Acid-fast bacterium detection and identification from paraffin-embedded tissues using a PCR-pyrosequencing method.
J Clin Pathol
; 71(2): 148-153, 2018 Feb.
Article
em En
| MEDLINE
| ID: mdl-28735303
ABSTRACT
AIMS:
Acid-fast bacterium (AFB) identification from formalin-fixed paraffin-embedded (FFPE) tissues is challenging and may not be readily available to the clinical laboratory. A method to detect and identify AFB from FFPE tissues using PCR and pyrosequencing (PCR-Seq) was developed and evaluated.METHODS:
The method was validated using spiked cell-clotted paraffin blocks before use with patients' specimens. DNA was extracted from tissue sections, and a 16S rRNA gene fragment was amplified and a signature sequence was produced on a PyroMark ID system. Sequences were aligned to established databases for AFB identification. Additional tissue sections were stained and examined for AFB.RESULTS:
Both sensitivity and specificity were 100% on spiked cell-clotted blocks without cross-reactivity with non-AFB. Of 302 FFPE tissues from patients, 116 (38%) were AFB-stain positive; 83 (72%) of these had AFB identified. The 21 AFB identified included Mycobacterium tuberculosis complex (14 cases), Mycobacterium leprae (3), Mycobacterium genavense (2), Mycobacterium marinum-ulcerans group (3) and 17 other AFB (61). Thirteen cases were AFB-stain indeterminate and 4 were positive by the PCR-Seq method. Of the AFB stain-negative cases, 167 were negative and 6 were positive by PCR-Seq.CONCLUSIONS:
The PCR-Seq method provided specific identification of various AFB species or complexes from FFPE tissues.Palavras-chave
Texto completo:
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Tema:
Epidemiologia
/
Geral
Bases de dados:
MEDLINE
Assunto principal:
DNA Bacteriano
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Reação em Cadeia da Polimerase
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Fixação de Tecidos
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Inclusão em Parafina
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Análise de Sequência de DNA
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Mycobacterium
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Nocardia
Tipo de estudo:
Diagnostic_studies
Idioma:
En
Revista:
J Clin Pathol
Ano de publicação:
2018
Tipo de documento:
Article