipo , JordâniaRESUMO
ABSTRACT Objective: To investigate potential associations of four substitutions in NAT2 gene and of acetylator phenotype of NAT2 with systemic lupus erythematosus (SLE) and clinical phenotypes. Methods: Molecular analysis of 481C>T, 590G>A, 857G>A, and 191G>A substitutions in the NAT2 gene was performed by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) technique, from DNA extracted from peripheral blood samples obtained from patients with SLE (n = 91) and controls (n = 97). Results and conclusions: The 857GA genotype was more prevalent among nonwhite SLE patients (OR = 4.01, 95% CI = 1.18-13.59). The 481T allele showed a positive association with hematological disorders that involve autoimmune mechanisms, specifically autoimmune hemolytic anemia or autoimmune thrombocytopenia (OR = 1.97; 95% CI = 1.01-3.81).
RESUMO Objetivo: Investigar potenciais associações de quatro substituições do gene NAT2 (N-acetiltransferase 2) e do fenótipo acetilador de NAT2 com o lúpus eritematoso sistêmico (LES) e os fenótipos clínicos. Métodos: A análise molecular das substituições 481C > T, 590G > A, 857G > A e 191G > A do gene NAT2 foi feita com a técnica de PCR-RFLP, usando DNA extraído de amostras de sangue periférico obtidas de pacientes com LES (n = 91) e controles (n = 97). Resultados e conclusões: O genótipo 857GA foi mais prevalente entre pacientes com LES não brancas (OR = 4,01, IC 95% = 1,18-13,59). O alelo 481 T apresentou associação positiva com as alterações hematológicas que envolvem mecanismos autoimunes, especificamente anemia hemolítica autoimune ou trombocitopenia autoimune (OR = 1,97; IC 95% = 1,01-3,81).
Assuntos
Humanos , Arilamina N-Acetiltransferase/genética , Polimorfismo de Fragmento de Restrição/genética , Lúpus Eritematoso Sistêmico/genética , Predisposição Genética para Doença/genética , GenótipoRESUMO
Abstract: Background: Psoriasis is a multigenic and multifactorial dermatological disease linked to cardiovascular diseases. Increased levels of homocysteine in patients with psoriasis have been demonstrated in many studies. The most frequently investigated genetic defect that plays a role in homocysteine metabolism is single point substitution (C to T) located on the 677th nucleotide of the methylenetetrahydrofolate reductase gene (MTHFR). Objective: In this study, we aimed to investigate methylenetetrahydrofolate C677T polymorphism in psoriasis patients in Turkey. Methods: The study included 96 patients with psoriasis and 77 controls from southern Turkey. Methylenetetrahydrofolate C677T polymorphism was analysed using the Polymerase Chain Reaction-Restriction Fragment Length Polymorphism methods. Results: In the psoriasis group, 34 CC (35.4%), 46 CT (47.9%) and 16 TT (16.7%) genotypes were found, respectively; while in the control group, the figures were 39 (50.6%), 35 (45.5%), 3 (3.9%). Homozygote and heterozygote T alleles of methylenetetrahydrofolate C677T polymorphism were significantly higher in the psoriasis than in the control group (p=0.013). Conclusion: We firstly found a correlation between methylenetetrahydrofolate C677T polymorphism and psoriasis among the southern Turkish population.
Assuntos
Humanos , Masculino , Feminino , Adulto , Pessoa de Meia-Idade , Polimorfismo Genético/genética , Psoríase/enzimologia , Metilenotetra-Hidrofolato Redutase (NADPH2)/genética , Psoríase/genética , Turquia , Polimorfismo de Fragmento de Restrição/genética , Fatores de RiscoRESUMO
ABSTRACT Objectives: Innate immunity is involved in the physiopathology of ankylosing spondylitis (AS), with the participation of Gram-negative bacteria, modulation of human leukocyte antigen (HLA) B27 and the involvement of pattern recognition receptors, such as Toll-like receptors (TLRs). The aim of this study was to investigate the clinical characteristics and frequency of TLR4 polymorphisms (Asp299Gly and Thr 399Ile) in a cohort of Brazilian patients with AS. Methods: A cross-sectional study was carried out involving 200 patients with a diagnosis of AS and a healthy control group of 200 individuals. Disease activity, severity and functional capacity were measured. The study of TLR4 polymorphisms was performed using the restriction fragment length polymorphism method. HLA-B27 was analyzed by conventional polymerase chain reaction. The IBM SPSS Statistics 20 program was used for the statistical analysis, with p-values less than 0.05 considered significant. Results: Mean age and disease duration were 43.1 ± 12.7 and 16.6 ± 9.2 years, respectively. The sample was predominantly male (71%) and non-Caucasian (52%). A total of 66% of the group of patients were positive for HLA-B27. The sample of patients was characterized by moderate functional impairment and a high degree of disease activity. No significant association was found between the two TLR4 polymorphisms and susceptibility to AS. Conclusions: TLR4 polymorphisms 399 and 299 were not more frequent in patients with AS in comparison to the health controls and none of the clinical variables were associated with these polymorphisms.
RESUMO Objetivos: A imunidade inata está envolvida na fisiopatologia da espondilite anquilosante (EA), com a participação de bactérias gram-negativas, modulação do antígeno leucocitário humano (HLA) B27 e o envolvimento de receptores de reconhecimento de padrões, como os receptores Toll-like (TLR). O objetivo deste estudo foi investigar as características clínicas e a frequência de polimorfismos em TLR4 (Asp299Gly e Thr399Ile) em uma coorte de pacientes brasileiros com EA. Métodos: Fez-se um estudo transversal que envolveu 200 pacientes com diagnóstico de EA e um grupo controle saudável de 200 indivíduos. Mediram-se a atividade da doença, a gravidade e a capacidade funcional. O estudo dos polimorfismos em TLR4 foi feito com o método de polimorfismo de fragmentos de restrição. O HLA-B27 foi analisado por reação em cadeia da polimerase convencional. Usou-se o programa SPSS Statistics 20 da IBM para a análise estatística e foram considerados significativos valores de p inferiores a 0,05. Resultados: A média de idade e a duração da doença foram de 43,1 ± 12,7 e 16,6 ± 9,2 anos, respectivamente. A amostra foi predominantemente do sexo masculino (71%) e de não brancos (52%). Do grupo de pacientes 66% eram HLA-B27 positivos. A amostra de pacientes foi caracterizada por uma alteração funcional moderada e um elevado grau de atividade da doença. Não foi encontrada associação estatisticamente significativa entre os polimorfismos em TLR4 e a susceptibilidade à EA. Conclusões: Os polimorfismos em TLR4 399 e 299 não foram mais frequentes em pacientes com EA em comparação com controles saudáveis e nenhuma das variáveis clínicas esteve associada a esses polimorfismos.
Assuntos
Humanos , Masculino , Feminino , Adulto , Espondilite Anquilosante/genética , Polimorfismo de Fragmento de Restrição/genética , Antígeno HLA-B27/genética , Receptor 4 Toll-Like/genética , Brasil , Estudos de Casos e Controles , Estudos Transversais , Predisposição Genética para DoençaRESUMO
Introduction The pathogenesis of Parkinson’s disease (PD) involves both genetic susceptibility and environmental factors, with focus on the mutation in the alpha-synuclein gene (SNCA).Objective To analyse the polymorphism SNCA-A53T in patients with familial PD (FPD) and sporadic PD (SPD).Method A total of 294 individuals were studied, regardless of sex and with mixed ethnicity. The study group with 154 patients with PD, and the control group included 140 individuals without PD. The genotyping of SNCA-A53T was performed by PCR/RFLP. Significance level was p < 0.05.Results Among all patients, 37 (24%) had FPD and 117 (75.9%) had SPD. The absence of SNCA-A53T mutation was observed in all individuals.Conclusion SPD is notably observed in patients. However, the SNCA-A53T mutation was absent in all individuals, which does not differ controls from patients. This fact should be confirmed in a Brazilian study case with a more numerous and older population.
Introdução A patogênese da doença de Parkinson (DP) envolve fatores ambientais e suscetibilidade genética, destacando-se a mutação de alfa-sinucleína (SNCA.)Objetivos Analisar a variante genética SNCA-A53T em pacientes com DP familiar (DPF) e DP esporádica (DPE).Método Foram estudados 294 indivíduos, independente de sexo, com etnia miscigenada, sendo 154 com DP e 140 sem a doença (grupo controle). A genotipagem de SNCA-A53T foi realizada por PCR/RFLP. Nível de significância para p < 0,05.Resultados Entre os pacientes, 37(24%) tinham DPF e 117 (75,9%) DPE. A ausência da mutação SNCA-A53T em todos os indivíduos.Conclusão DPE é destacada entre os pacientes, no entanto a mutação SNCA-A53T ausente em todos os indivíduos, não diferenciando os grupo controle e pacientes, o que deve ser confirmado em população brasileira, considerando uma ampla casuística, além da ancestralidade.
Assuntos
Adulto , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Mutação , Doença de Parkinson/genética , Polimorfismo de Fragmento de Restrição/genética , alfa-Sinucleína/genética , Brasil , Estudos de Casos e Controles , Estudos de Associação Genética , Predisposição Genética para Doença , Genótipo , Reação em Cadeia da Polimerase , Fatores SexuaisRESUMO
Background: NAT genes are considered candidate genes for the genetic predisposition to non-syndromic Cleft lip with or without cleft palate (NSCLP), since they codify for N-acetyltransferases, enzymes responsible for the biotransformation of arylamines, hydrazine drugs, and a great number of toxins and carcinogens present in diet, cigarette smoke, and environment. Aim: To determine the association between alleles determining slow acetylator phenotype and the risk of NSCLP. Material and Methods: We analyzed *5 (481C>T), *6 (590G>A) and *7 (857G>A) alleles which determine the slow acetylator phenotype and *4 (wild type) allele by polymerase chain reaction/restriction fragment length polymorphism in 97 progenitor-case trios of NSCLP in Argentinian Obstetric Wards. We evaluated the transmission disequilibrium (TDT). Results: TDT showed a positive association between allele *5 and NSCLP (odds ratio = 1,6; p = 0,03). Conclusions: The presence of *5 allele is significantly higher in cases with congenital NSCLP.
Assuntos
Feminino , Humanos , Masculino , Arilamina N-Acetiltransferase/genética , Fenda Labial/genética , Fissura Palatina/genética , Polimorfismo de Fragmento de Restrição/genética , Alelos , Análise do Polimorfismo de Comprimento de Fragmentos Amplificados , Análise de Variância , Argentina , Pai , Predisposição Genética para Doença , Genótipo , Triagem de Portadores Genéticos , Desequilíbrio de Ligação , MãesRESUMO
Objetivo Analisar a influência da associação do polimorfismo G54D (rs1800450) do gene MBL2 no diabetes melito gestacional (DMG) quanto à necessidade de tratamento complementar e ocorrência de recém-nascidos grandes para a idade gestacional. Sujeitos e métodos Cento e cinco pacientes com DMG segundo parâmetro da OMS (Organização Mundial da Saúde) foram avaliadas no período de novembro de 2010 a outubro de 2012. As gestantes foram divididas em dois grupos correspondentes à presença (n = 37) ou à ausência (n = 68) do alelo mutante. As variantes do polimorfismo G54D foram identificadas por meio da técnica de polimorfismos de comprimentos de fragmentos de restrição (RFLP). Parâmetros antropométricos e bioquímicos da mãe e do recém-nascido (RN) e a necessidade de terapia complementar associada à dietoterapia foram avaliados como desfechos primários. Resultados Das pacientes analisadas, 35,2% carregavam pelo menos um alelo mutante do polimorfismo G54D. Os dois grupos não apresentaram diferença significativa quanto a ganho de peso, paridade, idade, índice de massa corporal e idade gestacional de chegada à maternidade. Os grupos de pacientes portadoras ou não do alelo mutante não diferiram quanto à necessidade de tratamento complementar à dietoterapia (16,2% vs. 26,7%) respectivamente e à ocorrência de recém-nascidos grandes para a idade gestacional (24,3% vs. 13,2%). Conclusão Nossos dados demonstraram que o polimorfismo G54D do gene MBL2 não teve efeito sobre a ...
Objective To assess the association of the G54D (rs1800450) polymorphism of the gene MBL2 in the gestational diabetes mellitus with the need for additional treatment and the occurrence of large newborns for the gestational age. Subjects and methods One hundred and five patients recruited in Joinville – Brazil were evaluated between November 2010 and October 2012. Pregnant women were divided in two groups correspondents to the presence (n = 37) or absence (n = 68) of the mutant allele. The variants of the polymorphism G54D were identified by restriction fragment lengths polymorphisms (RFLP). Anthropometric and biochemical parameters of the mother and the newborn, and the necessity of additional therapy associated with diet were assessed as the primary outcomes. Results Thirty-five point two percent of the evaluated patients carried at least one mutated allele of G54D polymorphism. There were no significant differences in weight gain, parity, age, body mass index and gestational age of arrival at maternity between the two groups. The groups of patients with or without the mutated allele did not differ in the need for additional treatment associated with diet (16.2% vs. 26.7%) respectively and with the occurrence of large newborns for gestational age (24.3% vs. 13.2%). Conclusion Our data showed that the polymorphism G54D of the gene MBL2 had no effect in the need for additional treatment associated with the diet-based therapy and in the occurrence of large newborns for gestational age in the studied population. Arq Bras Endocrinol Metab. 2014;58(9):900-5 .
Assuntos
Adulto , Feminino , Humanos , Recém-Nascido , Gravidez , Adulto Jovem , Peso ao Nascer/genética , Diabetes Gestacional/genética , Macrossomia Fetal/genética , Lectina de Ligação a Manose/genética , Polimorfismo Genético/genética , Polimorfismo de Fragmento de Restrição/genética , Alelos , Índice de Massa Corporal , Glicemia/análise , Idade Gestacional , Frequência do Gene/genética , Estudos Prospectivos , Aumento de PesoRESUMO
Associations between polymorphisms of the CD36 gene and susceptibility to coronary artery heart disease (CHD) are not clear. We assessed allele frequencies and genotype distributions of CD36 gene polymorphisms in 112 CHD patients and 129 control patients using semi-quantitative polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) analysis. Additionally, we detected CD36 mRNA expression by real-time quantitative PCR, and we quantified plasma levels of oxidized low-density lipoprotein (ox-LDL) using an enzyme-linked immunosorbent assay (ELISA). There were no significant differences between the two groups (P>0.05) in allele frequencies of rs1761667 or in genotype distribution and allele frequencies of rs3173798. The genotype distribution of rs1761667 significantly differed between CHD patients and controls (P=0.034), with a significantly higher frequency of the AG genotype in the CHD group compared to the control group (P=0.011). The plasma levels of ox-LDL in patients with the AG genotype were remarkably higher than those with the GG and AA genotypes (P=0.010). In a randomized sample taken from patients in the two groups, the CD36 mRNA expression of the CHD patients was higher than that of the controls. In CHD patients, the CD36 mRNA expression in AG genotype patients was remarkably higher than in those with an AA genotype (P=0.005). After adjusted logistic regression analysis, the AG genotype of rs1761667 was associated with an increased risk of CHD (OR=2.337, 95% CI=1.336-4.087, P=0.003). In conclusion, the rs1761667 polymorphism may be closely associated with developing CHD in the Chongqing Han population of China, and an AG genotype may be a genetic susceptibility factor for CHD.
Assuntos
Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , /genética , Doença da Artéria Coronariana/genética , Predisposição Genética para Doença , Polimorfismo Genético/genética , Povo Asiático/genética , Estudos de Casos e Controles , China/etnologia , Doença da Artéria Coronariana/sangue , Doença da Artéria Coronariana/etnologia , Ensaio de Imunoadsorção Enzimática , Frequência do Gene , Genótipo , Predisposição Genética para Doença/etnologia , Modelos Logísticos , Lipoproteínas LDL/sangue , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição/genética , Reação em Cadeia da Polimerase em Tempo Real , Fatores de Risco , RNA Mensageiro/análiseRESUMO
The characteristics of tuberculosis (TB) patients related to a chain of recent TB transmissions were investigated. Mycobacterium tuberculosis (MTB) isolates (120) were genotyped using the restriction fragment length polymorphism-IS6110 (R), spacer oligotyping (S) and mycobacterial interspersed repetitive units-variable number of tandem repeats (M) methods. The MTB isolates were clustered and the clusters were grouped according to the similarities of their genotypes. Spearman’s rank correlation coefficients between the groups of MTB isolates with similar genotypes and those patient characteristics indicating a risk for a pulmonary TB (PTB) chain transmission were ana- lysed. The isolates showing similar genotypes were distributed as follows: SMR (5%), SM (12.5%), SR (1.67%), MR (0%), S (46.67%), M (5%) and R (0%). The remaining 35 cases were orphans. SMR exhibited a significant correlation (p < 0.05) with visits to clinics, municipalities and comorbidities (primarily diabetes mellitus). S correlated with drug consumption and M with comorbidities. SMR is needed to identify a social network in metropolitan areas for PTB transmission and S and M are able to detect risk factors as secondary components of a transmission chain of TB.
Assuntos
Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Adulto Jovem , Técnicas de Genotipagem/métodos , Mycobacterium tuberculosis/genética , Tuberculose Pulmonar/microbiologia , Tuberculose Pulmonar/transmissão , Cidades , Comorbidade , DNA Bacteriano/isolamento & purificação , Genótipo , Sequências Repetitivas Dispersas/genética , Testes de Sensibilidade Microbiana , México/epidemiologia , Epidemiologia Molecular/métodos , Mycobacterium tuberculosis/classificação , Mycobacterium tuberculosis/isolamento & purificação , Polimorfismo de Fragmento de Restrição/genética , Fatores de Risco , Fatores Sociológicos , Estatísticas não Paramétricas , Sequências de Repetição em Tandem/genética , Tuberculose Pulmonar/epidemiologia , Tuberculose Pulmonar/genética , População UrbanaRESUMO
OBJECTIVE: to determine whether C. trachomatis was present in neonates with infection, but without an isolated pathogen, who died during the first week of life. METHODS: early neonatal death cases whose causes of death had been previously adjudicated by the institutional mortality committee were randomly selected. End-point and real-time polymerase chain reaction of the C. trachomatis omp1 gene was used to blindly identify the presence of chlamydial DNA in the paraffinized samples of five organs (from authorized autopsies) of each of the dead neonates. Additionally, differential diagnoses were conducted by amplifying a fragment of the 16S rRNA of Mycoplasma spp. RESULTS: in five cases (35.7%), C. trachomatis DNA was found in one or more organs. Severe neonatal infection was present in three cases; one of them corresponded to genotype D of C. trachomatis. Interestingly, another case fulfilled the same criteria but had a positive polymerase chain reaction for Mycoplasma hominis, a pathogen known to produce sepsis in newborns. CONCLUSION: the use of molecular biology techniques in these cases of early infant mortality demonstrated that C. trachomatis could play a role in the development of severe infection and in early neonatal death, similarly to that observed with Mycoplasma hominis. Further study is required to determine the pathogenesis of this perinatal infection. .
OBJETIVO: determinar se a C. trachomatis está presente em neonatos com infecção, porém sem patógeno isolado, que morreram durante a primeira semana de vida. MÉTODOS: casos de óbito neonatal precoce cujas causas de óbito haviam sido anteriormente determinadas pelo Comitê de Mortalidade da instituição foram aleatoriamente selecionados. Foram utilizadas as reações em cadeia da polimerase convencional e em tempo real do gene omp1 da C. trachomatis, para identificar, às cegas, a presença de DNA de clamídia nas amostras desparafinizadas de cinco órgãos (de autópsias autorizadas) de cada um dos neonatos mortos. Além disso, foram realizados diagnósticos diferenciais por amplificação de um fragmento do rRNA 16S de Mycoplasma ssp. RESULTADOS: em cinco casos (35,7%) a presença de DNA de C. trachomatis foi detectada em um ou mais órgãos. Havia infecção neonatal grave em três casos; um deles correspondente ao genótipo D de C. trachomatis. Curiosamente, outro caso preencheu os mesmos critérios, porém possuía uma reação em cadeia da polimerase positiva para Mycoplasma hominis, um patógeno conhecido por causar sepse em recém-nascidos. CONCLUSÃO: a utilização de técnicas de biologia molecular nos casos de mortalidade infantil precoce mostrou que a C. trachomatis poderia desempenhar um papel no desenvolvimento de infecção grave e no óbito neonatal precoce semelhante ao observado com a Mycoplasma hominis. São necessários estudos adicionais para determinar a patogênese dessa infecção perinatal. .
Assuntos
Feminino , Humanos , Recém-Nascido , Masculino , Infecções por Chlamydia/microbiologia , Infecções por Chlamydia/mortalidade , Chlamydia trachomatis/genética , DNA Bacteriano/isolamento & purificação , Autopsia , Mycoplasma/isolamento & purificação , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição/genéticaRESUMO
Fundamentos: O polimorfismo C825T do gene GNB3 está associado à hipertensão arterial sistêmica (HAS) em algumas populações já analisadas, porém alguns estudos se mostram controversos no que se refere a esta relação. Objetivo: Avaliar a relação do polimorfismo C825T do gene GNB3 com a HAS de difícil controle medicamentoso em hipertensos de Campos Gerais, PR - Brasil. Métodos: Em relação ao polimorfismo C825T de GNB3, foram determinados os genótipos de 60 hipertensos, os quais foram estratificados em dois grupos (fácil e difícil controle medicamentoso), por meio da técnica de PCR-RFLP (Polymerase Chain Reaction - Restriction Fragment Lenght Polymorphism). Foram avaliadas as frequências alélicas e genotípicas, utilizando-se o teste do qui-quadrado de Pearson, com correção de Yates e odds ratio (OR). Resultados: Não houve diferenças entre os grupos, quando comparadas as frequências alélicas e genotípicas, indicando que a população está em equilíbrio. A probabilidade de o paciente possuir o polimorfismo e a HAS de difícil controle foi 53,5 % (OR=1,15; IC95 % = 0,41-3,26), analisando-se os genótipos. Já a análise dos alelos, separadamente, mostrou uma associação de 55,4 % (OR=1,24; IC95 % = 0,59-2,57). Conclusão: Nesta população não foi encontrada relação entre o polimorfismo C825T do gene GNB3 e a HAS de difícil controle, indicando que outros fatores estão influenciando a manifestação dessa doença nestes pacientes.
Background: C825T polymorphism of the GNB3 gene is associated with systemic arterial hypertension (SAH) in some studied populations, although certain studies are controversial in terms of this relationship. Objective: To evaluate the relationship between C825T polymorphism of the GNB3 gene and difficult-to-treat SAH among hypertensive patients in Campos Gerais, Paraná State, Brazil. Methods: With regard to C825T polymorphism of the GNB3 gene, the genotypes were defined for sixty hypertensive patients divided in 2 groups (easy and difficult-to-treat with drugs), using the Polymerase Chain Reaction - Restriction Fragment Length Polymorphism (PCR-RFLP) technique. The allele and genotype frequencies were assessed through the Pearson chi-square test, with Yates correction and odds ratio (OR). Results: There were no differences between the groups when comparing the allele and genotype frequencies, indicating that the population is in equilibrium. The probability that a patient has polymorphism with difficult-to-treat SAH reached 53.5% (OR=1.15, 95%CI = 0.41-3.26), analyzing the genotypes. A separate allele analysis showed an association of 55.4% (OR=1.24, 95%CI = 0.59-2.57). Conclusion: No relationship was found in this population between C825T polymorphism of the GNB3 gene and difficult-to-treat SAH, indicating that other factors are influencing the appearance of this disease among these patients.
Assuntos
Humanos , Diuréticos/administração & dosagem , Hipertensão/complicações , Polimorfismo de Fragmento de Restrição/genética , Sinvastatina , Estudos de Casos e Controles , Sistema Renina-AngiotensinaRESUMO
OBJECTIVE: The aim of the study was to investigate whether adiposity and metabolic markers, such as leptin, glucose, and lipids, are influenced by leptin (LEP) and leptin receptor (LEPR) gene polymorphisms in a sample of our population. SUBJECTS AND METHODS: A group of 326 individuals of Caucasian-European descent, aged 30 to 80 years, 87 men and 239 women, 148 obese and 178 non-obese, was randomly selected at two clinical hospitals in the city of Sao Paulo, Brazil. All individuals declared their ethnic group as white during the initial interview. Anthropometric measurements, body mass index (BMI), and fat mass were evaluated. Blood samples were drawn for DNA extraction and measurements of leptin, soluble leptin receptor, glucose, and lipids. LEP -2548G>A and LEPR Lys109Arg (c.326A>G), Gln233Arg (c.668A>G) and Lys656Asn (c.1968G>C) polymorphisms were detected by PCR-RFLP. RESULTS: Increased leptin and serum lipids, and LEPR Arg223Arg (GG genotype) were associated with higher risk for obesity (p < 0.05), while reduced risk was found in LEPR Arg109Arg (GG genotype) carriers (OR: 0.38, 95%CI: 0.19-0.77, p = 0.007). Multiple linear regression analysis showed a relationship between LEPR 223Arg, increased waist circumference, and leptinemia (p < 0.05), while LEPR 109Arg was associated with high total cholesterol and triglycerides (p < 0.05). LEPR haplotype 3 (AGG: 109Lys/233Arg/656Lys) carriers have increased risk for obesity (OR: 2.56, 95% CI: 1.19-5.49, p = 0.017). Moreover, this haplotype was associated with increased BMI, waist circumference, and leptinemia (p < 0.05). CONCLUSIONS: LEPR polymorphisms are associated with obesity, hyperleptinemia, and atherogenic lipid profile, suggesting their potential role for leptin resistance and cardiovascular risk. Moreover, LEPR haplotype 3 confers susceptibility to adiposity and hyperleptinemia in our population.
OBJETIVO: O estudo teve por objetivo investigar a influência de polimorfismos nos genes da leptina (LEP) e do receptor de leptina (LEPR) na adiposidade e em marcadores metabólicos, como leptina, glicose e lipídeos, em uma amostra de nossa população. SUJEITOS E MÉTODOS: Um grupo de 326 indivíduos com idade de 30 a 80 anos, 87 homens e 239 mulheres, 148 obesos e 178 não obesos, e de etnia branca foi selecionado aleatoriamente em dois hospitais clínicos da cidade de São Paulo, Brasil. Medidas antropométricas, índice de massa corporal (IMC) e gordura corporal foram avaliados. Amostras de sangue foram obtidas para extração de DNA e determinações de leptina, receptor de leptina solúvel, glicose e lipídeos. Os polimorfismos LEP -2548G>A e LEPR Lys109Arg (c.326A>G), Gln233Arg (c.668A>G) e Lys656Asn (c.1968G>C) foram detectados por PCR-RFLP. RESULTADOS: Leptina e lipídeos séricos aumentados e LEPR Arg223Arg (genótipo GG) foram associados com maior risco de obesidade (p < 0,05), enquanto foi encontrado risco reduzido de obesidade, em portadores de LEPR Arg109Arg (genótipo GG) (OR: 0,38, 95%CI: 0,19-0,77, p = 0,007). A análise de regressão linear múltipla mostrou relação entre o alelo LEPR 223Arg e circunferência abdominal e leptinemia aumentadas (p < 0,05), enquanto o alelo LEPR 109Arg foi associado com aumento de colesterol total e triglicerídeos (p < 0,05). Os portadores do haplotipo 3 do LEPR (AGG: 109Lys/233Arg/656Lys) tiveram maior risco aumentado para obesidade (OR: 2.56, 95% CI: 1.19-5.49, p = 0,017). Além disso, esse haplótipo foi associado com IMC, circunferência abdominal e leptinemia aumentados (p < 0,05). CONCLUSÕES: Polimorfismos de LEPR são associados com obesidade, hiperleptinemia e perfil lipídico aterogênico sugerindo seu papel potencial para a resistência à leptina e risco cardiovascular.
Assuntos
Adulto , Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Adiposidade/genética , Leptina/genética , Obesidade/genética , Polimorfismo de Fragmento de Restrição/genética , Receptores para Leptina/genética , Análise de Variância , Brasil , Biomarcadores/sangue , Glicemia/metabolismo , Distribuição de Qui-Quadrado , Frequência do Gene , Glucose/metabolismo , Leptina/sangue , Obesidade/sangue , Reação em Cadeia da Polimerase , Fatores de Risco , Receptores para Leptina/sangue , Circunferência da Cintura/genéticaRESUMO
OBJECTIVE: The JAK2 46/1 haplotype has recently been described as a major contributing factor to the development of myeloproliferative neoplasm, whether positive or negative forthe JAK2 V617F mutation. The G allele, identified by a single-nucleotide polymorphism known as JAK2 rs10974944, is part of the JAK2 46/1 haplotype. The aim of this study was to verify the association between the presence of the G allele and the development of BCR-ABL-negative chronic myeloproliferative neoplasms in our population. METHODS: Blood and oral mucosa swab samples were obtained from 56 patients of two local Brazilian hospitals who had previously been diagnosed with BCR-ABL-negative chronic myeloproliferative neoplasms. Blood samples from 90 local blood donors were used as controls. The presence of the G allele was assessed using a PCR-RFLP assay after extracting DNA from the samples. RESULTS: The presence of the G allele was strongly associated with the presence of BCR-ABL-negative chronic myeloproliferative neoplasms (p = 0.0001; OR = 2.674; 95% CI = 1.630-4.385) in the studied population. CONCLUSION: In agreement with previous reports, the JAK2 46/1 haplotype, represented in this study by the presence of the G allele, is an important predisposing factor in the oncogenetic development of these neoplasms in our population.
Assuntos
Idoso , Idoso de 80 Anos ou mais , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Adulto Jovem , Haplótipos/genética , /genética , Transtornos Mieloproliferativos/genética , Brasil , Distribuição de Qui-Quadrado , Doença Crônica , Proteínas de Fusão bcr-abl/genética , Frequência do Gene , Mutação/genética , Reação em Cadeia da Polimerase , Polimorfismo de Fragmento de Restrição/genética , Distribuição por SexoRESUMO
OBJECTIVE: The aim of this study was to evaluate, by PCR-RFLP and real-time PCR, the yield and quality of genomic DNA collected from buccal cells by mouthwash after different storage times at room temperature. MATERIAL AND METHODS: A group of volunteers was recruited to collect buccal cells using a mouthwash solution. The collected solution was divided into 3 tubes, one tube were used for immediate extraction and the remaining received ethanol and were kept at room temperature for 4 and 8 days followed by dna extraction. The concentration, purity and integrity of the dna were determined using spectrophotometry and electrophoresis. DNA quality differences among the three incubation times were also evaluated for genotyping EGF +61 a/g (rs 4444903) polymorphism by PCR-RFLP and for IRF6 polymorphism (rs 17015215) using real-time PCR. RESULTS: There was no significant difference of dna yield (p=0.75) and purity (p=0.86) among the three different incubation times. DNA obtained from different incubation times presented high-molecular weight. The PCR-RFLP and real time pcr reactions were successfully performed for all DNA samples, even those extracted after 8 days of incubation. All samples genotyped by real-time pcr presented c allele for irf6 gene polymorphism (homozygous: cc; heterozygous: Ct) and the C allele was used as a reference for Ct values. The samples presented the same genotype for the different times in both techniques. CONCLUSION: We demonstrated that the method described herein is simple and low cost, and that DNA can be extracted and pcr amplified after storage in mouthwash solution at room temperature.
Assuntos
Adulto , Feminino , Humanos , Pessoa de Meia-Idade , DNA , Técnicas de Genotipagem/métodos , Boca/citologia , Polimorfismo de Fragmento de Restrição/genética , Reação em Cadeia da Polimerase em Tempo Real/métodos , Análise de Variância , Eletroforese , Reprodutibilidade dos Testes , Saliva , Espectrofotometria , Fatores de TempoRESUMO
Obstructive sleep apnea syndrome (OSAS) is one of the most complex disorders of sleep; it involves several genetic factors that contribute to the phenotype. Serotonin (5-HT) regulates a variety of visceral and physiological functions, including sleep. Gene 5-HTR2A polymorphisms may change the transcription of several receptors in the serotoninergic system, thereby contributing to OSAS. AIM: To investigate the prevalence of T102C and -1438G/A polymorphisms in the 5-HTR2A gene of patients with and without OSAS . MATERIAL AND METHOD: A molecular study of 100 index-cases and 100 controls of both genders. DNA was extracted from blood leukocytes samples and the regions that enclose both polymorphisms were amplified with PCR-RFLP. STUDY DESIGN: A cross-sectional case study. RESULTS: There was a significant prevalence of males in index cases compared to controls (p<0.0001). No significant genotypic differences between cases and controls were found in T102C polymorphisms (p=1.000).There were significant differences between the AA genotype of -1438G/A polymorphisms and patients with OSAS (OR:2.3; CI95 percent:1.20-4.38, p=0.01). CONCLUSION: Serotonergic mechanisms may be related to OSAS. There were no differences in the prevalence of T102C polymorphisms in patients with OSAS and the control group. There is evidence of an association between the -1438G /A polymorphism and OSAS.
A síndrome da apneia obstrutiva do sono (SAOS) é um dos distúrbios mais complexos do sono, envolvendo múltiplos fatores genéticos contribuintes para o fenótipo. A serotonina (5-HT) está envolvida na regulação de uma variedade de funções viscerais e fisiológicas, inclusive o sono. Polimorfismos no gene 5-HTR2A podem alterar a transcrição, afetando o número de receptores do sistema serotoninérgico, contribuindo para a SAOS. OBJETIVO: Investigar a prevalência dos polimorfismos T102C e -1438G/A no gene HTR2A em pacientes com e sem SAOS. MATERIAL E MÉTODO: Estudo molecular em 100 pacientes como casos-índice e em 100 como grupo controle, de ambos os gêneros. O DNA foi extraído de leucócitos de sangue periférico e realizada a amplificação das regiões que abrangem ambos os polimorfismos pelas técnicas da PCR-RFLP. DESENHO DO ESTUDO: Estudo de caso/controle em corte transversal. Resultados: Houve prevalência significativa do gênero masculino nos casos-índice em relação aos controles (p<0,0001). Para o polimorfismo T102C, não houve diferença genotípica significante entre casos e controles (p=1,000). Houve diferença significativa entre o genótipo AA do polimorfismo -1438G/A e pacientes com SAOS (OR:2,3; IC95 por cento:1,20-4,38; p=0,01). CONCLUSãO: Os mecanismos serotoninérgicos parecem estar relacionados a SAOS. Não há diferenças na prevalência do polimorfismo T102C entre os pacientes com SAOS e o grupo controle. Há evidências de associação entre o polimorfismo -1438G/A e a SAOS.
Assuntos
Adolescente , Adulto , Idoso , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Adulto Jovem , Polimorfismo de Fragmento de Restrição/genética , /genética , Apneia Obstrutiva do Sono/genética , Estudos de Casos e Controles , Estudos Transversais , Genótipo , Reação em Cadeia da Polimerase , Índice de Gravidade de DoençaRESUMO
Background & objectives Endothelial nitric oxide is a potent vasodilator and impairment of its generation brought about by gene polymorphism is considered a major predictor for several diseases. A single nucleotide polymorphism G894T within exon 7 of endothelial nitric oxide synthase (eNOS-7) gene, resulting in a replacement of glutamic acid by aspartic acid, has been studied as a putative candidate gene for cardiovascular diseases. The pattern of eNOS-7 Glu298Asp variant in the Indian population is poorly known. The present study was planned to determine the prevalence of the variant of this gene among tea garden community in Assam, North-East India with high prevalence of hypertension. Methods Study participants of both sex aged ≥18 yr were recruited randomly from temporary field clinics established in tea gardens of Dibrugarh, Assam. Genomic DNA was extracted from 409 subjects by the conventional phenol-chloroform method. The prevalence of the eNOS exon 7 Glu298Asp variant was determined by polymerase chain reaction and restriction fragment length polymorphism analysis. Results The study population was in Hardy-Weinberg Equilibrium. The frequency of the eNOS GG, GT and TT genotypes was found to be 75, 22 and 3 per cent respectively and did not show any significant difference in gender wise analysis. Interpretation & conclusions Our results showed that the prevalence of the homozygous GG genotype was high (75%) and the rare mutant genotype (homozygous, TT) was 3 per cent in a population at risk with cardiovascular disease. Such population-based data on various polymorphisms can ultimately be exploited in pharmacogenomics.
Assuntos
Adulto , Idoso , Idoso de 80 Anos ou mais , Doenças Cardiovasculares/enzimologia , Doenças Cardiovasculares/epidemiologia , Doenças Cardiovasculares/genética , Cromossomos Humanos Par 7/genética , Feminino , Predisposição Genética para Doença/epidemiologia , Predisposição Genética para Doença/genética , Genótipo , Haplótipos/genética , Humanos , Índia/epidemiologia , Masculino , Pessoa de Meia-Idade , Mutação de Sentido Incorreto/genética , Óxido Nítrico Sintase Tipo III/genética , Farmacogenética/métodos , Polimorfismo de Fragmento de Restrição/genética , Polimorfismo de Nucleotídeo Único/genética , PrevalênciaRESUMO
Background & objectives: El Tor Vibrio cholerae O1 carrying ctxBC trait, so-called El Tor variant that causes more severe symptoms than the prototype El Tor strain, first detected in Bangladesh was later shown to have emerged in India in 1992. Subsequently, similar V. cholerae strains were isolated in other countries in Asia and Africa. Thus, it was of interest to investigate the characteristics of V. cholerae O1 strains isolated chronologically (from 1986 to 2009) in Thailand. Methods: A total of 330 V. cholerae O1 Thailand strains from hospitalized patients with cholera isolated during 1986 to 2009 were subjected to conventional biotyping i.e., susceptibility to polymyxin B, chicken erythrocyte agglutination (CCA) and Voges-Proskauer (VP) test. The presence of ctxA, ctxB, zot, ace, toxR, tcpAC, tcpAE, hlyAC and hlyAE were examined by PCR. Mismatch amplification mutation assay (MAMA) - and conventional- PCRs were used for differentiating ctxB and rstR alleles. Results: All 330 strains carried the El Tor virulence gene signature. Among these, 266 strains were typical El Tor (resistant to 50 units of polymyxin B and positive for CCA and VP test) while 64 had mixed classical and El Tor phenotypes (hybrid biotype). Combined MAMA-PCR and the conventional biotyping methods revealed that 36 strains of 1986-1992 were either typical El Tor, hybrid, El Tor variant or unclassified biotype. The hybrid strains were present during 1986-2004. El Tor variant strains were found in 1992, the same year when the typical El Tor strains disappeared. All 294 strains of 1993-2009 carried ctxBC ; 237 were El Tor variant and 57 were hybrid. Interpretation & conclusions: In Thailand, hybrid V. cholerae O1 (mixed biotypes), was found since 1986. Circulating strains, however, are predominantly El Tor variant (El Tor biotype with ctxBC).
Assuntos
Formas Bacterianas Atípicas/genética , Técnicas de Tipagem Bacteriana/métodos , Quimera/genética , Cólera/epidemiologia , Cólera/genética , Cólera/microbiologia , Toxina da Cólera/genética , DNA Bacteriano/genética , Variação Genética , Genótipo , Humanos , Epidemiologia Molecular/métodos , Fenótipo , Polimorfismo de Fragmento de Restrição/genética , Tailândia/epidemiologia , Vibrio cholerae O1/classificação , Vibrio cholerae O1/genética , Vibrio cholerae O1/isolamento & purificaçãoRESUMO
Sichuan basin, situated in the west of China, is the fourth biggest basin in China. In order to describe a systematic study of the cry2-type genes resources from Bacillus thuringiensis strains of Sichuan basin, a total of 791 Bacillus thuringiensis strains have been screened from 2650 soil samples in different ecological regions. The method of PCR-restriction fragment length polymorphism (PCR-RFLP) was used to identify the type of cry2 genes. The results showed that 322 Bacillus thuringiensis strains harbored cry2-type genes and four different RFLP patterns were found. The combination of cry2Aa/cry2Ab genes was the most frequent (90.4 percent), followed by cry2Aa (6.8 percent) and cry2Ab alone (2.5 percent), and only one novel type of cry2 gene was cloned from one isolate (JF19-2). The full-length of this novel gene was obtained by the method of thermal asymmetric interlaced PCR (Tail-PCR), which was designated as cry2Ag1 (GenBank No. ACH91610) by the Bt Pesticide Crystal Protein Nomenclature Committee. In addition, the result of scanning electron microscopic (SEM) observation showed that these strains had erose, spherical, bipyramidal, and square crystal. And the results of sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) demonstrated that these strains harbored about one to three major proteins. These strains exhibited a wide range of insecticidal spectrum toxic to Aedes aegypti (Diptera) and Pieris rapae Linnaeus, 1758 (Lepidoptera). Particularly, JF19-2 contained cry2Ag gene had the highest insecticidal activity. All these researches mentioned above revealed the diversity and particularity of cry2-type gene resources from Bacillus thuringiensis strains in Sichuan basin.