Abstract
Lymph node tuberculosis (TB) is the leading cause of extrapulmonary tuberculosis and is the most frequently identified type in Aguascalientes, Mexico. Conventional diagnosis has serious limitations for rapid detection of extrapulmonary tuberculosis in clinical samples. Here PCR and modified FISH have been tested as complementary diagnosis methods for extrapulmonary tuberculosis. The specific insertion sequence IS6110 for Mycobacterium tuberculosis complex was used to perform PCR and build DNA and PNA FISH probes (20bp). PCR and modified DNA and PNA FISH assays were performed to evaluate 41 lymph node paraffin-embedded tissue samples, in comparison with the histopathology diagnosis, which was considered the gold standard (22 positive and 19 negative). In comparison with histopathology diagnosis PCR showed 62.5 % sensitivity and 77.8 % specificity (χ(2) = 4.583 p < 0.05). Modified DNA FISH showed 71.4% sensitivity and 84.6% specificity (χ(2) = 11.21 p < 0.05). PNA FISH showed 66.7% sensitivity and 60.0% specificity (χ(2) = 2.93 p > 0.05). Ziehl Neelsen stain was positive in only four cases of 22 lymph node samples positive to histopathology. In contrast, PCR and modified DNA FISH were positive in 20 cases of the same group. The negative cases were coincident in all tests. PCR and DNA FISH showed a significant increase in the number of cases detected and also showed higher sensitivity and specificity compared with data reported by traditional methodology. In developing countries, these techniques could help to complement the early diagnosis and timely treatment of extrapulmonary tuberculosis.
Highlights
Lymph node tuberculosis (TB) is the leading cause of extrapulmonary tuberculosis and is the most frequently identified type in Aguascalientes, Mexico
On the histopathological examination positive (HEP), PCR IS6110 was positive in 20 samples and negative in two samples
Fluorescent in-situ hybridization (FISH) As a first test of specificity, IS6110 DNA and peptide nucleic acid (PNA) oligonucleotide fluorescent probes were applied to pure cultures, including M. tuberculosis H37Rv (American Type Culture Collection, USA,) and M. bovis BCG ATCC 35734, which were used as positive inner controls
Summary
Lymph node tuberculosis (TB) is the leading cause of extrapulmonary tuberculosis and is the most frequently identified type in Aguascalientes, Mexico. Methodology: The specific insertion sequence IS6110 for Mycobacterium tuberculosis complex was used to perform PCR and build DNA and PNA FISH probes (20bp). PCR and modified DNA and PNA FISH assays were performed to evaluate 41 lymph node paraffin-embedded tissue samples, in comparison with the histopathology diagnosis, which was considered the gold standard (22 positive and 19 negative). Conclusions: PCR and DNA FISH showed a significant increase in the number of cases detected and showed higher sensitivity and specificity compared with data reported by traditional methodology. In developing countries, these techniques could help to complement the early diagnosis and timely treatment of extrapulmonary tuberculosis. This diagnosis depends mainly on the bacilli culture obtained after fine needle aspiration biopsy or excisional biopsy, with the microscopic demonstration of the acid-fast bacilli or Ziehl
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