Nucleo de Pos Graduacao e Pesquisa da Santa Casa de Belo Horizonte

Belo Horizonte, Brazil

Nucleo de Pos Graduacao e Pesquisa da Santa Casa de Belo Horizonte

Belo Horizonte, Brazil
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Rocha-Silva F.,Nucleo de Pos Graduacao e Pesquisa da Santa Casa de Belo Horizonte | Gracielle-Melo C.,Nucleo de Pos Graduacao e Pesquisa da Santa Casa de Belo Horizonte | Goes A.M.,Federal University of Minas Gerais | Caligiorne R.B.,Nucleo de Pos Graduacao e Pesquisa da Santa Casa de Belo Horizonte
Recent Patents on Endocrine, Metabolic and Immune Drug Discovery | Year: 2016

Background: Paracoccidioidomycosis (PCM) is a systemic mycosis caused by dimorphic fungi Paracoccidioides brasiliensis and Paracoccidioides lutzii. It is prevalent in Latin American, mainly in Brazil. Therefore, PCM has fundamental impact on the Brazilian global economy, especially in public health system, since it is affecting economical active population in different country regions. Objective: The present study aimed to standardize the Real Time-Polymerase Chain Reaction (rt-PCR) for an efficient and safe PCM diagnosis amplifying the recombinant protein PB27 gene, only expressed by specimens of Paracoccidioides genus. Methods: To standardize a methodology of rt-PCR using species-specific primers and probe designed for annealing in this specific region of the fungi´s genome, amplifying the recombinant protein PB27 gene, only expressed by specimens of Paracoccidioides genus. Followed by design in silico, experiments were performed in vitro to determine rt-PCR specificity, efficiency and genome detection limit. Results: The primers and probe sequences were deposited in Brazilian Coordination of Technological Innovation and Transfer (CTIT), under patent reference number BR1020160078830. The present study demonstrated the rt-PCR applicability for support on diagnosis of paracoccidioidomycosis, presenting low cost, which makes it affordable for public health services in developing countries as Brazil. It is noteworthy that it is necessary to validate this methodology using clinical samples before to use as a safe method of diagnosis. A review of all patents related to this topic was performed and it was shown that, to date, there are no records of patent on kits for paracoccidioidomycosis´s diagnostic. Indeed, there is still a lot to go to reach this goal. Conclusion: The reaction developed was standardized and patented, opening perspectives to molecular diagnosis development for paracoccidioidomycosis, since rt-PCR can be applied to a broad spectrum of infectious diseases. It would need to be tested in biological samples in order to validate this method and then generate a diagnostic kit for Paracoccidioidomycosis. © 2016 Bentham Science Publishers.


Sandrim V.C.,Nucleo de Pos Graduacao e Pesquisa da Santa Casa de Belo Horizonte | Palei A.C.T.,University of Campinas | Sertorio J.T.,University of Sao Paulo | Cavalli R.C.,University of Sao Paulo | And 2 more authors.
Molecular Human Reproduction | Year: 2010

Pre-eclampsia (PE) is associated with decreased nitric oxide (NO) formation. However, no previous study has examined whether genetic variations in the endothelial NO synthase (eNOS) affect this alteration. We hypothesized that PE decreases NO formation depending on eNOS polymorphisms. We examined how three eNOS polymorphisms [T-786C, rs2070744; Glu298Asp, rs1799983; 27 bp variable number of tandem repeats (VNTR) in intron 4] affect plasma nitrite concentrations in 205 pregnant women [107 healthy pregnant (HP) and 98 PE]. Genotypes were determined and eNOS haplotypes were inferred using the PHASE 2.1 program. The plasma nitrite concentrations were determined using an ozone-based chemiluminescence assay. The Glu298Asp polymorphism had no effects on the plasma nitrite concentrations. Higher nitrite levels were found in HP women with the CC versus TT genotype for the T-786C polymorphism (277.9±19.5 versus 140.6±8.2 nM; P < 0.05). Lower nitrite levels were found in healthy women with the 4a4a versus 4b4b genotype for the VNTR polymorphism (95.1±3.3 versus 216.1±16.8 nM; P < 0.05). No effects of genotypes were found in PE women (all P > 0.05). The 'C Glu b' haplotype was more frequent in the HP group than in the PE group (20 versus 5; P = 0.0044). This haplotype was associated with higher nitrite concentrations than the other haplotypes in healthy pregnancies (P < 0.05). No differences in nitrite concentrations were found among PE women with different eNOS haplotypes (P > 0.05). These findings indicate that eNOS polymorphisms affect endogenous NO formation in normal pregnancy, but not in PE, and that the 'C Glu b' haplotype may protect against the development of PE by increasing endogenous NO formation. © The Author 2010. Published by Oxford University Press on behalf of the European Society of Human Reproduction and Embryology. All rights reserved. For Permissions, please email: journals.permissions@oxfordjournals.org.

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