Abstract
Broad-range PCR has become a valuable tool for the identification of microorganisms in the clinical laboratory over the last years. It was primarily used to identify slow-growing and fastidious microorganisms with poor biochemical activity. Nowadays, it is also used to identify microorganisms directly from clinical samples such as blood or punctuates from primarily sterile body sites. In these specimens, the usage of broad-range PCR is challenging regarding contamination and standardization. To overcome these problems, a new test system, the SepsiTest™, was introduced recently employing broad-range PCR for the identification of microorganisms in septic patients. In this chapter, the test system is described and the equipment necessary listed.
This is a preview of subscription content, log in via an institution.
Buying options
Tax calculation will be finalised at checkout
Purchases are for personal use only
Learn about institutional subscriptionsReferences
Clarridge JE 3rd (2004) Impact of 16S rRNA gene sequence analysis for identification of bacteria on clinical microbiology and infectious diseases. Clin Microbiol Rev 17:840–862, table of contents
Sontakke S, Cadenas MB, Maggi RG, Diniz PP, Breitschwerdt EB (2009) Use of broad range16S rDNA PCR in clinical microbiology. J Microbiol Methods 76:217–225
Atkins SD, Clark IM (2004) Fungal molecular diagnostics: a mini review. J Appl Genet 45:3–15
Klappenbach JA, Saxman PR, Cole JR, Schmidt TM (2001) rrndb: the ribosomal RNA operon copy number database. Nucleic Acids Res 29:181–184
CLSI, Clinical and Laboratory Standards Institute (2008) Interpretive criteria for identification of bacteria and fungi by DNA target sequencing; approved guideline. CLSI dokument MM18-A (ISBN 1-56238-664-6)
Wellinghausen N, Kochem AJ, Disque C, Muhl H, Gebert S, Winter J, Matten J, Sakka SG (2009) Diagnosis of bacteremia in whole-blood samples by use of a commercial universal 16S rRNA gene-based PCR and sequence analysis. J Clin Microbiol 47:2759–2765
Leitner E, Kessler HH, Spindelboeck W, Hoenigl M, Putz-Bankuti C, Stadlbauer-Kollner V, Krause R, Grisold AJ, Feierl G, Stauber RE (2013) Comparison of two molecular assays with conventional blood culture for diagnosis of sepsis. J Microbiol Methods 92:253–255
Wiesinger-Mayr H, Jordana-Lluch E, Martro E, Schoenthaler S, Noehammer C (2011) Establishment of a semi-automated pathogen DNA isolation from whole blood and comparison with commercially available kits. J Microbiol Methods 85:206–213
Sleigh J, Cursons R, La Pine M (2001) Detection of bacteraemia in critically ill patients using 16S rDNA polymerase chain reaction and DNA sequencing. Intensive Care Med 27:1269–1273
Fenollar F, Raoult D (2007) Molecular diagnosis of bloodstream infections caused by non-cultivable bacteria. Int J Antimicrob Agents 30(Suppl 1):S7–S15
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Science+Business Media New York
About this protocol
Cite this protocol
Leitner, E., Kessler, H.H. (2015). Broad-Range PCR for the Identification of Bacterial and Fungal Pathogens from Blood: A Sequencing Approach. In: Mancini, N. (eds) Sepsis. Methods in Molecular Biology, vol 1237. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1776-1_13
Download citation
DOI: https://doi.org/10.1007/978-1-4939-1776-1_13
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-1775-4
Online ISBN: 978-1-4939-1776-1
eBook Packages: Springer Protocols