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1.
J. bras. pneumol ; 45(2): e20180278, 2019. tab
Article in English | LILACS | ID: biblio-1002433

ABSTRACT

ABSTRACT Objective: Pulmonary nontuberculous mycobacterial infections are caused by nontuberculous mycobacteria (NTM), the microbiological diagnosis of which involves the isolation and identification of the same species in at least two sputum samples, one BAL fluid sample, or one lung biopsy sample. The objective of the present study was to determine the frequency at which the various NTM species are identified among selected individuals and in potential cases of pulmonary nontuberculous mycobacterial infection. Methods: This was a retrospective analysis of the data on species isolated from respiratory specimens collected from 2,843 individuals between 2011 and 2014. Potential NTM infection cases were identified on the basis of the international microbiological criteria adopted in the state of São Paulo. Results: A total of 50 species were identified using the molecular method PCR-restriction enzyme analysis. Samples collected from 1,014 individuals were analyzed in relation to the microbiological criteria, and 448 (44.18%) had a presumptive diagnosis of pulmonary nontuberculous mycobacterial infection, the species identified most frequently being, in descending order, Mycobacterium kansasii, M. abscessus, M. intracellulare, M. avium, and M. szulgai. Conclusions: Although various NTM species were identified among the individuals studied, those presumptively identified most frequently on the basis of the microbiological criteria adopted in the state of São Paulo were the ones that are most commonly associated with pulmonary nontuberculous mycobacterial infection worldwide or in specific geographic regions.


RESUMO Objetivo: As micobacterioses pulmonares são doenças causadas por micobactérias não tuberculosas (MNTs), cujo diagnóstico microbiológico envolve o isolamento e a identificação da mesma espécie a partir de pelo menos duas amostras de escarro, uma de lavado brônquico ou uma de biópsia pulmonar. O objetivo do presente estudo foi determinar as frequências das diferentes espécies de MNTs em indivíduos selecionados e em potenciais casos de micobacterioses pulmonares. Métodos: Análise retrospectiva dos dados de identificação de espécies isoladas a partir de espécimes clínicos pulmonares de 2.843 indivíduos incluídos no estudo entre 2011 e 2014. A identificação dos potenciais casos baseou-se nos critérios microbiológicos internacionais adotados no estado de São Paulo. Resultados: Um total de 50 espécies foi identificado utilizando-se o método molecular PCR-restriction enzyme analysis. Dos 1.014 indivíduos analisados quanto aos critérios microbiológicos, 448 (44,18%) tiveram o diagnóstico presuntivo de micobacteriose pulmonar, sendo as maiores frequências de casos, em ordem decrescente, Mycobacterium kansasii, M. abscessus, M. intracellulare, M. avium e M. szulgai. Conclusões: Embora tenham sido identificadas diversas espécies de MNTs entre os indivíduos estudados, as que tiveram as maiores frequências de casos presuntivamente identificados pelos critérios microbiológicos adotados no estado de São Paulo foram as que mais frequentemente estão associadas a micobacterioses pulmonares mundialmente ou em várias regiões geográficas.


Subject(s)
Humans , Male , Female , Nontuberculous Mycobacteria/isolation & purification , Mycobacterium Infections, Nontuberculous/diagnosis , Mycobacterium Infections, Nontuberculous/microbiology , Brazil/epidemiology , Restriction Mapping , Polymerase Chain Reaction , Retrospective Studies , Lung/microbiology , Mycobacterium Infections, Nontuberculous/epidemiology
2.
Acta sci., Biol. sci ; 38(3): l3327-332, jul.-set. 2016. tab, map
Article in English | LILACS, VETINDEX | ID: biblio-1460781

ABSTRACT

Tibouchina hatschbachii Wurdack (Melastomataceae) is an autogamous shrub restricted to granite (GO) and sandstone (SO) rock outcrops from subtropical Brazil. We designed primers for the amplification of microsatellite regions for T. hatschbachii, and characterized these primers to estimate genetic diversity parameters and contemporary genetic structure patterns. Eight loci were successfully amplified and were characterized using 70 individuals from three natural populations. Polymorphic information content ranged from 0.200 to 0.772 per locus. All loci were polymorphic, with allele numbers ranging from two to eight. The low degree of polymorphism may be explained by the fact that T. hatschbachii has disjunct populations and a recent genetic bottleneck, and also that it is self-pollinated. The observed and expected heterozygosities ranged from 0.115 to 1.000 and from 0.112 to 0.800, respectively. We observed private alleles in all loci. These are important features that enable us to identify population differentiation and help to us understand gene flow patterns for T. hatschbachii in subtropical Brazil. Eight microsatellite loci from other species of Tibouchina amplified positively in T. hatschbachii.


Tibouchina hatschbachii Wurdack (Melastomataceae) é um arbusto autógamo, com ocorrência restrita em afloramentos rochosos graníticos (GO) e areníticos (SO) na região subtropical do Brasil. Neste trabalho, foram desenvolvidos marcadores para a amplificação de regiões microssatélites para T. hatschbachii e caracterizados esses primers para estimar parâmetros de diversidade genética. Oito loci foram amplificados com sucesso e caracterizados, utilizando 70 indivíduos de três populações naturais. O conteúdo de informação polimórfica variou de 0,200 a 0,772 por locus. Todos os loci foram polimórficos, com números de alelos que variam de dois a oito. O baixo grau de polimorfismo pode ser explicado pelo fato de que T. hatschbachii possui populações disjuntas e uma história recente de gargalo genético populacional, e também pelo fato de apresentar um sistema reprodutivo de autopolinização, tendendo a favorecer a baixa variação. As heterozigosidades observadas e esperadas variaram entre 0,115-1,000 e 0,112-0,800, respectivamente. Também foi observada a presença de alelos privados em todos os loci. Estas são características importantes que nos permitirão identificar a diferenciação entre populações e poderão ajudar na compreensão dos padrões de fluxo gênico atual de T. hatschbachii na região subtropical do Brasil. Oito loci microssatélites de outras espécies de Tibouchina amplificaram


Subject(s)
Animals , Melastomataceae/growth & development , Melastomataceae/genetics , Microsatellite Repeats , Restriction Mapping/veterinary
3.
Rev. latinoam. enferm ; 22(6): 973-979, 16/12/2014. tab
Article in English | LILACS, BDENF | ID: lil-732953

ABSTRACT

OBJECTIVE: to relate neck circumference with metabolic syndrome and its criteria among college students. METHOD: cross-sectional study conducted with 702 college students in Fortaleza, CE, Brazil from September 2010 to June 2011. Socio-demographic data, waist circumference and neck circumference were collected together with blood pressure, fasting blood sugar, triglyceride levels, and HDL-C. RESULTS: 1.7% of the studied sample presented metabolic syndrome. Of these, 58.3% presented altered neck circumference (p<0.006). As neck circumference decreases, pressure levels improve (p<0.001). Additionally, college students with high fasting blood sugar (p=0.003) and high triglyceride levels (p<0.001) presented higher values of neck circumference. CONCLUSION: neck circumference is a potential predictive marker in the detection of metabolic syndrome and its components among college students. .


OBJETIVO: relacionar a circunferência do pescoço com a síndrome metabólica e seus critérios em universitários. MÉTODO: estudo transversal, realizado com 702 universitários de Fortaleza, CE, Brasil, no período de setembro de 2010 a junho de 2011. Coletaram-se dados sociodemográficos, circunferência da cintura, circunferência do pescoço, níveis de pressão arterial e glicemia plasmática de jejum, triglicerídeos e lipoproteína de alta densidade. RESULTADOS: 1,7% da amostra investigada tinha a síndrome metabólica. Desses, 58,3% apresentaram circunferência do pescoço alterada (p<0,006). Na medida em que decresce a circunferência do pescoço, os valores pressóricos dos universitários melhoram (p<0,001). Também, observou-se que universitários com valores de glicemia de jejum plasmática (p=0,003) e triglicerídeos (p<0,001) elevados apresentaram maiores valores de circunferência do pescoço. CONCLUSÃO: a circunferência do pescoço mostrou-se um possível marcador preditivo para detecção da síndrome metabólica e seus componentes em universitários. .


OBJETIVO: relacionar la circunferencia del cuello con el síndrome metabólico y sus criterios en universitarios. MÉTODO: estudio transversal realizado con 702 universitarios de Fortaleza-CE, Brasil, en el período de septiembre de 2010 a junio de 2011. Se recolectaron datos sociodemográficos, circunferencia de la cintura, circunferencia del cuello, niveles de presión arterial y glucemia plasmática de ayuno, triglicéridos y HDL-C. RESULTADOS: 1,7% de la muestra investigada tenían el síndrome metabólico. De estos, 58,3% presentaron circunferencia del cuello alterada (p<0,006). A medida que decrece la circunferencia del cuello mejoran los valores de la presión de los universitarios (p<0,001). También, se observó que los universitarios con valores de glucemia de ayuno plasmática (p=0,003) y triglicéridos (p<0,001) elevados presentaron mayores valores de circunferencia del cuello. CONCLUSIÓN: la circunferencia del cuello se mostró un posible indicador de predicción para la detección del síndrome metabólico y sus componentes, en universitarios. .


Subject(s)
Humans , Animals , Cathepsins/physiology , Lysosomes/metabolism , Proteins/metabolism , Amino Acid Sequence , Autophagy , Base Sequence , Cathepsins/antagonists & inhibitors , Cathepsins/genetics , Cell Compartmentation , Cycloheximide/pharmacology , Cystatins/physiology , Gene Expression Regulation , Leucine/analogs & derivatives , Leucine/pharmacology , Lysosomes/enzymology , Molecular Sequence Data , Muscular Diseases/physiopathology , Restriction Mapping
4.
Mem. Inst. Oswaldo Cruz ; 109(8): 1081-1085, 12/2014. graf
Article in English | LILACS | ID: lil-732602

ABSTRACT

We present here three expression plasmids for Trypanosoma cruzi adapted to the Gateway® recombination cloning system. Two of these plasmids were designed to express trypanosomal proteins fused to a double tag for tandem affinity purification (TAPtag). The TAPtag and Gateway® cassette were introduced into an episomal (pTEX) and an integrative (pTREX) plasmid. Both plasmids were assayed by introducing green fluorescent protein (GFP) by recombination and the integrity of the double-tagged protein was determined by western blotting and immunofluorescence microscopy. The third Gateway adapted vector assayed was the inducible pTcINDEX. When tested with GFP, pTcINDEX-GW showed a good response to tetracycline, being less leaky than its precursor (pTcINDEX).


Subject(s)
Gene Expression/genetics , Genetic Vectors/genetics , Plasmids , Restriction Mapping/methods , Trypanosoma cruzi/genetics , Blotting, Western , Expressed Sequence Tags/metabolism , Green Fluorescent Proteins , Life Cycle Stages/genetics , Mutagenesis, Insertional , Tetracycline/pharmacology , Trypanosoma cruzi/drug effects
5.
São Paulo; SMS; nov. 2014. 2 p. map, ilus.
Monography in Portuguese | LILACS, ColecionaSUS, CEINFO-Producao, SMS-SP, SMS-SP | ID: biblio-940651

ABSTRACT

O folder apresenta estudos e ferramentas realizados e desenvolvidas pela GISA - Gerência de Geoprocessamento e Informações Socioambientais (Coordenação de Epidemiologia e Informação) da Secretaria Municipal da Saúde


Subject(s)
Male , Female , Humans , Health Status Indicators , Restriction Mapping , Unified Health System
6.
Clinical Pediatric Hematology-Oncology ; : 55-58, 2013.
Article in Korean | WPRIM | ID: wpr-788482

ABSTRACT

We report a long-term follow-up of unstable hemoglobin (Hb) patient. He was diagnosed as Hb Madrid [beta115(G17)Ala-->Pro] by direct DNA sequencing and restriction enzyme analysis. Hydroxyurea had been given for beta-chain hemoglobinopathies through activation of gamma(gamma)-chain synthesis. Nowadays he still needs transfusion three or four times per year, but he had been free of hemolytic crisis after hydroxyurea. Although he has been treated for hemochromatosis with parenteral and oral iron-chelating agents, liver cirrhosis complicated by esophageal varix was developed and treated with endoscopic ligation. In addition, he is on warfarin maintenance for anticoagulation therapy for extensive portal vein and superior mesenteric vein thrombosis which presented with abdominal pain and diagnosed by CT scan. In management of unstable Hb patients, physician should monitor and control the serum ferritin level with iron-chelating agents, and be aware of possible long-term complication including hemochromatosis, cirrhosis or thromboembolism.


Subject(s)
Humans , Abdominal Pain , Esophageal and Gastric Varices , Ferritins , Fibrosis , Follow-Up Studies , Hemochromatosis , Hemoglobinopathies , Hemoglobins , Hemoglobins, Abnormal , Hydroxyurea , Ligation , Liver Cirrhosis , Mesenteric Veins , Organothiophosphorus Compounds , Portal Vein , Restriction Mapping , Sequence Analysis, DNA , Thromboembolism , Thrombosis , Warfarin
7.
Journal of Medicinal Plants. 2013; 12 (48): 40-53
in Persian | IMEMR | ID: emr-148724

ABSTRACT

Fennel [Foeniculum vulgar] is a medicinal plant species in the Apiaceae family with culinary and medicinal uses. This study was conducted to evaluate the effects of enzymatic digestion of PCR product in improvement of the efficiency of RAPD markers. Nine RAPD primers were used to amplify the genomic DNA of fifteen accessions of Fennel. Following amplification, a part of PCR products was digested with two restriction enzymes [EcoRl and MseI]. Both of digested and undigested PCR products were separated on agarose gel electrophoresis. The accessions were grouped by cluster analysis and polymorphic information content index was calculated for each marker. Also percentage and component of essential oil were indicated by CC/MS analysis. The comparison of banding patterns of digested and undigested PCR products revealed that digestion of RAPD-PCR product using a four base cutter enzyme such as Mse I shows a higher level of polymorphism as compared to standard RAPD. Cluster analysis based on data obtained by modified RAPD classified accessions more suitable as compared to standard RAPD data. There was no correlation between genetic diversity and metabolic yield. Restriction enzymes have enormous potential to improve the efficiency of RAPD markers in evaluation of genetic diversity across genome


Subject(s)
Plants, Medicinal , Polymerase Chain Reaction , Random Amplified Polymorphic DNA Technique , Polymorphism, Genetic , Restriction Mapping , Genetic Variation , Oils, Volatile
8.
Clinics ; 68(2): 179-184, 2013. ilus, tab
Article in English | LILACS | ID: lil-668804

ABSTRACT

OBJECTIVE: To determine the incidence of Mycobacterium tuberculosis complex and non-tuberculous mycobacterial isolates in the routine setting of a large general hospital using an "in-house" multiplex polymerase chain reaction method and to establish a paradigm for the definitive identification of mycobacteria isolated using semi-automated equipment. METHODS: Established tests, including polymerase chain reaction restriction enzyme analysis, PNB, and NAP inhibition tests as the gold standard, showed 100% agreement with an IS6110/hsp65 multiplex polymerase chain reaction when used to identify stock strains (n = 117). RESULTS: In a subsequent study, 8,790 clinical specimens producing 476 isolates were evaluated with multiplex PCR and also showed 100% agreement in identification using PRA-polymerase chain reaction as the gold standard. The application of this technique to routine analysis was demonstrated in this study. A method was established with the initial application of multiplex PCR for all positive liquid cultures and the subsequent identification of non-tuberculous mycobacteria by polymerase chain reaction restriction enzyme analysis. In total, 77% of isolates belonged to the Mycobacterium tuberculosis complex, and 23% were non-tuberculous mycobacteria. CONCLUSIONS: Several non-tuberculous mycobacterial species were identified, primarily M. avium, but other potentially pathogenic species were also frequently observed, including M. fortuitum, M. abscessus, and M. kansasii. The expeditious communication of these data to the clinical staff was fundamental for the diagnosis of clinical cases. Even in settings where tuberculosis is of major importance, the incidence of non-tuberculous mycobacteria infection is substantial.


Subject(s)
Humans , Mycobacterium tuberculosis/isolation & purification , Nontuberculous Mycobacteria/isolation & purification , Brazil , Hospitals, General , Multiplex Polymerase Chain Reaction , Mycobacterium tuberculosis/classification , Nontuberculous Mycobacteria/classification , Restriction Mapping
9.
Clinical Pediatric Hematology-Oncology ; : 55-58, 2013.
Article in Korean | WPRIM | ID: wpr-221895

ABSTRACT

We report a long-term follow-up of unstable hemoglobin (Hb) patient. He was diagnosed as Hb Madrid [beta115(G17)Ala-->Pro] by direct DNA sequencing and restriction enzyme analysis. Hydroxyurea had been given for beta-chain hemoglobinopathies through activation of gamma(gamma)-chain synthesis. Nowadays he still needs transfusion three or four times per year, but he had been free of hemolytic crisis after hydroxyurea. Although he has been treated for hemochromatosis with parenteral and oral iron-chelating agents, liver cirrhosis complicated by esophageal varix was developed and treated with endoscopic ligation. In addition, he is on warfarin maintenance for anticoagulation therapy for extensive portal vein and superior mesenteric vein thrombosis which presented with abdominal pain and diagnosed by CT scan. In management of unstable Hb patients, physician should monitor and control the serum ferritin level with iron-chelating agents, and be aware of possible long-term complication including hemochromatosis, cirrhosis or thromboembolism.


Subject(s)
Humans , Abdominal Pain , Esophageal and Gastric Varices , Ferritins , Fibrosis , Follow-Up Studies , Hemochromatosis , Hemoglobinopathies , Hemoglobins , Hemoglobins, Abnormal , Hydroxyurea , Ligation , Liver Cirrhosis , Mesenteric Veins , Organothiophosphorus Compounds , Portal Vein , Restriction Mapping , Sequence Analysis, DNA , Thromboembolism , Thrombosis , Warfarin
10.
Journal of Lung Cancer ; : 77-83, 2012.
Article in English | WPRIM | ID: wpr-178022

ABSTRACT

PURPOSE: We applied a simplified method using polymerase chain reaction (PCR)-based enzymatic digestion for the detection of epidermal growth factor receptor (EGFR) mutation. MATERIALS AND METHODS: We selected 74 samples of adenocarcinoma of the lung with EGFR exons 19 and 21 that had been previously sequenced. We designed PCR primers and chose a DNA restriction enzyme. Seventy four additional lung cancer samples were tested as a test set. For test sets, the PCR-based method was performed first, followed by validation of the result by DNA sequencing. RESULTS: In the first sample group, we found 15 (20.3%) mutations in exon 19, and 9 (12.2%) mutations in exon 21 using the sequencing method. By using the PCR-based method, we were able to identify all of the mutated samples detected by the sequencing method. The PCR-based method also detected mutations in exon 19 in three additional samples and in exon 21 in one additional sample. In the second sample group, by performing the PCR-based method, we found 10 (13.5%) and 7 (9.5%) mutations in exons 19 and 21, respectively. Additional mutations in exon 19 were identified in 2 samples by the sequencing method. However, the sequencing method failed to identify a mutation in exon 21 in one sample. CONCLUSION: The sensitivity of the PCR-based enzymatic digestion method seems to be comparable to that of the traditional sequencing method for detecting EGFR mutations. Our method can be widely used as a screening test to select patients who may benefit from EGFR targeted therapy.


Subject(s)
Humans , Adenocarcinoma , Carcinoma, Non-Small-Cell Lung , Digestion , DNA , DNA Restriction Enzymes , Epidermal Growth Factor , Exons , Genes, erbB-1 , Lung , Lung Neoplasms , Mass Screening , Polymerase Chain Reaction , ErbB Receptors , Restriction Mapping
11.
Chinese Journal of Virology ; (6): 439-443, 2012.
Article in Chinese | WPRIM | ID: wpr-354711

ABSTRACT

Serratia marcescens jn01 was employed as the host for the isolation of phages from environmental sewage. One strain of phage named SmPjn was purified by picking transparent plaque with 2mm diameter and clear edge on the double-layer agar repeatedly. Electron micrographs indicated that the phage head was icosahedral with head size and tail length of (58 +/- 2.16) x (55 +/- 0.47) nm and (7 +/- 1.25) nm, respectively. On the basis of the morphology, this phage belongs to the family Podoviridae. Host-range determination revealed that the phage was capable of infecting the other two isolates of S. marcescens, P25 and CMCC41002. The optimal multiplicity of infection was 1. A one-step growth curve of SmPjn indicated that the latent period and burst size were estimated at 50 min and 1,125 pfu/cell, respectively . Genomic DNA of SmPjn was above 27kb in size and could be digested by Hind Ill and EcoR I into 11 and 9 visible fragments after electrophoresis, respectively. A novel Podoviridae-phage infecting S. marcescens was firstly reported in China.


Subject(s)
China , DNA, Viral , Genetics , Metabolism , Host Specificity , Podoviridae , Genetics , Restriction Mapping , Serratia marcescens , Physiology
12.
Rev. Soc. Bras. Med. Trop ; 44(5): 551-554, Sept.-Oct. 2011. ilus, tab
Article in English | LILACS | ID: lil-602919

ABSTRACT

INTRODUCTION: Human cytomegalovirus is an opportunistic betaherpesvirus that causes persistent and serious infections in immunodeficient patients. Recurrent infections occur due to the presence of the virus in a latent state in some cell types. It is possible to examine the virus using molecular methods to aid in the immunological diagnosis and to generate a molecular viral profile in immunodeficient patients. The objective of this study was to characterize cytomegalovirus genotypes and to generate the epidemiological and molecular viral profile in immunodeficient patients. METHODS: A total of 105 samples were collected from immunodeficient patients from the City of Belém, including newborns, hemodialysis patients, transplant recipients and HIV+ patients. An IgG and IgM antibody study was completed using ELISA, and enzymatic analysis by restriction fragment length polymorphism (RFLP) was performed to characterize viral genotypes. RESULTS: It was observed that 100 percent of the patients had IgG antibodies, 87 percent of which were IgG+/IgM-, consistent with a prior infection profile, 13 percent were IgG+/IgM+, suggestive of recent infection. The newborn group had the highest frequency (27 percent) of the IgG+/IgM+ profile. By RFLP analysis, only one genotype was observed, gB2, which corresponded to the standard AD169 strain. CONCLUSIONS: The presence of IgM antibodies in new borns indicates that HCMV continues to be an important cause of congenital infection. The low observed genotypic diversity could be attributed to the small sample size because newborns were excluded from the RFLP analysis. This study will be continued including samples from newborns to extend the knowledge of the general and molecular epidemiology of HCMV in immunodeficient patients.


INTRODUÇÃO: O citomegalovírus é um betaherpesvírus oportunista, causador de infecções persistentes e graves em pacientes imunodeficientes. As infecções recorrentes ocorrem devido à presença do vírus em estado de latência, em alguns tipos celulares, o que possibilita a pesquisa viral por métodos moleculares para auxiliar nos diagnósticos imunológicos, assim como traçar o perfil epidemiológico e molecular viral em pacientes imunodeficientes. O objetivo deste estudo foi caracterizar os genótipos de citomegalovírus e traçar o perfil epidemiológico e molecular viral em pacientes imunodeficientes. MÉTODOS: Um total de 105 amostras foi coletado de pacientes imunodeficientes da Cidade de Belém, incluindo recém-nascidos, hemodialisados, transplantados e pacientes HIV+. Foi realizada a pesquisa de anticorpos IgG e IgM pelo método ELISA e análise enzimática pelo método restriction fragment length polymorphism (RFLP) para caracterização dos genótipos virais. RESULTADOS: Foi observado que 100 por cento dos pacientes apresentavam anticorpos IgG, 87 por cento eram IgG+/IgM-perfil de infecção pregressa; e 13 por cento IgG+/ IgM+ sugestivo de infecção recente. O grupo dos recém-nascidos apresentou maior frequência (27 por cento) do perfil IgG+/IgM+. Na análise por RFLP, foi observado um único genótipo, o gB2, que corresponde ao padrão genotípico da cepa AD169. CONCLUSÕES: A presença de anticorpos IgM nos recém-nascidos indica que o vírus CMV continua sendo causa importante de infecção congênita; a baixa diversidade genotípica pode ser atribuída ao tamanho amostral devido a exclusão dos recém-nascidos na análise por RFLP. Esse estudo será continuado incluindo amostras de recém-nascidos a fim de contribuir para um amplo conhecimento da epidemiologia geral e molecular do citomegalovírus em pacientes imunodeficientes da Cidade de Belém.


Subject(s)
Adult , Humans , Infant, Newborn , Cytomegalovirus Infections/virology , Cytomegalovirus/genetics , Genome, Viral/genetics , HIV Infections/immunology , Immunocompromised Host/immunology , Kidney Transplantation/immunology , Brazil , Dialysis , Immunoglobulin G/blood , Immunoglobulin M/blood , Restriction Mapping/methods
13.
J. bras. pneumol ; 37(5): 628-635, set.-out. 2011. tab
Article in Portuguese | LILACS | ID: lil-604390

ABSTRACT

OBJETIVO: Identificar as espécies de micobactérias encontradas no escarro de pacientes com suspeita de tuberculose pulmonar e analisar o impacto dessas identificações na abordagem terapêutica. MÉTODOS: Foram avaliados 106 pacientes com suspeita de tuberculose pulmonar encaminhados para o serviço de pneumologia de um hospital público em Teresina, Piauí. Espécimes de escarro matinal foram avaliados quanto à presença de micobactérias por baciloscopia e cultura. Foram utilizadas PCR e análise de restrição enzimática do gene hsp65 (PRA-hsp65) para a identificação das cepas de micobactérias isoladas em cultura. RESULTADOS: Foram analisadas 206 amostras de escarro. A idade dos pacientes variou de 15 a 87 anos, sendo 67 por cento do gênero masculino. Tosse ocorreu em 100 por cento dos casos. O padrão radiográfico predominante foi de lesão moderada, observada em 70 por cento. A positividade no esfregaço foi de 76 por cento, e isolamento em cultura ocorreu em 91 por cento das culturas executadas. Testes tradicionais identificaram micobactérias não tuberculosas (MNT) em 9 por cento dos isolados. O método PRA-hsp65 confirmou esses dados, mostrando sete padrões de bandas capazes de identificar as espécies de MNT isoladas: Mycobacterium kansasii; M. abscessus 1; M. abscessus 2; M. smegmatis; M. flavescens 1; M. gordonae 5 e M. gordonae 7. Todos os pacientes com MNT tinham mais de 60 anos, e observaram-se bronquiectasias em 88 por cento das radiografias. Houve dois casos de reinfecção, identificados inicialmente como infecção por M. abscessus e M. kansasii. CONCLUSÕES: As MNT causam infecção pulmonar em pacientes imunocompetentes, e a identificação das MNT é importante para estabelecer o diagnóstico correto e a decisão terapêutica mais adequada. O método PRA-hsp65 é útil para identificar espécies de MNT e pode ser implantado em laboratórios de biologia molecular não especializados em micobactérias.


OBJECTIVE: To identify mycobacterial species in the sputum of patients suspected of having pulmonary tuberculosis and to determine the impact that the acquisition of this knowledge has on the therapeutic approach. METHODS: We evaluated 106 patients suspected of having pulmonary tuberculosis and referred to the pulmonology department of a public hospital in the city of Teresina, Brazil. Morning sputum specimens were evaluated for the presence of mycobacteria by sputum smear microscopy and culture. We used PCR and restriction enzyme analysis of the hsp65 gene (PRA-hsp65) to identify the strains of mycobacteria isolated in culture. RESULTS: A total of 206 sputum samples were analyzed. Patient ages ranged from 15 to 87 years, and 67 percent were male. There was cough in 100 percent of the cases. The predominant radiographic pattern was moderate disease, observed in 70 percent. Smear positivity was 76 percent, and isolation in culture occurred in 91 percent of the cultures. Traditional tests identified nontuberculous mycobacteria (NTM) in 9 percent of the isolates. The PRA-hsp65 method confirmed these data, showing seven band patterns that were able to identify the isolated species of NTM: Mycobacterium kansasii; M. abscessus 1; M. abscessus 2; M. smegmatis; M. flavescens 1; M. gordonae 5; and M. gordonae 7. All of the patients with NTM were over 60 years of age, and bronchiectasis was seen in 88 percent of the X-rays. There were two cases of reinfection, initially attributed to M. abscessus and M. kansasii. CONCLUSIONS: In immunocompetent patients, NTM can infect the lungs. It is important to identify the specific NTM in order to establish the correct diagnosis and choose the most appropriate therapeutic regimen. The PRA-hsp65 method is useful in identifying NTM species and can be implemented in molecular biology laboratories that do not specialize in the identification of mycobacteria.


Subject(s)
Adolescent , Adult , Aged , Aged, 80 and over , Female , Humans , Male , Middle Aged , Young Adult , Bacterial Proteins/isolation & purification , /isolation & purification , Nontuberculous Mycobacteria/genetics , Restriction Mapping/methods , Sputum/microbiology , Tuberculosis, Pulmonary/microbiology , Brazil , Bacterial Proteins/genetics , /genetics , Nontuberculous Mycobacteria/classification , Nontuberculous Mycobacteria/isolation & purification
14.
J. bras. pneumol ; 37(4): 521-526, jul.-ago. 2011. ilus, tab
Article in Portuguese | LILACS | ID: lil-597204

ABSTRACT

OBJETIVO: Identificar micobactérias não tuberculosas (MNT) isoladas de sítios estéreis em pacientes internados no Hospital Universitário Clementino Fraga Filho, Rio de Janeiro (RJ) entre 2001 e 2006. MÉTODOS: Durante o período do estudo, 34 isolados de MNT de sítios estéreis de 14 pacientes, a maioria HIV positivos, foram submetidos a identificação fenotípica e hsp65 PCR-restriction enzyme analysis (PRA, análise por enzimas de restrição por PCR do gene hsp65). RESULTADOS: A maioria dos isolados foi identificada como Mycobacterium avium, seguida por M. monacense, M. kansasii e M. abscessus em menores proporções. CONCLUSÕES: A combinação de PRA, um método relativamente simples e de baixo custo, com algumas características fenotípicas pode fornecer a identificação correta de MNT na rotina de laboratórios clínicos.


OBJECTIVE: To identify nontuberculous mycobacteria (NTM) isolated from sterile sites in patients hospitalized between 2001 and 2006 at the Clementino Fraga Filho University Hospital, located in the city of Rio de Janeiro, Brazil. METHODS: During the study period, 34 NTM isolates from sterile sites of 14 patients, most of whom were HIV-positive, were submitted to phenotypic identification and hsp65 PCR-restriction enzyme analysis (PRA). RESULTS: Most isolates were identified as Mycobacterium avium, followed by M. monacense, M. kansasii, and M. abscessus. CONCLUSIONS: The combination of PRA, a relatively simple and inexpensive method, with the evaluation of a few phenotypic characteristics can allow NTM to be accurately identified in the routine of clinical laboratories.


Subject(s)
Adolescent , Adult , Humans , Middle Aged , Young Adult , Bacterial Proteins/analysis , /analysis , Genes, Bacterial/genetics , Mycobacterium Infections, Nontuberculous/microbiology , Nontuberculous Mycobacteria/genetics , Polymerase Chain Reaction/methods , Restriction Mapping/methods , Bacteriological Techniques , Brazil , DNA Restriction Enzymes , DNA, Bacterial/analysis , Hospitals, University , Inpatients , Mycobacterium avium Complex/isolation & purification , Mycobacterium avium-intracellulare Infection/microbiology , Nontuberculous Mycobacteria/isolation & purification
15.
Article in English | IMSEAR | ID: sea-135604

ABSTRACT

Background & objectives: The Janus-associated Kinase-2 mutation JAK2 V617F in chronic myeloproliferative disorders (CMPDs) has been described as a frequent genetic event in majority of patients with polycythemia vera (PV), essential thrombocythemia (ET) and idiopathic myelofibrosis (IMF). Its frequency varies in different populations but there are no data from India. We therefore, looked for JAK2 V617F mutation in Indian patients with chronic myeloproliferative disorders. Methods: Mutation screening for JAK2 V617F in patients with polycythemia vera, essential thrombocythemia and idiopathic myelofibrosis was performed in 75 patients attending Haematology clinic in a tertiary care hospital in north India, by polymerase chain reaction and restriction enzyme-based assay. Results: JAK2 V617F mutation was found in 51 of 75 cases (68%) of CMPD, 82 per cent in PV, 70 per cent in ET and 52 per cent of IMF. The presence of JAK2 V617F mutation was associated with a higher haemoglobin level (P<0.05), a higher white blood cell count (P<0.01) and higher age (P<0.05). Interpretation & conclusion: The JAK2 V617F mutation was detected in 86 per cent of patients with CMPD disorders. Peripheral blood mutation screening for JAK2 V617F can be incorporated into the initial evaluation of patients suspected to have CMPD.


Subject(s)
Age Factors , Electrophoresis, Agar Gel , Genetic Predisposition to Disease/genetics , Hemoglobins/analysis , Humans , India/epidemiology , Janus Kinase 2/genetics , Leukocyte Count , Mutation, Missense/genetics , Myeloproliferative Disorders/epidemiology , Myeloproliferative Disorders/genetics , Polymerase Chain Reaction , Prevalence , Restriction Mapping , Statistics, Nonparametric
16.
Braz. j. med. biol. res ; 43(4): 338-344, Apr. 2010. tab
Article in English | LILACS | ID: lil-543580

ABSTRACT

Lactobacilli isolated from the vaginal tract of women with and without bacterial vaginosis (BV) were identified and characterized for the production of antagonists. Bacterial samples were isolated from healthy women (N = 16), from patients with clinical complaints but without BV (N = 30), and from patients with BV (N = 32). Identification was performed using amplified ribosomal DNA restriction analysis. Production of antagonistic compounds was evaluated by the double-layer diffusion technique using Gram-positive (N = 9) and Gram-negative bacteria (N = 6) as well as yeast (N = 5) as indicator strains. Of a total of 147 isolates, 133 were identified as pertaining to the genus Lactobacillus. Lactobacillus crispatus was the species most frequently recovered, followed by L. johnsonii and L. jensenii. Statistical analysis showed that L. crispatus was more frequent in individuals without BV (P < 0.05). A higher production of antagonists was noted in L. crispatus isolates from healthy women (P < 0.05). More acidic local pH and higher H2O2 production by isolated lactobacilli from healthy women suggest these mechanisms as the possible cause of this antagonism. In conclusion, a significant correlation was detected between the presence and antagonistic properties of certain species of Lactobacillus and the clinical status of the patients.


Subject(s)
Female , Humans , Hydrogen Peroxide/metabolism , Lactobacillus/metabolism , Vagina/microbiology , Vaginosis, Bacterial/microbiology , Bacterial Typing Techniques/methods , DNA, Ribosomal/analysis , DNA, Ribosomal/genetics , Lactobacillus/classification , Lactobacillus/isolation & purification , Restriction Mapping
17.
Scientific and Research Journal of Army University of Medical Sciences-JAUMS. 2010; 8 (3): 193-199
in Persian | IMEMR | ID: emr-146305

ABSTRACT

Staphylococcus aureus has emerged over the past several decades as a leading cause of hospital- and community acquired infection. A significant component in the success has been its acquisition of antibiotic Resistant factors. We have developed a rapid and simplified approach for the strain characterization of staph.aureus on the basis of multilocussequence typing. MLRFT for s.aureus involes amplification of seven house keeping gene locus, the amplicons are then digested directly with one or two restriction Enzyme and the restriction fragments are resolved by agarose gel electrophoresis. These isolates had been additionally characterized for their susceptibilities to antibiotics. MLRFT resolved 146 isolates into 19 RFT that 48 isolates have the same molecular patterns. Their susceptibilities to antibiotics were showed 90% resistance to methicillin. MLRFT provides convenient procedures for molecular epidemiology it requires minimal laboratory facilities and is relatively simple and inexpensive to perform.63% of cluster [24 isolates] take placed two fold cluster that 90% of them was resistant to methicillin, 55% belong to twofold cluster, there fore methicillin resistant isolates can transmit easily


Subject(s)
Humans , Molecular Epidemiology , Hospitals, Military , Restriction Mapping , Methicillin-Resistant Staphylococcus aureus , Methicillin Resistance , Personnel, Hospital , Patients
18.
Chinese Journal of Medical Genetics ; (6): 140-143, 2010.
Article in Chinese | WPRIM | ID: wpr-349025

ABSTRACT

<p><b>OBJECTIVE</b>To identify the RUNX2 gene mutation in two unrelated Chinese families with cleidocranial dysplasia (CCD), and to assess the feasibility of gene diagnosis for patients with CCD.</p><p><b>METHODS</b>Genomic DNA was isolated from peripheral blood samples of 4 patients and 4 healthy members in the two pedigrees as well as 102 unrelated healthy controls. All 7 coding exons and their flanking intronic sequences of the RUNX2 gene were amplified by PCR, then the PCR products were sequenced bi-directionally. The sequencing results were compared with normal sequences in GenBank to identify the mutation. The mutation was confirmed by RFLP with restriction endonuclease.</p><p><b>RESULTS</b>In one family, a novel heterozygous missense mutation c.346T to A (W116R) in exon 1 of the RUNX2 gene was detected in the two affected individuals, and the mutation was further confirmed with Bsr I restriction endonuclease digestion. In the other family, a novel nonsense mutation c.610A TO T (K204X) was identified in the two patients. No above sequence change was found in the 102 healthy controls.</p><p><b>CONCLUSION</b>Two novel RUNX2 mutations were found in two unrelated Chinese families with cleidocranial dysplasia. The identification of these mutations further extended the mutation spectrum of RUNX2 gene and will facilitate prenatal diagnosis and gene diagnosis of CCD.</p>


Subject(s)
Adult , Female , Humans , Infant , Male , Asian People , Genetics , Base Sequence , Case-Control Studies , Cleidocranial Dysplasia , Genetics , Core Binding Factor Alpha 1 Subunit , Genetics , DNA Mutational Analysis , Mutation , Pedigree , Restriction Mapping
19.
Journal of Veterinary Science ; : 221-225, 2010.
Article in English | WPRIM | ID: wpr-79616

ABSTRACT

The aim of this study was to evaluate the presence of Helicobacter (H.) spp. in swine affected by gastric ulceration. Stomachs from 400 regularly slaughtered swine were subjected to gross pathological examination to evaluate the presence of gastric ulcers. Sixty-five samples collected from ulcerated pars esophagea and 15 samples from non-ulcerated pyloric portions were submitted to histopathological and molecular analyses, to detect Helicobacter spp., H. suis and H. pylori by PCR. Feces and saliva swabs were also collected from 25 animals in order to detect in vivo the presence of Helicobacter spp.. Gastric ulcers were detected in 373 cases (93%). The presence of ulcers in association with inflammatory processes was further confirmed by histological examination. Forty-nine percent (32/65) of the ulcerated esophageal portions as well as 53% (8/15) of the non-ulcerated pyloric portions were positive for Helicobacter spp. by PCR. The Helicobacter spp. positive samples were also positive for H. suis, while H. pylori was not detected. These results were confirmed by restriction enzyme analysis. With regard to feces and saliva samples, 15/25 (60%) and 16/25 (64%) were positive for Helicobacter spp. PCR, respectively but all were negative in H. suis and H. pylori specific PCR.


Subject(s)
Animals , Feces/microbiology , Helicobacter/isolation & purification , Polymerase Chain Reaction/veterinary , Restriction Mapping/veterinary , Saliva/microbiology , Stomach/microbiology , Stomach Ulcer/microbiology , Swine , Swine Diseases/microbiology
20.
Indian J Exp Biol ; 2009 Apr; 47(4): 245-9
Article in English | IMSEAR | ID: sea-62384

ABSTRACT

The genome segment 7 of two Indian isolates of bluetongue virus (BTV) from Avikanagar (BTV-1-western India) and Hyderabad (BTV-Untyped Hyderabad-southern India) was amplified by RT-PCR using two sets of VP7 specific primers. A sequence of 1137 bp comprising the complete coding sequence of the VP7 gene from Avikanagar isolate and a 1154 bp full-length sequence from BTV-UT Hyderabad isolate were amplified. Further, restriction enzyme digestion of these full-length amplicons, using EcoRI, PstI and TaqI revealed that genome segment 7 from both isolates were different from each other by absence of any EcoRI site in the BTV-UT Hyderabad isolate. There were also variations in the number and position of restriction sites for TaqI enzyme in these two isolates. Taql has two sites in the Avikanagar isolate whereas four sites in BTV-UT Hyderabad. The restriction digestion fragments obtained after PstI digestion were differentiated on the basis of their distinct sizes in both isolates. Comparison of their in silico restriction profiles with that of other isolates from different countries revealed that the two Indian isolates belonging to different parts of India had variations in their VP7 gene which was also distinguishable from at least some isolates from Australia and South Africa. Hence the restriction enzyme (RE) based analysis might be a useful tool in determining the genetic diversity in genome segment 7 and also for tracing their evolutionary relationships.


Subject(s)
Animals , Bluetongue virus/genetics , Bluetongue virus/isolation & purification , Computational Biology , Genes, Viral , India , Polymerase Chain Reaction , Restriction Mapping , Viral Core Proteins/genetics
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