Navegando por Palavras-chave "Real-time PCR"

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    Differentiation of Klebsiella pneumoniae carbapenemase (KPC) variants by pyrosequencing
    (Elsevier B.V., 2014-05-01) Monteiro, Jussimara [UNIFESP]; Widen, Raymond H.; Pignatari, Antonio C. C. [UNIFESP]; Kubasek, Carly; Silbert, Suzane; Tampa Gen Hosp; Universidade Federal de São Paulo (UNIFESP)
    A fast and reliable protocol using the pyrosequencing technique was developed to identify 11 different types of the KPC enzyme. A total of 65 b/alpha(Kpc) positive bacterial isolates were tested and characterized. in the end, the pyrosequencing proved to be a powerful tool for epidemiological studies of KPC producer isolates. (c) 2014 Elsevier B.V. All rights reserved.
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    Frequency of Toxoplasma gondii in the retina in eye banks in Brazil
    (Springer, 2017) Costa, Deise F. [UNIFESP]; Nascimento, Heloisa [UNIFESP]; Sutili, Aline [UNIFESP]; Nobrega, Fernando A. J.; Fowler, Flavio [UNIFESP]; Nobrega, Mario Junqueira; Garrido, Cristina; Dias, Janaina de Oliveira; Adan, Consuelo B. D.; Rizzo, Luiz Vicente; Silveira, Claudio; Belfort, Rubens, Jr. [UNIFESP]; Commodaro, Alessandra G. [UNIFESP]
    Ocular toxoplasmosis is the main cause of posterior uveitis worldwide frequently leading to vision loss. In Brazil, the seroprevalence of Toxoplasma gondii infection ranges from 50 to 80% depending of the region studied. The frequency of toxoplasmic retinal scar may reach 18% of the adults in the South of Brazil. Our goal was to determine the frequency of T. gondii DNA in retinas from eye banks from different regions in Brazil. A total of 162 eyes were obtained from eye banks in Manaus (n = 60), Sao Paulo (n = 60), Chapeco (n = 26), and Joinville (n = 16). The retinas were macroscopically analyzed and collected for DNA extraction. Real-time PCR (qPCR) was performed using the T. gondii B1 marker. By qPCR, a higher frequency of T. gondii DNA in the retinas from the eye bank of Joinville (25%) was found when compared to Manaus (5%). The retinas from Sao Paulo and Chapeco were qPCR negative. Clinical examination determined the retina lesions to be compatible with toxoplasmosis in the following frequencies: Joinville (62.5%), Manaus (10%), Sao Paulo (6.7%), and Chapeco (15.4%).
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    High resolution melting versus sequenciamento genético de nova geração na detecção de mutação do gene KRAS em câncer colorretal
    (Universidade Federal de São Paulo (UNIFESP), 2017-05-30) Miranda, Raelson Rodrigues [UNIFESP]; Forones, Nora Manoukian [UNIFESP]; Silva, Tiago Donizetti da [UNIFESP]; http://lattes.cnpq.br/0950776650887037; http://lattes.cnpq.br/7314943504526739; http://lattes.cnpq.br/5692801803165299; Universidade Federal de São Paulo (UNIFESP)
    Introduction: Frequency of KRAS gene mutations in colorectal cancer is around 34% and confers poor prognosis. The most frequency are on exon 2, 3 and 4. The knowledge about KRAS profile plays crucial role in the management of this disease, because the use of MABs like cetuximab and panitumumab are recommended exclusively for KRAS wild-type tumors. In other words, it has important predictive value. PCR-High-Resolution melting (HRM) analysis is a novel technology able to detect mutations with high sensitivity through of variations in melting temperature of DNA with low cost than sequencing methods. Therefore, could be a good tool for screening of KRAS mutations. Objective: Comparing capacity of HRM to detect exons 2, 3 and 4 KRAS mutations with Next Generation Sequencing. Methods: We performed PCR-HRM runs for 47 patients with confirmed colorectal adenocarcinoma in StepOne plus Real-time PCR (Thermo-Fisher/Life Technology©) and compared the melting temperature and HRM curve between mutated and wild-type groups. We used the Illumina HiSeq platform® as reference methods for the Next Generation Sequencing. The statistical analysis was performed through SPSS software v20.1. Results: For exon 2, the melt curves were different between mutant and wild-type groups. The mean of melting temperature was 78.13 ºC for wild-type and 77.87 ºC for mutant profile (p= 0.001; CI 95%: 0,11-0,41; T-test). Sensitivity was 93.8%, specificity 96.6%, PPV 92.2%, NPV 98.51%, accuracy 91.5% and false positive and negative rates 3,4% for PCR-HRM. In terms of exons 3 and 4, melt curve were not different. Conclusions: In this study, the high resolution melting curve profile proved be a good test for screening of exon 2 KRAS mutations in colorectal cancer with high sensitivity and specificity. However, for exons 3 and 4, HRM was not able to detect mutations.
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    Influenza virus prevalence in asymptomatic and symptomatic subjects during pandemic and postpandemic periods
    (Elsevier B.V., 2015-05-01) Melchior, Thaís Boim [UNIFESP]; Perosa, Ana Helena [UNIFESP]; Camargo, Clarice Neves [UNIFESP]; Granato, Celso [UNIFESP]; Bellei, Nancy [UNIFESP]; Universidade Federal de São Paulo (UNIFESP)
    Background: in 2009, Influenza A(H1N1) pdm09 had a major effect on global health, causing thousands of deaths. However, the results of asymptomatic infection in transmission has not been well studied. We analyzed asymptomatic influenza infection in individuals with different risk factors for acquiring influenza in a university hospital.Methods: Respiratory samples from 100 children with respiratory symptoms, their asymptomatic care-givers, 100 asymptomatic HIV-infected patients, and 100 health care workers collected during 2009-2011 were tested for influenza by real time real-time polymerase chain reaction.Results: Influenza infection rate in symptomatic children was 44% and in asymptomatic adults it was 8.3% (P < .01). Children older than age 5 years had a 3.4 times greater chance of being infected during influenza season than younger children. Among the asymptomatic group, influenza was more frequent in caregivers (14%; P = .032) and a higher rate (31.8%) was observed if a child was infected during the 2009 pandemic. Contact with an infected child was an important risk factor for influenza acquisition (odds ratio, 3.8; 95% confidence interval, 1.1-13.2) among caregivers. the mean cluster of differentiation 4 T-cell count of HIV-positive patients infected with influenza was not statistically different from uninfected patients (P = .29).Conclusions: Asymptomatic household contacts of symptomatic children may play an important role in community transmission and a more proactive intervention should be considered during future pandemics. Copyright (C) 2015 by the Association for Professionals in Infection Control and Epidemiology, Inc. Published by Elsevier Inc. All rights reserved.