Detection of Paracoccidioides brasiliensis in Tissue Samples by a Nested PCR Assay
AUTOR(ES)
Bialek, Ralf
FONTE
American Society for Microbiology
RESUMO
A nested PCR assay for the detection of Paracoccidioides brasiliensis DNA was evaluated, using a sequence of the immunogenic gp43 gene as a target. This gene encodes an outer membrane protein unique to this dimorphic fungus. DNA from six clinical isolates and the ATCC strain 60885 of P. brasiliensis, as well as DNA from closely related fungi, was examined to determine detection limits and cross-reactions. PCR was done on DNA extracts of lung homogenates from 23 experimentally P. brasiliensis-infected and two uninfected BALB/c mice and from 20 Histoplasma capsulatum-infected ICR mice. The results were compared to quantitative cultures. A detection limit of 0.5 fg of specific DNA was determined using cloned plasmid DNA. In all seven P. brasiliensis isolates, the expected 196-bp nested PCR product was found. Their sequences were 100% identical to the gp43 gene sequence in GenBank. DNA extracts of all other, related fungi were negative. The PCR assay was positive in 21 out of 23 culture-positive lung homogenates with concentrations of 1 × 103 to 1.3 × 107 CFU of P. brasiliensis per g of lung. Uninfected BALB/c mice and H. capsulatum-infected mice samples gave negative results. The high sensitivity and specificity of this new nested PCR assay for the detection of P. brasiliensis in tissue samples were demonstrated. The assay may be useful for diagnosis in human tissue samples.
ACESSO AO ARTIGO
http://www.pubmedcentral.nih.gov/articlerender.fcgi?artid=87153Documentos Relacionados
- An optimized one-tube, semi-nested PCR assay for Paracoccidioides brasiliensis detection
- Detection of Sporothrix schenckii in Clinical Samples by a Nested PCR Assay
- Detection of Cryptococcus neoformans DNA in Tissue Samples by Nested and Real-Time PCR Assays
- Improved Detection of Rhinoviruses in Clinical Samples by Using a Newly Developed Nested Reverse Transcription-PCR Assay
- Semi-nested PCR para a detecção molecular do fungo Paracoccidioides brasiliensis