Comparison of three phenotypic and deoxyribonucleic acid extraction methods for isolation and Identification of Nocardia spp


1 Department of Bacteriology and Virology, Medical School, Isfahan University of Medical Sciences, Isfahan, Iran

2 Department of Infectious Diseases, Infectious and Tropical Diseases center, Isfahan University of Medical Sciences, Isfahan, Iran


Background: The aerobic Actinomycetes are a large group of soil-indwelling bacteria that are distributed in world-wide. These Gram-positive bacteria are most commonly associated with opportunistic infections in both immunocompromised and immunocompetent hosts.
Materials and Methods: In this study, three phenotypic and deoxyribonucleic acid (DNA) extraction methods for isolation and identification of Nocardia genus were compared. Samples were taken in five different locations of Isfahan's suburb from hospitals area, parks, agricultural lands, gardens, arid lands with different soil temperature and pH.
Results: In this study, showed that slip-buried-method was better than two other phenotypic methods; 14 out of 70 soil samples (20%) were positive for Nocardia spp. DNA of positive samples were extracted with three techniques and DNA extraction by microwave technique was better than others. This technique was confirmed with observation of DNA bands on 1% agarose gel.
Conclusions: These bacteria are important in immune deficient patients such as cancer patients, transplant recipients, tuberculosis; acquired immunodeficiency syndrome etc., Their affluence is unsteady in different zones of the world. In this study, among the three phenotypic methods for the isolation of Nocardia slip-buried method was better than other methods. Among DNA extraction techniques, DNA extraction by microwave method would be selective method for DNA extraction of Nocardia spp. compared with others techniques.


1. Kachuei R, Emami M, Mirnejad R, Khoobdel M. Diversity and frequency of Nocardia spp. in the soil of Isfahan Province, Iran. Asian Pac J Trop Biomed 2012;2:474-8.  Back to cited text no. 1
2. Kämpfer P, Huber B, Buczolits S, Thummes K, Grün-Wollny I, Busse HJ. Nocardia acidivorans sp. nov., isolated from soil of the island of Stromboli. Int J Syst Evol Microbiol 2007;57:1183-7.  Back to cited text no. 2
3. Sabuncuoðlu H, Cibali Açikgo ZZ, Caydere M, Ustün H, Semih Keskil I. Nocardia farcinica brain abscess: A case report and review of the literature. Neurocirugia (Astur) 2004;15:600-3.  Back to cited text no. 3
4. Xing K, Qin S, Fei SM, Lin Q, Bian GK, Miao Q, et alNocardia endophytica sp. nov., an endophytic Actinomycete isolated from the oil-seed plant Jatropha curcas L. Int J Syst Evol Microbiol 2011;61:1854-8.  Back to cited text no. 4
5. Xu P, Li WJ, Tang SK, Jiang Y, Gao HY, Xu LH, et alNocardia lijiangensis sp. nov., a novel Actinomycete strain isolated from soil in China. Syst Appl Microbiol 2006;29:308-14.  Back to cited text no. 5
6. Saubolle MA, Sussland D. Nocardiosis: Review of clinical and laboratory experience. J Clin Microbiol 2003;41:4497-501.  Back to cited text no. 6
7. Aghamirian MR, Ghiasian SA. Isolation and characterization of medically important aerobic Actinomycetes in soil of Iran (2006-2007). Open Microbiol J 2009;3:53-7.  Back to cited text no. 7
8. Khan ZU, Neil L, Chandy R, Chugh TD, Al-Sayer H, Provost F, et alNocardia asteroides in the soil of Kuwait. Mycopathologia 1997;137:159-63.  Back to cited text no. 8
9. Singh M, Sandhu RS, Randhawa HS. Comparison of paraffin baiting and conventional culture techniques for isolation of Nocardia asteroides from sputum. J Clin Microbiol 1987;25:176-7.  Back to cited text no. 9
10. Baron EJ, Peterson LR, Finegold SM. Bailey and Scott′s Diagnostic Microbiology. St. Louis: Mosby; 1994.  Back to cited text no. 10
11. Winn WC, Allen SD, Janda WM, Koneman EW, Schreckenberger PC, Procop GW, et al. Koneman′s Color Atlas and Textbook of Diagnostic Microbiology. Philadelphia: Lippincott Williams and Wilkins; 2005.  Back to cited text no. 11
12. van Gelderen de Komaid A, Durán EL. The efficiency of 1 M NH4Cl and 2 M NaCl for the isolation of pathogenic Nocardia from soil. Mycopathologia 1989;108:117-23.  Back to cited text no. 12
13. McTaggart LR, Richardson SE, Witkowska M, Zhang Sx. Phylogeny and identification of Nocardia species on the basis of multilocus sequence analysis. J Clin Microbiol 2010;48:4525-33.  Back to cited text no. 13
14. Kageyama A, Yazawa K, Kudo T, Taniguchi H, Nishimura K, Mikami Y. First isolates of Nocardia abscessus from humans and soil in Japan. Nihon Ishinkin Gakkai Zasshi 2004;45:17-21.  Back to cited text no. 14
15. Rodríguez-Nava V, Couble A, Devulder G, Flandrois JP, Boiron P, Laurent F. Use of PCR-restriction enzyme pattern analysis and sequencing database for hsp65 gene-based identification of Nocardia species. J Clin Microbiol 2006;44:536-46.  Back to cited text no. 15
16. Sahathevan M, Harvey FA, Forbes G, O′Grady J, Gimson A, Bragman S, et al. Epidemiology, bacteriology and control of an outbreak of Nocardia asteroides infection on a liver unit. J Hosp Infect 1991;18 Suppl A: 473-80.  Back to cited text no. 16
17. Salgado A, Ramirez N, Sandoval E, Sandoval H. Fast method for DNA extraction in Nocardia and Saccharomonospora. J Mycol Med/J Med Mycol 2008;18:100-2.  Back to cited text no. 17
18. Ayyar S, Tendolkar U, Deodhar L. A comparison of three media for isolation of Nocardia species from clinical specimens. J Postgrad Med 1992;38:70-2.  Back to cited text no. 18
[PUBMED]  Medknow Journal  
19. Amaro A, Duarte E, Amado A, Ferronha H, Botelho A. Comparison of three DNA extraction methods for Mycobacterium bovisMycobacterium tuberculosis and Mycobacterium avium subsp. avium. Lett Appl Microbiol 2008;47:8-11.  Back to cited text no. 19
20. Stapleton F, Keay LJ, Sanfilippo PG, Katiyar S, Edwards KP, Naduvilath T. Relationship between climate, disease severity, and causative organism for contact lens-associated microbial keratitis in Australia. Am J Ophthalmol 2007;144:690-8.  Back to cited text no. 20
21. Vetlugina LA, Adiiatova ZhF, Khozhamuratova SSh, Rymzhanova ZA, Trenozhnikova LP, Kopytina MN. Isolation of Actinomycetales from the soil of Kazakhstan on selective media with antibiotics. Antibiot Khimioter 1990;35:3-5.  Back to cited text no. 21
22. Goel S, Kanta S. Prevalence of Nocardia species in the soil of Patiala area. Indian J Pathol Microbiol 1993;36:53-60.  Back to cited text no. 22
23. van Gelderen de Komaid A, Runco de Laborda R, Salim R. Natural occurrence of Nocardia in soil of Tucumán: Physiological characteristics. Mycopathologia 1987;99:15-9.  Back to cited text no. 23
24. Negroni R, López Daneri G, Arechavala A, Bianchi MH, Robles AM. Clinical and microbiological study of mycetomas at the Muñiz Hospital of Buenos Aires between 1989 and 2004. Rev Argent Microbiol 2006;38:13-8.  Back to cited text no. 24
25. Denguezli M, Kourda M, Ghariani N, Belajouza C, Mokni B, Chebil F, et al. Mycetomas in central Tunisia. Ann Dermatol Venereol 2003;130:515-8.  Back to cited text no. 25
26. Conville PS, Witebsky FG. Nocardia and other aerobic Actinomycetes. Topley and Wilson′s Microbiology and Microbial Infections. Hodder Amold; Washington, DC.:Distributed in the United States of America by ASM Press, 2005.  Back to cited text no. 26
27. Butler WR, Kilburn JO, Kubica GP. High-performance liquid chromatography analysis of mycolic acids as an aid in laboratory identification of Rhodococcus and Nocardia species. J Clin Microbiol 1987;25:2126-31.  Back to cited text no. 27
28. Kim BY. Biosystematics of the genus Dactylosporangium and some other filamentous actinomycetes. Thesis. School of Biology, Faculty of Science, Agriculture and Engineering, Newcastle University, Newcastle upon Tyne, United Kingdom, 2010.  Back to cited text no. 28
29. Naidoo D, Kumari S, Bux F. Characterization of Nocardia farcinica, a filamentous bacterium isolated from foaming activated sludge samples. Water Environ Res 2011;83:527-31.  Back to cited text no. 29
30. Lamm AS, Khare A, Conville P, Lau PC, Bergeron H, Rosazza JP. Nocardia iowensis sp. nov., an organism rich in biocatalytically important enzymes and nitric oxide synthase. Int J Syst Evol Microbiol 2009;59:2408-14.  Back to cited text no. 30
31. Reddy AK, Garg P, Kaur I. Spectrum and clinicomicrobiological profile of Nocardia keratitis caused by rare species of Nocardia identified by 16S rRNA gene sequencing. Eye (Lond) 2010;24:1259-62.  Back to cited text no. 31