Detection of Histoplasma Capsulatum by Nested-PCR in Clinical Specimens
Histoplasmosis is a systemic mycosis whose diagnosis is based on clinical, mycological and immunological criteria. Identification of its etiological agent in biological samples is difficult to perform. We evaluated a nested-PCR technique to identify Histoplasma capsulatum in clinical specimens (bloo...
| Main Authors: | , , , , , , , , |
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| Format: | Article |
| Language: | English |
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Universidad del Zulia,Facultad de Medicina,Departamento de Enfermedades Infecciosas y Tropicales
2007-12-01
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| Series: | Kasmera |
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| Online Access: | https://produccioncientificaluz.org/index.php/kasmera/article/view/4794 |
| _version_ | 1827846627339010048 |
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| author | María Teresa Colella Sofía Mata Claudia Hartung Celina Pérez Arantza Roselló Carolina Olaizola Carmen Fernández Silvia Magaldi Félix Toro |
| author_facet | María Teresa Colella Sofía Mata Claudia Hartung Celina Pérez Arantza Roselló Carolina Olaizola Carmen Fernández Silvia Magaldi Félix Toro |
| author_sort | María Teresa Colella |
| collection | DOAJ |
| description | Histoplasmosis is a systemic mycosis whose diagnosis is based on clinical, mycological and immunological criteria. Identification of its etiological agent in biological samples is difficult to perform. We evaluated a nested-PCR technique to identify Histoplasma capsulatum in clinical specimens (blood and bone marrow) and reference strains. Three different methods for DNA isolation were tested based on the reagents Guanidine thiocyanate, Benzoyl chloride and Triton X-100. Among these procedures, Guanidine thiocyanate-based method yielded by utmost good qualityDNAand was easy to perform when used with clinical specimens. However, a better recovery of DNA was accomplished with the Triton-X-100 method. Reference strains of Paracoccidioides brasiliensis, Candida glabrata, Cryptococcus neoformans and Coccidioides immitis (C posadasii) were tested to assess PCR assay specificity. The samples were amplified by nested-PCR and analyzed by 3% agarose gel electrophoresis. Each of the reference strains showed the same signal upon amplification with a DNA band of 210 bases pairs, as well as 5 blood samples and 2 bone marrow specimens, with high sensitivity (10 picograms of Histoplasma capsulatum DNA) and specificity (no cross reactivity with P. brasiliensis, C. glabrata, C. neoformans and C. immitis). This technique has proved to be useful as a diagnostic technique. |
| first_indexed | 2024-03-12T09:18:48Z |
| format | Article |
| id | doaj.art-0d31ff9116a94e549776ef6d2b4a1900 |
| institution | Directory Open Access Journal |
| issn | 0075-5222 2477-9628 |
| language | English |
| last_indexed | 2024-03-12T09:18:48Z |
| publishDate | 2007-12-01 |
| publisher | Universidad del Zulia,Facultad de Medicina,Departamento de Enfermedades Infecciosas y Tropicales |
| record_format | Article |
| series | Kasmera |
| spelling | doaj.art-0d31ff9116a94e549776ef6d2b4a19002023-09-02T14:39:44ZengUniversidad del Zulia,Facultad de Medicina,Departamento de Enfermedades Infecciosas y TropicalesKasmera0075-52222477-96282007-12-013521561634793Detection of Histoplasma Capsulatum by Nested-PCR in Clinical SpecimensMaría Teresa Colella0Sofía Mata1Claudia Hartung2Celina Pérez3Arantza Roselló4Carolina Olaizola5Carmen Fernández6Silvia Magaldi7Félix Toro8Sección de Micología Médica, Instituto de Medicina Tropical, Universidad Central de Venezuela, Caracas, Venezuela.Sección de Micología Médica, Instituto de Medicina Tropical, Universidad Central de Venezuela, Caracas, Venezuela.Sección de Micología Médica, Instituto de Medicina Tropical, Universidad Central de Venezuela, Caracas, Venezuela.Sección de Micología Médica, Instituto de Medicina Tropical, Universidad Central de Venezuela, Caracas, Venezuela.Sección de Micología Médica, Instituto de Medicina Tropical, Universidad Central de Venezuela, Caracas, Venezuela.Cátedra de Medicina Tropical, Escuela Luis Razetti, Facultad de Medicina. Universidad Central de Venezuela.Cátedra de Medicina Tropical, Escuela Luis Razetti, Facultad de Medicina. Universidad Central de Venezuela.Cátedra de Medicina Tropical, Escuela Luis Razetti, Facultad de Medicina. Universidad Central de Venezuela.Laboratorio de Biología Molecular, Instituto de Inmunología, Universidad Central de Venezuela, Caracas, Venezuela.Histoplasmosis is a systemic mycosis whose diagnosis is based on clinical, mycological and immunological criteria. Identification of its etiological agent in biological samples is difficult to perform. We evaluated a nested-PCR technique to identify Histoplasma capsulatum in clinical specimens (blood and bone marrow) and reference strains. Three different methods for DNA isolation were tested based on the reagents Guanidine thiocyanate, Benzoyl chloride and Triton X-100. Among these procedures, Guanidine thiocyanate-based method yielded by utmost good qualityDNAand was easy to perform when used with clinical specimens. However, a better recovery of DNA was accomplished with the Triton-X-100 method. Reference strains of Paracoccidioides brasiliensis, Candida glabrata, Cryptococcus neoformans and Coccidioides immitis (C posadasii) were tested to assess PCR assay specificity. The samples were amplified by nested-PCR and analyzed by 3% agarose gel electrophoresis. Each of the reference strains showed the same signal upon amplification with a DNA band of 210 bases pairs, as well as 5 blood samples and 2 bone marrow specimens, with high sensitivity (10 picograms of Histoplasma capsulatum DNA) and specificity (no cross reactivity with P. brasiliensis, C. glabrata, C. neoformans and C. immitis). This technique has proved to be useful as a diagnostic technique.https://produccioncientificaluz.org/index.php/kasmera/article/view/4794histoplasmosisPCRADN |
| spellingShingle | María Teresa Colella Sofía Mata Claudia Hartung Celina Pérez Arantza Roselló Carolina Olaizola Carmen Fernández Silvia Magaldi Félix Toro Detection of Histoplasma Capsulatum by Nested-PCR in Clinical Specimens Kasmera histoplasmosis PCR ADN |
| title | Detection of Histoplasma Capsulatum by Nested-PCR in Clinical Specimens |
| title_full | Detection of Histoplasma Capsulatum by Nested-PCR in Clinical Specimens |
| title_fullStr | Detection of Histoplasma Capsulatum by Nested-PCR in Clinical Specimens |
| title_full_unstemmed | Detection of Histoplasma Capsulatum by Nested-PCR in Clinical Specimens |
| title_short | Detection of Histoplasma Capsulatum by Nested-PCR in Clinical Specimens |
| title_sort | detection of histoplasma capsulatum by nested pcr in clinical specimens |
| topic | histoplasmosis PCR ADN |
| url | https://produccioncientificaluz.org/index.php/kasmera/article/view/4794 |
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