16S rRNA and Secondary Gene Target Sequencing on Pure Cultures

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*Accredited by the Standards Council of Canada to Laboratory no. 594 - CAN-P-4E (ISO/IEC 17025)

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Reference Details

Description:

16S rRNA gene PCR*, sequencing, and phylogenetic analysis. Secondary gene target sequencing and phylogenetic analysis is used complementarily with 16S gene sequencing. The choice of specific secondary gene target is dependent on 16S gene sequencing results.

Test Category:
Sequencing
Pathogen:
Bacterial pathogens
Illnesses and Diseases:
  • Various
Specimen:

Pure cultures of isolate implicated in clinical illness or environmental isolates.

Collection Method:

Slants or plates of any suitable media and swabs in transport media are all acceptable. Samples submitted for isolation and/or identification of strict anaerobes must be submitted under oxygen-free conditions. Pre-reduced anaerobically sterilized (PRAS)/Cary and Blair transport medium or PRAS Cooked Meat broth are acceptable.

Specimen Processing, Storage and Shipping:

Store and ship samples at room temperature.

Transportation of Dangerous Goods:

Shipping of specimens shall be done by a TDG certified individual in accordance with TDG regulations. For additional information regarding classification of specimens for the purposes of shipping, consult either Part 2 Appendix 3 of the TDG Regulations or section 3.6.2 of the IATA Dangerous Goods Regulations as applicable.

For additional guidance on the transport of infectious substances in other languages, please click on the link below.

http://www.who.int/ihr/capacity-strengthening/infectious-substances/en/

 

Patient Criteria:

Clinical illness with symptoms suggestive of bacterial infection.

Accompanying Documentation:

Completed Special Bacteriology requisition form detailing all patient information and relevant clinical information. If possible, attach lab results that have already been done at local or provincial laboratories.

Comments:

All patient and strain history must be included. Enteric bacteria, Mycobacteria, common nosocomial agents (MRSA, VRE), Neisseria, Eukaryotes, and Risk Group 3 bacteria are NOT accepted by Special Bacteriology. Please note that multiple samples isolated from the same source and patient, and identified as the same organism by the sender, will need an explanation in order for testing to be done on all the samples. Acceptance and/or rejection of the additional samples is at the discretion of the Special Bacteriology lab.   Contact lab for further information.  Please refer to the Guide to Services for test services offered by the NML.  *The 16S rRNA PCR test is accredited to CAN-P-4E (ISO/IEC 17025:2005) standards.

Methods and Interpretation of Results:

16S gene and secondary gene target (if applicable) sequencing results are compared against publically-available databases (GenBank) and known type strains.

Turnaround Time:

15 calendar days for the 16S/secondary gene target final report. Report may be delayed in cases where staff or resources are limited, or for poor or slow growing organisms.  In cases where further testing is required, the final report may be delayed and status of request will be forwarded in a preliminary report. Final report is sent once all testing is complete.

Contact:
Phone: (204) 789-2137
Fax: (204) 784-7509
References:
  1. Conville PS, Zelazny AM, Witebsky FG. 2006. Analysis of secA1 gene sequences for identification of Nocardia species. J Clin Microbiol. 44(8):2760-6
  2. Fry, N. K., S. Warwick, N. A. Saunders, and T.M. Embley. 1991. The use of 16S ribosomal RNA analyses to investigate the phylogeny of the family Legionellaceae. J. Gen. Microbiol. 137:1215-1222.
  3. Khamis, A, Raoult, D, La Scola, B. 2004. rpoB Gene Sequencing for Identification of Corynebacterium Species. JCM 42: 3925-3931
  4. Kolbert CP, Rys PN, Hopkins M, Lynch DT, Germer JJ, O’Sullivan CE, et al. 16S ribosomal DNA sequence analysis for identification of bacteria in a clinical microbiology laboratory.  In: Persing DH, Tenover FD, Versalovic J, Tang Y-W, Unger ER, Relman DA, White TJ, eds. Molecular microbiology: diagnostic principles and practice. Washington, DC: ASM Press 2004. p 361–77.
  5. Ratcliff RM, Lanser JA, Manning PA, Heuzenroeder MW.  1998.  Sequence-based classification scheme for the genus Legionella targeting the mip gene.  J Clin Microbiol. 36(6):1560-7.
  6. Yamamoto, S, PJM Bouvet, and S Harayama.  1999.  Phylogenetic structures of the genus Acinetobacter based on gyrB sequences: comparison with the grouping by DNA-DNA hybridization.  International Journal of Systematic Bacteriology.  49: 87-95.
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