Evaluation of Peptide Nucleic Acid Probe-Based Fluorescence In Situ Hybridization for the Detection of Mycobacterium tuberculosis Complex and Nontuberculous Mycobacteria in Clinical Respiratory Specimens
2
조회
0
다운로드
0
인용
0
참고문헌
키워드
초록
Background: Tuberculosis is globally the most important cause of death from single pathogen. Rapid and accurate identification of mycobacteria is essential for the control of tuberculosis. We evaluated a fluorescence in situ hybridization (FISH) method using peptide nucleic acid (PNA) probes for the differentiation of Mycobacterium tuberculosis complex (MTB) and nontuberculous mycobacteria (NTM) in direct smears of sputum specimens. Methods: The cross-reactivity of MTB- and NTM-specific PNA probes was examined with reference strains of M. tuberculosis ATCC 13950, Mycobacterium kansasii ATCC 12479, Mycobacterium fortuitum ATCC 6841, several clinical isolates of mycobacteria (Mycobacterium abscessus, Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium gordonae and Mycobacterium chelonae), and 11 frequently isolated respiratory bacterial species other than mycobacteria. A series of 128 sputa (89 MTB culture positive, 29 NTM culture positive, and 10 under treatment culture negative) with grades of trace to 4+ were used to evaluate the performance of the method. Results: The MTB- and NTM-specific PNA probes showed specific reactions with the reference strains of MTB and M. kansasii and clinical isolates of mycobacteria except M. fortuitum ATCC 6841, and no cross-reactivity with other tested bacteria. The PNA probe-based FISH assay for detection of MTB had a sensitivity and specificity of 100%, respectively. The sensitivity and specificity of the NTM-specific PNA probe was 100%. The smear grades of the PNA FISH test were same as with those of the fluorescence AFB stain in 2+ or higher grade. Conclusion: Detection and differentiation based on PNA FISH is sensitive and accurate for detecting mycobacteria and for differentiating MTB from NTM in clinical sputum smears. (Ann Clin Microbiol 2015;18:-43)
Abstract
Background: Tuberculosis is globally the most important cause of death from single pathogen. Rapid and accurate identification of mycobacteria is essential for the control of tuberculosis. We evaluated a fluorescence in situ hybridization (FISH) method using peptide nucleic acid (PNA) probes for the differentiation of Mycobacterium tuberculosis complex (MTB) and nontuberculous mycobacteria (NTM) in direct smears of sputum specimens. Methods: The cross-reactivity of MTB- and NTM-specific PNA probes was examined with reference strains of M. tuberculosis ATCC 13950, Mycobacterium kansasii ATCC 12479, Mycobacterium fortuitum ATCC 6841, several clinical isolates of mycobacteria (Mycobacterium abscessus, Mycobacterium avium, Mycobacterium intracellulare, Mycobacterium gordonae and Mycobacterium chelonae), and 11 frequently isolated respiratory bacterial species other than mycobacteria. A series of 128 sputa (89 MTB culture positive, 29 NTM culture positive, and 10 under treatment culture negative) with grades of trace to 4+ were used to evaluate the performance of the method. Results: The MTB- and NTM-specific PNA probes showed specific reactions with the reference strains of MTB and M. kansasii and clinical isolates of mycobacteria except M. fortuitum ATCC 6841, and no cross-reactivity with other tested bacteria. The PNA probe-based FISH assay for detection of MTB had a sensitivity and specificity of 100%, respectively. The sensitivity and specificity of the NTM-specific PNA probe was 100%. The smear grades of the PNA FISH test were same as with those of the fluorescence AFB stain in 2+ or higher grade. Conclusion: Detection and differentiation based on PNA FISH is sensitive and accurate for detecting mycobacteria and for differentiating MTB from NTM in clinical sputum smears. (Ann Clin Microbiol 2015;18:-43)
참고문헌 (18)
- "WHO web sites on infectious diseases. Global Tuberculosis Report 2013" (2013)[URL]
- Griffith DE, "An official ATS", IDSA statement: diagnosis, treatment, and prevention of nontuberculous mycobacterial diseases, 2007, pp.175, 367~416
- Richter E, "Manual of Clinical Microbiology", pp.503~524 (2011), ASM Press
- Shinnick TM, "National plan for reliable tuberculosis laboratory services using a systems approach. Recommendations from CDC and the Association of Public Health Laboratories Task Force on Tuberculosis Laboratory Services", MMWR Recomm Rep, 54, pp.1~12 (2005)
- Fitzgerald DW, "Mandell, Douglas, and Bennett’s Principles and Practice of Infectious Disease", pp.3129~3163 (2010), Churchill Livingstone Elsevier
- Drobniewski FA, "Differentiation of Mycobacterium tuberculosis complex and nontuberculous mycobacterial liquid cultures by using peptide nucleic acid-fluorescence in situ hybridization probes", J Clin Microbiol, 38, pp.444~447 (2000)
- Padilla E, "Evaluation of a fluorescence hybridization assay using peptide nucleic acid probes for identification and differentiation of tuberculous and non-tuberculous mycobacteria in liquid cultures", Eur J Clin Microbiol Infect Dis, 19, pp.140~145 (2000)
- Stender H, "Fluorescence in situ hybridization assay using peptide nucleic acid probes for differentiation between tuberculous and nontuberculous mycobacterium species in smears of mycobacterium cultures", J Clin Microbiol, 37, pp.2760~2765 (1999)
- Stender H, "Direct detection and identification of Mycobacterium tuberculosis in smear-positive sputum samples by fluorescence in situ hybridization(FISH)using peptide nucleic acid(PNA)probes", Int J Tuberc Lung Dis, 3, pp.830~837 (1999)
- 최영진, "Evaluation of Peptide Nucleic Acid Probe-based Real-time PCR for Detection of Mycobacterium tuberculosis Complex and Nontuberculous Mycobacteria in Respiratory Specimens", Annals of Laboratory Medicine, 32(4), pp.257~263 (2012), 대한진단검사의학회[DOI]
- Rodriguez-Nuñez J, "Mycobacterium tuberculosis complex detected by modified fluorescent in situ hybridization in lymph nodes of clinical samples", J Infect Dev Ctries, 6, pp.58~66 (2012)
- Lefmann M, "Evaluation of peptide nucleic acid-fluorescence in situ hybridization for identification of clinically relevant mycobacteria in clinical specimens and tissue sections", J Clin Microbiol, 44, pp.3760~3767 (2006)
- Choi YJ, "Evaluation of peptide nucleic acid array for the detection of hepatitis B virus mutations associated with antiviral resistance", Arch Virol, 156, pp.1517~1524 (2011)
- Porcheddu A, "Peptide nucleic acids (PNAs), a chemical overview", Curr Med Chem, 12, pp.2561~2599 (2005)
- Egholm M, "Recognition of guanine and adenine in DNA by cytosine and thymine containing peptide nucleic acids(PNA)", J Am Chem Soc, 114, pp.9677~9678 (1992)
- "Diagnostic Standards and Classification of Tuberculosis in Adults and Children. This official statement of the American Thoracic Society and the Centers for Disease Control and Prevention was adopted by the ATS Board of Directors, July 1999. This statement was endorsed by the Council of the Infectious Disease Society of America, September 1999", Am J Respir Crit Care Med, 161, pp.1376~1395 (2000)
- Hongmanee P, "Evaluation of a fluorescence in situ hybridization assay for differentiation between tuberculous and nontuberculous Mycobacterium species in smears of Lowenstein-Jensen and Mycobacteria Growth Indicator Tube cultures using peptide nucleic acid probes", J Clin Microbiol, 39, pp.1032~1035 (2001)
- Perry-O'Keefe H, "Identification of indicator microorganisms using a standardized PNA FISH method", J Microbiol Methods, 47, pp.281~292 (2001)