Real-time pcr and nested-pcr assays for detection of Pneumocystis sp. in lung tissues of bats / Real-time pcr and nested-pcr assays for detection of Pneumocystis sp. in lung tissues of bats
Maria Cavallini Sanches, Edna; Ferreiro, Laerte; Pinto de Andrade, Caroline; Missel Pacheco, Susi; Morais Santurio, Janio; Lopes Almeida, Laura; Spanamberg, Andréia; Wissmann, Gustavo.
Acta sci. vet. (Online);
40(4): 01-05, 2012.
Artigo
em Inglês
| VETINDEX
| ID: vti-475553
Resumo
Background: Pneumocystis constitutes a highly diversifi ed biological group, with numerous species, which are strongly host-specifi c and well adapted to live inside the lungs of a diverse range of mammals. The detection of DNA from Pneumocystis in clinical specimens by PCR assays is leading to important advances in pneumonia caused by Pneumocystis and its epidemiology. The aim of this study was to analyze two different diagnostic methods, real-time PCR (qPCR) using primers based in the Major Surface Glycoprotein (MSG) of Pneumocystis sp. and conventional nested PCR using primers designed to the small subunit of mitochondrial rRNA (mtSSU rRNA) for detection of Pneumocystis DNA in lung tissue from bats.Materials, Methods & Results: Bats (195 samples) were captured (2007-2009) in caves, forests, and urban areas, were obtained from the Program of Rabies Control of two states in Brazil: Mato Grosso and Rio Grande do Sul, located respectively in the Mid-Western and Southern regions of the country approximately 2000 km apart. Lung tissue (250 mg) was fi nely minced, homogenized with crushing and DNA extraction was carried out with commercial kit. DNA samples the lung tissue of bats were analyzed by nested PCR, using oligonucleotide primers designed for the gene encoding the mitochondrial small subunit r RNA (mtSSU rRNA) and Taqman probe and primers for qPCR were selected based onBackground: Pneumocystis constitutes a highly diversifi ed biological group, with numerous species, which are strongly host-specifi c and well adapted to live inside the lungs of a diverse range of mammals. The detection of DNA from Pneumocystis in clinical specimens by PCR assays is leading to important advances in pneumonia caused by Pneumocystis and its epidemiology. The aim of this study was to analyze two different diagnostic methods, real-time PCR (qPCR) using primers based in the Major Surface Glycoprotein (MSG) of Pneumocystis sp. and conventional nested PCR using primers designed to the small subunit of mitochondrial rRNA (mtSSU rRNA) for detection of Pneumocystis DNA in lung tissue from bats.Materials, Methods & Results: Bats (195 samples) were captured (2007-2009) in caves, forests, and urban areas, were obtained from the Program of Rabies Control of two states in Brazil: Mato Grosso and Rio Grande do Sul, located respectively in the Mid-Western and Southern regions of the country approximately 2000 km apart. Lung tissue (250 mg) was fi nely minced, homogenized with crushing and DNA extraction was carried out with commercial kit. DNA samples the lung tissue of bats were analyzed by nested PCR, using oligonucleotide primers designed for the gene encoding the mitochondrial small subunit r RNA (mtSSU rRNA) and Taqman probe and primers for qPCR were selected based on
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