Identification of fungemia agents using the polymerase chain reaction and restriction fragment length polymorphism analysis
Prompt and specific identification of fungemia agents is important in order to define clinical treatment. However, in most cases conventional culture identification can be considered to be time-consuming and not without errors. The aim of the present study was to identify the following fungemia agen...
| Main Authors: | , , , , |
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| Format: | Article |
| Language: | English |
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Associação Brasileira de Divulgação Científica
2010-08-01
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| Series: | Brazilian Journal of Medical and Biological Research |
| Subjects: | |
| Online Access: | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2010000800003 |
| _version_ | 1828736918375890944 |
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| author | M.S. Santos E.S. Souza R.M.S. Junior S. Talhari J.V.B. Souza |
| author_facet | M.S. Santos E.S. Souza R.M.S. Junior S. Talhari J.V.B. Souza |
| author_sort | M.S. Santos |
| collection | DOAJ |
| description | Prompt and specific identification of fungemia agents is important in order to define clinical treatment. However, in most cases conventional culture identification can be considered to be time-consuming and not without errors. The aim of the present study was to identify the following fungemia agents: Candida albicans, Candida parapsilosis, Candida tropicalis, Candida glabrata, Cryptococcus neoformans, Cryptococcus gattii, and Histoplasma capsulatum using the polymerase chain reaction and restriction fragment length polymorphism analysis (PCR/RFLP). More specifically: a) to evaluate 3 different amplification regions, b) to investigate 3 different restriction enzymes, and c) to use the best PCR/RFLP procedure to indentify 60 fungemia agents from a culture collection. All 3 pairs of primers (ITS1/ITS4, NL4/ITS5 and Primer1/Primer2) were able to amplify DNA from the reference strains. However, the size of these PCR products did not permit the identification of all the species studied. Three restriction enzymes were used to digest the PCR products: HaeIII, Ddel and Bfal. Among the combinations of pairs of primers and restriction enzymes, only one (primer pair NL4/ITS5 and restriction enzyme Ddel) produced a specific RFLP pattern for each microorganism studied. Sixty cultures of fungemia agents (selected from the culture collection of Fundação de Medicina Tropical do Amazonas - FMTAM) were correctly identified by PCR/RFLP using the prime pair NL4/ITS5 and Ddel. We conclude that the method proved to be both simple and reproducible, and may offer potential advantages over phenotyping methods. |
| first_indexed | 2024-04-12T23:35:08Z |
| format | Article |
| id | doaj.art-837ec17f8a5c4ac895886d7fdfc12724 |
| institution | Directory Open Access Journal |
| issn | 0100-879X 1414-431X |
| language | English |
| last_indexed | 2024-04-12T23:35:08Z |
| publishDate | 2010-08-01 |
| publisher | Associação Brasileira de Divulgação Científica |
| record_format | Article |
| series | Brazilian Journal of Medical and Biological Research |
| spelling | doaj.art-837ec17f8a5c4ac895886d7fdfc127242022-12-22T03:12:10ZengAssociação Brasileira de Divulgação CientíficaBrazilian Journal of Medical and Biological Research0100-879X1414-431X2010-08-01438712716Identification of fungemia agents using the polymerase chain reaction and restriction fragment length polymorphism analysisM.S. SantosE.S. SouzaR.M.S. JuniorS. TalhariJ.V.B. SouzaPrompt and specific identification of fungemia agents is important in order to define clinical treatment. However, in most cases conventional culture identification can be considered to be time-consuming and not without errors. The aim of the present study was to identify the following fungemia agents: Candida albicans, Candida parapsilosis, Candida tropicalis, Candida glabrata, Cryptococcus neoformans, Cryptococcus gattii, and Histoplasma capsulatum using the polymerase chain reaction and restriction fragment length polymorphism analysis (PCR/RFLP). More specifically: a) to evaluate 3 different amplification regions, b) to investigate 3 different restriction enzymes, and c) to use the best PCR/RFLP procedure to indentify 60 fungemia agents from a culture collection. All 3 pairs of primers (ITS1/ITS4, NL4/ITS5 and Primer1/Primer2) were able to amplify DNA from the reference strains. However, the size of these PCR products did not permit the identification of all the species studied. Three restriction enzymes were used to digest the PCR products: HaeIII, Ddel and Bfal. Among the combinations of pairs of primers and restriction enzymes, only one (primer pair NL4/ITS5 and restriction enzyme Ddel) produced a specific RFLP pattern for each microorganism studied. Sixty cultures of fungemia agents (selected from the culture collection of Fundação de Medicina Tropical do Amazonas - FMTAM) were correctly identified by PCR/RFLP using the prime pair NL4/ITS5 and Ddel. We conclude that the method proved to be both simple and reproducible, and may offer potential advantages over phenotyping methods.http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2010000800003PCR/RFLPFungemia agentsIdentificationDdel |
| spellingShingle | M.S. Santos E.S. Souza R.M.S. Junior S. Talhari J.V.B. Souza Identification of fungemia agents using the polymerase chain reaction and restriction fragment length polymorphism analysis Brazilian Journal of Medical and Biological Research PCR/RFLP Fungemia agents Identification Ddel |
| title | Identification of fungemia agents using the polymerase chain reaction and restriction fragment length polymorphism analysis |
| title_full | Identification of fungemia agents using the polymerase chain reaction and restriction fragment length polymorphism analysis |
| title_fullStr | Identification of fungemia agents using the polymerase chain reaction and restriction fragment length polymorphism analysis |
| title_full_unstemmed | Identification of fungemia agents using the polymerase chain reaction and restriction fragment length polymorphism analysis |
| title_short | Identification of fungemia agents using the polymerase chain reaction and restriction fragment length polymorphism analysis |
| title_sort | identification of fungemia agents using the polymerase chain reaction and restriction fragment length polymorphism analysis |
| topic | PCR/RFLP Fungemia agents Identification Ddel |
| url | http://www.scielo.br/scielo.php?script=sci_arttext&pid=S0100-879X2010000800003 |
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