Chapter 6.4 : Molecular Identification

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Currently, accurate identification of aerobic actinomycetes to the species level requires the use of molecular methods, specifically, gene sequencing and analysis. 16S rRNA gene sequence analysis can provide genus-level and, for most genera, species-level identification of clinically relevant aerobic actinomycetes ( ). Based on recommendations from Tindall et al. ( ), nearly complete high-quality gene sequences are necessary to maintain accurate genus/species assignment based on sequence similarity values as defined by CLSI ( ). Speciation of isolates suspected of being aerobic actinomycetes requires a 16S rRNA gene sequence with a minimum length of 1,440 bp. A BLAST analysis of a high-quality gene sequence against a DNA sequence database such as GenBank (www.ncbi.nlm.nih.gov/Genbank/) will allow for genus or species assignment based on sequence similarity results in comparison with a reference strain.

Citation: Leber A. 2016. Molecular Identification, p 6.4.1-6.4.3. In Clinical Microbiology Procedures Handbook, Fourth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818814.ch6.4
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1. CLSI. 2008. Interpretive Criteria for Identification of Bacteria and Fungi by DNA Target Sequencing: Approved Guideline. Approved standard MM18-A. CLSI, Wayne, PA.
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5. Conville PS, Witebsky FG. 2005. Multiple copies if the 16S rRNA gene in Nocardia nova isolates and implications for sequence-based identification procedures. J Clin Microbiol 43: 28812885.
6. Weisburg WG, Barnes SM, Pelletier DA, Lane DJ. 1991. 16S ribosomal DNA amplification for phylogenetic study. J Bacteriol 173: 697703.
7. Morey RE, Galloway RL, Bragg SL, Steigerwalt AG, Mayer LW, Levett PN. 2006. Species-specific identification of Leptospiraceae of 16S rRNA gene sequencing. J Clin Microbiol 44: 35103516.
8. Heuer H, Kresk M, Baker P, Smalla K, Wellington EM. 1997. Analysis of actinomycete communities by specific amplification of genes encoding 16S rRNA and gel-electrophoretic separation in denaturing gradients. Appl Environ Microbiol 63: 32333241.
9. Wuyts J, Van de Peer Y, Winklemans T, De Wachter R. 2002. The European database on small subunit ribosomal RNA. Nucleic Acids Res 30: 183185.
1. Verroken A, Janssens M, Berhin C, Bogaerts P, Huang TD, Wauters G, Glupczynski Y. 2010. Evaluation of matrix-assisted laser desorption ionization-time of flight mass spectrometry for identification of nocardia species. J Clin Microbiol 48: 40154021.
2. Clark AE, Kaleta EJ, Arora A, Wolk DM. 2013. Matrix-assisted laser desorption ionization–time of flight mass spectrometry: a fundamental shift in the routine practice of clinical microbiology. Clin Microbiol Rev 26: 547603.
3. Farfour E, Leto J, Barritault M, Barberis C, Meyer J, Dauphin B, Le Guern A-S, Leflèche A, Badell E, Guiso N, Leclercq A, Le Monnier A, Lecuit M, Rodriguez-Nava V, Bergeron E, Raymond J, Vimont S, Bille E, Carbonnelle E, Guet-Revillet H, Lécuyer H, Beretti J-L, Vay C, Berche P, Ferroni A, Nassif X, Join-Lambert O. 2012. Evaluation of the Andromas matrix-assisted laser desorption ionization–time of flight mass spectrometry system for identification of aerobically growing gram-positive bacilli. J Clin Microbiol 50: 27022707.


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Table 6.4–1

16S rRNA gene primers ( ) for amplification and sequencing aerobic actinomycetes

Citation: Leber A. 2016. Molecular Identification, p 6.4.1-6.4.3. In Clinical Microbiology Procedures Handbook, Fourth Edition. ASM Press, Washington, DC. doi: 10.1128/9781555818814.ch6.4

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