Molecular Identification and Conventional Susceptibility Testing of Iranian Clinical Mycobacterium fortuitum Isolates

Document Type: Original Article

Authors

1 Department of Microbiology, School of Medicine, Isfahan University of Medical Sciences, Isfahan, Iran

2 Research Center for Infectious Diseases & Tropical Medicine, alternatively, Department of Microbiology, School of Medicine and, Isfahan University of Medical Sciences, Isfahan, Iran

3 Department of Medical Microbiology, Tehran University of Medical Sciences, Tehran, Iran

4 Iranian Research Center for Infectious Diseases, Isfahan University of Medical Sciences, Isfahan, Iran

5 Laboratory expert; Iranian Research Center for Infectious Diseases, Isfahan University of Medical Sciences, Isfahan, Iran

Abstract

Objective(s)
Rapidly growing mycobacteria (RGM) are capable of producing diseases in humans. Since mycobacteria vary in their susceptibility, precise identification is critical for adoption of correct drug therapy. The main aim of this study was molecular identification and evaluation of antimicrobial susceptibility pattern of Iranian clinically isolated Myocbacterium fortuitum.
Materials and Methods
A total of 72 presumptively identified isolates of clinical atypical mycobacteria collected by Isfahan Research Center for Infectious Diseases & Tropical Medicine during 2006-2008 were included in the current study. A combination of conventional and molecular tests was applied to identify the isolates. Molecular methods including genus and group specific PCR and PCR-Restriction Algorithm (PRA) based on hsp65 gene were applied to achieve exact identification of mycobacterial strains. Antimicrobial susceptibility testing onM. fortuitum isolates was performed by in-house prepared broth microdilution test..
Results
Out of 72 collected atypical mycobacteria isolates, we identified 25 strains of M. fortuitum. All strains had the specific molecular markers of mycobacterial identity and similar species specific PRA pattern of the international type strain of M. fortuitum. Drug susceptibility testing showed that the M. fortuitum isolates are sensitive to amikacin, sulfamethoxazole and ciprofloxacin (100%), imipenem (92%), clarithromycin (76%), cefoxitin (56%) and doxycycline (16%).
Conclusion
Molecular identification of atypical mycobacteria based on PRA is a reliable and rapid approach which can identify mycobacterial strains to the species level. Our study showed that M. fortuitum plays a significant role in pulmonary and extrapulmonary infection in patients and should be given proper considerations when clinical samples are processed.

Keywords


1.Yang SC, Hsueh PR, Lai HC, Teng LJ, Huang LM, Chen JM, et al. High prevalence of antimicrobial resistance in rapidly growing mycobacteria in Taiwan. Antimicrob Agents Chemother 2003; 47:1958-1962.

2.Woods GL. Susceptibility testing for mycobacteria. Clin Infect Dis 2000; 31:1209-1215.

3.Woods GL, Bergmann JS, Witebsky FG, Fahle GA, Boulet B, Plaunt M, et al. Multisite reproducibility of E-test for susceptibility testing of Mycobacterium abscessus, M. chelonae, and M. fortuitum. J Clin Microbiol 2000; 38:656-661.

4.Kim H, Kim SH, Shim TS, Kim MN, Bai GH, Park YG, et al. PCR restriction fragment length polymorphism analysis (PRA)-algorithm targeting 644 bp Heat Shock Protein 65 (hsp65) gene for differentiation of Mycobacterium spp. J Microbiol Methods 2005; 62:199-209.

5.Martin-Casabona N, Bahrmand AR, Bennedsen J, Thomsen VO, Curcio M, Fauville-Dufaux M, et al. Non-tuberculous mycobacteria: patterns of isolation. A multi-country retrospective survey. Int J Tuberc Lung Dis 2004; 8:1186-1193.

6.Griffith DE, Aksamit T, Brown-Elliott BA, Catanzaro A, Daley C, Gordin F, et al. ATS mycobacterial diseases subcommittee; American Thoracic Society; Infectious Disease Society of America. An official ATS/IDSA statement: diagnosis, treatment, and prevention of nontuberculous mycobacterial diseases. Am J Respir Crit Care Med 2007; 175:365-416.

7.Brown-Elliott BA, Wallace RJJ. Clinical and taxonomic status of pathogenic nonpigmented or late-pigmenting rapidly growing mycobacteria. Clin Microbiol Rev 2002; 15:716-746.

8.Kent PT, Kubica GP. Public health mycobacteriology: a guide for the level III laboratory. US Department of Health and Human Services publication (CDC).no. 86-8230. Centers for Disease Control, Atlanta, Ga. US: Public Health Service publication; 1995.

9.Shojaei H, Magee JG, Freeman R, Yates M, Horadagoda NU, Goodfellow M. Mycobacterium elephantis sp. nov, a rapidly growing non-chromogenic Mycobacterium isolated from an elephant. Int J Syst Evol Microbiol 2000; 5:1817-1820.

10.Khan IU, Yadav JS. Development of a single-tube, cell lysis-based, genus-specific PCR method for rapid identification of mycobacteria: optimization of cell lysis, PCR primers and conditions, and restriction pattern analysis. J Clin Microbiol 2004; 42:453-457.

11.Sahm DF, Washington II JA. Antibacterial susceptibility tests: dilution methods. In: Balows A. editor. Manual of Clinical Microbiology. 5thed. Washington, DC: American Society for Microbiology; 1991.p.1105-1116.

12.NCCLS. Susceptibility testing of Mycobacteria, Nocardia, and other aerobic actinomycetes. Approved standard M24-A, Vol. 23. NCCLS, Villanova, Pa. 2003.

13.American Thoracic Society. Diagnosis and treatment of disease caused by nontuberculous mycobacteria. Am J Respir Crit Care Med 1997; 156:S1-S25.

14.Lee SM, Kim JM, Jeong J, Park YK, Bai GH, Lee EY, et al. Evaluation of the broth microdilution method using 2,3-Diphenyl-5-thienyl-(2)-tetrazolium chloride for rapidly growing mycobacteria susceptibility testing. J Korean Med Sci 2007; 22:784-790.

15.Foo H, Hal S van, Jelfs P, Gilbert GL. Antimicrobial resistance in non-tuberculous mycobacteria in New South Wales, 2002-2008. Int J Antimicrob Agents 2009; 34:182-184.

16.Wallace RJ, Dalovisio JR, Pankey GA. Disk diffusion testing of susceptibility of Mycobacterium fortuitum and Mycobacterium chelonei to antibacterial agents. Antimicrob Agents Chemother 1979; 16:611-614.