Chapter 4 : Laboratory Diagnosis and Susceptibility Testing

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This chapter talks about laboratory diagnosis and susceptibility testing of complex. Clinical microbiology laboratories currently have a number of methods available that provide an accurate and rapid laboratory diagnosis of tuberculosis. Mycobacterial culture of stool specimens may be of value in intestinal tuberculosis cases, but these are rare. This type of culture has been requested to detect - infections in patients with AIDS; however, given that intestinal involvement with - is thought to be a component of disseminated disease, a blood culture for mycobacteria is the specimen of choice in this setting. When colonies resembling mycobacteria are observed, an acid-fast smear and subculture for identification and susceptibility testing should be made. Nucleic acid probe testing or another comparable molecular method of identification can be performed on colonies as soon as they appear, and the definitive identification can be made if results are consistent with complex (, , BCG, , , and “”). Cultures are essential to exclude the possibility of mixed infections, which exist although are rare; in some instances, for further characterization or identification (most amplification tests give results only at the complex level); and, most importantly, for complete antimicrobial susceptibility testing. Susceptibility tests should be performed on all isolates of complex recovered from previously untreated patients and also on isolates from patients on therapy who have positive acid- fast smears or cultures after 2 months of treatment.

Citation: Procop G, Roberts G. 2011. Laboratory Diagnosis and Susceptibility Testing, p 66-74. In Schlossberg D (ed), Tuberculosis and Nontuberculous Mycobacterial Infections, Sixth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555817138.ch4

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1. Abdelaal, A.,, H. A. El-Ghaffar,, M. H. Zaghloul,, N. El Mashad,, E. Badran,, and A. Fathy. 2009. Genotypic detection of rifampicin and isoniazid resistant Mycobacterium tuberculosis strains by DNA sequencing: a randomized trial. Ann. Clin. Microbiol. Antimicrob. 8:4.
2. Alcaide, F.,, M. A. Benitez,, J. M. Escriba,, and R. Martin. 2000. Evaluation of the BACTEC MGIT 960 and the MB/BacT systems for recovery of mycobacteria from clinical specimens and for species identification by DNA AccuProbe. J. Clin. Microbiol. 38:398401.
3. Angeby, K. A.,, J. Werngren,, J. C. Toro,, G. Hedstrom,, B. Petrini,, and S. E. Hoffner. 2003. Evaluation of the BacT/ALERT 3D system for recovery and drug susceptibility testing of Mycobacterium tuberculosis. Clin. Microbiol. Infect. 9:11481152.
4. Bemer, P.,, T. Bodmer,, J. Munzinger,, M. Perrin,, V. Vincent,, and H. Drugeon. 2004. Multicenter evaluation of the MB/BACT system for susceptibility testing of Mycobacterium tuberculosis. J. Clin. Microbiol. 42:10301034.
5. Bennedsen, J.,, V. O. Thomsen,, G. E. Pfyffer,, G. Funke,, K. Feld-mann,, A. Beneke,, P. A. Jenkins,, M. Hegginbothom,, A. Fahr,, M. Hengstler,, G. Cleator,, P. Klapper,, and E. G. Wilkins. 1996. Utility of PCR in diagnosing pulmonary tuberculosis. J. Clin. Microbiol. 34:14071411.
6. Bergmann, J. S.,, and G. L. Woods. 1996. Clinical evaluation of the Roche AMPLICOR PCR Mycobacterium tuberculosis test for detection of M. tuberculosis in respiratory specimens. J. Clin. Microbiol. 34:10831085.
7. Bird, B. R.,, M. M. Denniston,, R. E. Huebner,, and R. C. Good. 1996. Changing practices in mycobacteriology: a follow-up survey of state and territorial public health laboratories. J. Clin. Microbiol. 34:554559.
8. Blanchard, J. S. 1996. Molecular mechanisms of drug resistance in Mycobacterium tuberculosis. Annu. Rev. Biochem. 65:215239.
9. Butler, W. R.,, S. P. O’Connor,, M. A. Yakrus,, and W. M. Gross. 1994. Cross-reactivity of genetic probe for detection of Mycobacterium tuberculosis with newly described species Mycobacterium celatum. J. Clin. Microbiol. 32:536538.
10. Cartuyvels, R.,, C. De Ridder,, S. Jonckheere,, L. Verbist,, and J. Van Eldere. 1996. Prospective clinical evaluation of Amplicor Mycobacterium tuberculosis PCR test as a screening method in a low-prevalence population. J. Clin. Microbiol. 34:20012003.
11. Catanzaro, A.,, B. Davidson,, P. Fujiwara,, M. Goldberger,, F. Gordin,, M. Salfinger,, J. Sbarbaro,, N. Schluger,, M. Sierra,, and G. Woods. 1997. Rapid diagnostic tests for tuberculosis. Am. J. Respir. Crit. Care Med. 155:18041814.
12. Centers for Disease Control and Prevention. 2009. Updated guidelines for the use of nucleic acid amplification tests in the diagnosis of tuberculosis. MMWR Morb. Mortal. Wkly. Rep. 58:710.
13. Christiansen, D. C.,, G. D. Roberts,, and R. Patel. 2004. Mycobacterium celatum, an emerging pathogen and cause of false positive amplified Mycobacterium tuberculosis direct test. Diagn. Microbiol. Infect. Dis. 49:1924.
14. Cowan, L. S.,, L. Diem,, M. C. Brake,, and J. T. Crawford. 2004. Transfer of a Mycobacterium tuberculosis genotyping method, Spoligotyping, from a reverse line-blot hybridization, membrane-based assay to the Luminex multianalyte profiling system. J. Clin. Microbiol. 42:474477.
15. Cruciani, M.,, C. Scarparo,, M. Malena,, O. Bosco,, G. Serpelloni,, and C. Mengoli. 2004. Meta-analysis of BACTEC MGIT 960 and BACTEC 460 TB, with or without solid media, for detection of mycobacteria. J. Clin. Microbiol. 42:23212325.
16. Drosten, C.,, M. Panning,, and S. Kramme. 2003. Detection of Mycobacterium tuberculosis by real-time PCR using panmycobacterial primers and a pair of fluorescence resonance energy transfer probes specific for the M. tuberculosis complex. Clin. Chem. 49:16591661.
17. Ellner, P. D.,, T. E. Kiehn,, R. Cammarata,, and M. Hosmer. 1988. Rapid detection and identification of pathogenic mycobacteria by combining radiometric and nucleic acid probe methods. J. Clin. Microbiol. 26:13491352.
18. Evans, K. D.,, A. S. Nakasone,, P. A. Sutherland,, L. M. de la Maza,, and E. M. Peterson. 1992. Identification of Mycobacterium tuberculosis and Mycobacterium avium-M. intracellulare directly from primary BACTEC cultures by using acridinium-ester-labeled DNA probes. J. Clin. Microbiol. 30:24272431.
19. Ford, E. G.,, S. J. Snead,, J. Todd,, and N. G. Warren. 1993. Strains of Mycobacterium terrae complex which react with DNA probes for M. tuberculosis complex. J. Clin. Microbiol. 31:28052806.
20. Gamboa, F.,, J. M. Manterola,, B. Vinado,, L. Matas,, M. Gimenez,, J. Lonca,, J. R. Manzano,, C. Rodrigo,, P. J. Cardona,, E. Padilla,, J. Dominguez,, and V. Ausina. 1997. Direct detection of Mycobacterium tuberculosis complex in nonrespiratory specimens by Gen-Probe Amplified Mycobacterium Tuberculosis Direct Test. J. Clin. Microbiol. 35:307310.
21. Garcia de Viedma, D. 2003. Rapid detection of resistance in Mycobacterium tuberculosis: a review discussing molecular approaches. Clin. Microbiol. Infect. 9:349359.
22. Gori, A.,, A. Bandera,, G. Marchetti,, A. Degli Esposti,, L. Catozzi,, G. P. Nardi,, L. Gazzola,, G. Ferrario,, J. D. van Embden,, D. van Soolingen,, M. Moroni,, and F. Franzetti. 2005. Spoligo-typing and Mycobacterium tuberculosis. Emerg. Infect. Dis. 11:12421248.
23. Gori, A.,, A. D. Esposti,, A. Bandera,, M. Mezzetti,, C. Sola,, G. Marchetti,, G. Ferrario,, F. Salerno,, M. Goyal,, R. Diaz,, L. Gazzola,, L. Codecasa,, V. Penati,, N. Rastogi,, M. Moroni,, and F. Franzetti. 2005. Comparison between spoligotyping and IS6110 restriction fragment length polymorphisms in molecular genotyping analysis of Mycobacterium tuberculosis strains. Mol. Cell. Probes 19:236244.
24. Goto, M.,, S. Oka,, K. Okuzumi,, S. Kimura,, and K. Shimada. 1991. Evaluation of acridinium-ester-labeled DNA probes for identification of Mycobacterium tuberculosis and Mycobacterium avium-Mycobacterium intracellulare complex in culture. J. Clin. Microbiol. 29:24732476.
25. Hall, L.,, K. A. Doerr,, S. L. Wohlfiel,, and G. D. Roberts. 2003. Evaluation of the MicroSeq system for identification of mycobacteria by 16S ribosomal DNA sequencing and its integration into a routine clinical mycobacteriology laboratory. J. Clin. Microbiol. 41:14471453.
26. Heifets, L. B. 1996. Clinical mycobacteriology. Drug susceptibility testing. Clin. Lab. Med. 16:641656.
27. Herold, C. D.,, R. L. Fitzgerald,, and D. A. Herold. 1996. Current techniques in mycobacterial detection and speciation. Crit. Rev. Clin. Lab. Sci. 33:83138.
28. Huang, T. S.,, Y. C. Liu,, H. H. Lin,, W. K. Huang,, and D. L. Cheng. 1996. Comparison of the Roche AMPLICOR MYCOBACTERIUM assay and Digene SHARP Signal System with in-house PCR and culture for detection of Mycobacterium tuberculosis in respiratory specimens. J. Clin. Microbiol. 34:30923096.
29. Jonas, V.,, and M. Longiaru. 1997. Detection of Mycobacterium tuberculosis by molecular methods. Clin. Lab. Med. 17:119128.
30. Kontos, F.,, M. Maniati,, C. Costopoulos,, Z. Gitti,, S. Nicolaou,, E. Petinaki,, S. Anagnostou,, I. Tselentis,, and A. N. Maniatis. 2004. Evaluation of the fully automated Bactec MGIT 960 system for the susceptibility testing of Mycobacterium tuberculosis to first-line drugs: a multicenter study. J. Microbiol. Methods 56:291294.
31. Kraus, G.,, T. Cleary,, N. Miller,, R. Seivright,, A. K. Young,, G. Spruill,, and H. J. Hnatyszyn. 2001. Rapid and specific detection of the Mycobacterium tuberculosis complex using fluorogenic probes and real-time PCR. Mol. Cell. Probes 15:375383.
32. Kubica, G. P.,, and G. L. Pool. 1960. Studies on the catalase activity of acid-fast bacilli. I. An attempt to subgroup these organisms on the basis of their catalase activities at different temperatures and pH. Am. Rev. Respir. Dis. 81:387391.
33. Laverdiere, M.,, L. Poirier,, K. Weiss,, C. Beliveau,, L. Bedard,, and D. Desnoyers. 2000. Comparative evaluation of the MB/BacT and BACTEC 460 TB systems for the detection of mycobacteria from clinical specimens: clinical relevance of higher recovery rates from broth-based detection systems. Diagn. Microbiol. Infect. Dis. 36:15.
34. Manterola, J. M.,, F. Gamboa,, E. Padilla,, J. Lonca,, L. Matas,, A. Hernandez,, M. Gimenez,, P. J. Cardona,, B. Vinado,, and V. Ausina. 1998. Comparison of a nonradiometric system with Bactec 12B and culture on egg-based media for recovery of mycobacteria from clinical specimens. Eur. J. Clin. Microbiol. Infect. Dis. 17:773777.
35. Metchock, B.,, and L. Diem. 1995. Algorithm for use of nucleic acid probes for identifying Mycobacterium tuberculosis from BACTEC 12B bottles. J. Clin. Microbiol. 33:19341937.
36. Moore, D. F.,, J. I. Curry,, C. A. Knott,, and V. Jonas. 1996. Amplification of rRNA for assessment of treatment response of pulmonary tuberculosis patients during antimicrobial therapy. J. Clin. Microbiol. 34:17451749.
37. Musser, J. M. 1995. Antimicrobial agent resistance in mycobacteria: molecular genetic insights. Clin. Microbiol. Rev. 8:496514.
38. Noordhoek, G. T.,, J. D. van Embden,, and A. H. Kolk. 1996. Reliability of nucleic acid amplification for detection of Mycobacterium tuberculosis: an international collaborative quality control study among 30 laboratories. J. Clin. Microbiol. 34:25222525.
39. Pfyffer, G. E.,, P. Kissling,, E. M. Jahn,, H. M. Welscher,, M. Salfinger,, and R. Weber. 1996. Diagnostic performance of amplified Mycobacterium tuberculosis direct test with cerebrospinal fluid, other nonrespiratory, and respiratory specimens. J. Clin. Microbiol. 34:834841.
40. Piersimoni, C.,, A. Callegaro,, D. Nista,, S. Bornigia,, F. De Conti,, G. Santini,, and G. De Sio. 1997. Comparative evaluation of two commercial amplification assays for direct detection of Mycobacterium tuberculosis complex in respiratory specimens. J. Clin. Microbiol. 35:193196.
41. Piersimoni, C.,, and C. Scarparo. 2003. Relevance of commercial amplification methods for direct detection of Mycobacterium tuberculosis complex in clinical samples. J. Clin. Microbiol. 41:53555365.
42. Piersimoni, C.,, C. Scarparo,, A. Callegaro,, C. P. Tosi,, D. Nista,, S. Bornigia,, M. Scagnelli,, A. Rigon,, G. Ruggiero,, and A. Goglio. 2001. Comparison of MB/BacT ALERT 3D system with radiometric BACTEC system and Löwenstein-Jensen medium for recovery and identification of mycobacteria from clinical specimens: a multicenter study. J. Clin. Microbiol. 39:651657.
43. Pomputius, W. F., III,, J. Rost,, P. H. Dennehy,, and E. J. Carter. 1997. Standardization of gastric aspirate technique improves yield in the diagnosis of tuberculosis in children. Pediatr. Infect. Dis. J. 16:222226.
44. Procop, G. W. 2007. Molecular diagnostics for the detection and characterization of microbial pathogens. Clin. Infect. Dis. 45(Suppl. 2):S99S111.
45. Rickman, T. W.,, and N. P. Moyer. 1980. Increased sensitivity of acid-fast smears. J. Clin. Microbiol. 11:618620.
46. Roberts, G. D.,, N. L. Goodman,, L. Heifets,, H. W. Larsh,, T. H. Lindner,, J. K. McClatchy,, M. R. McGinnis,, S. H. Siddiqi,, and P. Wright. 1983. Evaluation of the BACTEC radiometric method for recovery of mycobacteria and drug susceptibility testing of Mycobacterium tuberculosis from acid-fast smear-positive specimens. J. Clin. Microbiol. 18:689696.
47. Sandin, R. L. 1996. Polymerase chain reaction and other amplification techniques in mycobacteriology. Clin. Lab. Med. 16:617639.
48. Scarparo, C.,, P. Piccoli,, A. Rigon,, G. Ruggiero,, P. Ricordi,, and C. Piersimoni. 2002. Evaluation of the BACTEC MGIT 960 in comparison with BACTEC 460 TB for detection and recovery of mycobacteria from clinical specimens. Diagn. Microbiol. Infect. Dis. 44:157161.
49. Shrestha, N. K.,, M. J. Tuohy,, G. S. Hall,, U. Reischl,, S. M. Gordon,, and G. W. Procop. 2003. Detection and differentiation of Mycobacterium tuberculosis and nontuberculous mycobacterial isolates by real-time PCR. J. Clin. Microbiol. 41:51215126.
50. Telenti, M.,, J. F. de Quiros,, M. Alvarez,, M. J. Santos Rionda,, and M. C. Mendoza. 1994. The diagnostic usefulness of a DNA probe for Mycobacterium tuberculosis complex (Gen-Probe) in Bactec cultures versus other diagnostic methods. Infection 22:1823.
51. Tjhie, J. H.,, A. F. van Belle,, M. Dessens-Kroon,, and D. van Soolingen. 2001. Misidentification and diagnostic delay caused by a false-positive amplified Mycobacterium tuberculosis direct test in an immunocompetent patient with a Mycobacterium celatum infection. J. Clin. Microbiol. 39:23112312.
52. Torres, M. J.,, A. Criado,, M. Ruiz,, A. C. Llanos,, J. C. Palomares,, and J. Aznar. 2003. Improved real-time PCR for rapid detection of rifampin and isoniazid resistance in Mycobacterium tuberculosis clinical isolates. Diagn. Microbiol. Infect. Dis. 45:207212.
53. Tortoli, E.,, M. Benedetti,, A. Fontanelli,, and M. T. Simonetti. 2002. Evaluation of automated BACTEC MGIT 960 system for testing susceptibility of Mycobacterium tuberculosis to four major antituberculous drugs: comparison with the radio-metric BACTEC 460TB method and the agar plate method of proportion. J. Clin. Microbiol. 40:607610.
54. Tortoli, E.,, P. Cichero,, C. Piersimoni,, M. T. Simonetti,, G. Gesu,, and D. Nista. 1999. Use of BACTEC MGIT 960 for recovery of mycobacteria from clinical specimens: multicenter study. J. Clin. Microbiol. 37:35783582.
55. Tuohy, M. J.,, G. S. Hall,, M. Sholtis,, and G. W. Procop. 2005. Pyrosequencing as a tool for the identification of common isolates of Mycobacterium sp. Diagn. Microbiol. Infect. Dis. 51:245250.
56. Turenne, C. Y.,, L. Tschetter,, J. Wolfe,, and A. Kabani. 2001. Necessity of quality-controlled 16S rRNA gene sequence databases: identifying nontuberculous Mycobacterium species. J. Clin. Microbiol. 39:36373648.
57. Wilson, M. L.,, B. L. Stone,, M. V. Hildred,, and R. R. Reves. 1995. Comparison of recovery rates for mycobacteria from BACTEC 12B vials, Middlebrook 7H11-selective 7H11 bi-plates, and Lowenstein-Jensen slants in a public health mycobacteriology laboratory. J. Clin. Microbiol. 33:25162518.
58. Woods, G.,, B. Brown-Elliot,, E. Desmond,, G. Hall,, L. Heifets,, G. Pfyffer,, J. Ridderhof,, R. J. Wallace,, N. Warren,, and F. Witebsky. 2010. Susceptibility Testing of Mycobacteria, Nocardiae, and Other Aerobic Actinomycetes. Approved standard M-24A, 2nd ed. Clinical and Laboratory Standards Institute, Wayne, PA.

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