Desenvolvimento e avaliação da acurácia do sistema de filtração (BacFil 6.0) na detecção de bacilos álcool-ácido resistentes em amostras de lavado broncoalveolar de pacientes com suspeita de tuberculose
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Data
2014-07-22
Autores
Almeida Junior, Pedro Sousa de
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Universidade Federal do Espírito Santo
Resumo
The smear microscopy is the main diagnostic method of tuberculosis (TB) due to its low cost, fastness and easy execution. However, its sensitivity is low especially in the paucibacillary forms of the disease. Therefore, new methods that enhance the sensitivity of smear microscopy are required. In this study, we proposed to improve and standardize the BacFil filtration system and to evaluate its performance and accuracy in bronchoalveolar lavage (BAL). To achieve these goals we developed a study in two stages. In the first stage we conducted assays to define: 1) use of polycarbonate white membranes to visualize fluorescent AFB stained by Auramine- O; 2) increase the volume of NALC, addition of glass beads in the digestion procedures and changes in the prefilter support; 3) ways to fixate the membrane on the microscopy slides; 4) the detection limit of the technique; 5) increased concentration of NaOCl and its influence to visualize AFB on the filtration’s membrane. It was verified that: the polycarbonate white membrane has the same efficiency of the polycarbonate black membrane to visualize fluorescent AFB; the modified digestion method using higher volume of NALC and glass beads plus prefilter support system based on holes allowed 98% of the samples to pass trough the filtration system; the best way to fixate the membrane on the slides was with double-side tape; the detection limit of the technique using the BacFil system version 6.0 was 2 to 9 AFB per milliliter of a bacterial suspension with O.D. = 0,250 ± 0,01; the use of NaOCl 10% (v/v) helps the passage of a higher volume of samples through the system, however it causes AFB loss on the filtration membrane after the time needed to finish all the process. In the second stage it was evaluated the accuracy of the standardized technique in 101 BAL of patients with suspected TB coming from Hospital Universitário Cassiano Antônio de Moraes. We observed a higher sensitivity of filtration technique using the BacFil version 6.0 system (76%) when compared to the concentrated smear microscopy (59%), and this difference is statistically significant (p = 0,001). However, it was detected a lower specificity of the filtration technique (95%) in comparison with centrifuged smear microscopy (99%). We concluded that the modifications made on the filtration system allowed us to solve the techinicals issues reported in previous versions, and to enhance the sensitivity of smear microscopy in paucibacillary samples (BAL) for the TB diagnosis.