R Trotha1, T Hanck, W König, B König. 1. Institute for Medical Microbiology, Otto-von-Guericke University, Magdeburg, Germany.
Abstract
BACKGROUND: Rapid and reliable identification of microorganisms is a prerequisite for the diagnosis and subsequent treatment of infectious diseases. The identification of pathogenic bacteria is traditionally based on their isolation from clinical samples and propagation on culture medium in the routine laboratory. However, despite clinical signs of infection, culture of the pathogenic agent often fails. This may be due to a low number of microorganisms, prior antibiotic treatment, nonculturable microorganisms or specific culture requirements for presently unknown pathogens. Amplification and sequencing of the entire prokaryotic 16S-rRNA is time consuming, labor intensive and expensive. MATERIALS AND METHODS: We describe here a procedure for the identification of a wide range of known and unknown clinically relevant microorganisms by sequencing a small, but highly informative region of the prokaryotic 16S-rRNA gene. This rapid ribosequencing method was evaluated with various reference strains and with clinical samples including eye anterior chamber fluid, cerebrospinal fluid (CSF) and blood cultures. RESULTS: All sequences obtained from the reference strains corresponded to the sequences in databases. We correlated severe eye infection with the isolation of Pseudomonas putida, neurological disorder with Tropheryma whippelii and disseminated visceral abscesses in a child with Blastobacter denitrificans. CONCLUSION: We consider the rapid ribosequencing method as a promising new tool for the analysis of infectious agents in primarily sterile body fluids where conventional culturing of microorganisms fails.
BACKGROUND: Rapid and reliable identification of microorganisms is a prerequisite for the diagnosis and subsequent treatment of infectious diseases. The identification of pathogenic bacteria is traditionally based on their isolation from clinical samples and propagation on culture medium in the routine laboratory. However, despite clinical signs of infection, culture of the pathogenic agent often fails. This may be due to a low number of microorganisms, prior antibiotic treatment, nonculturable microorganisms or specific culture requirements for presently unknown pathogens. Amplification and sequencing of the entire prokaryotic 16S-rRNA is time consuming, labor intensive and expensive. MATERIALS AND METHODS: We describe here a procedure for the identification of a wide range of known and unknown clinically relevant microorganisms by sequencing a small, but highly informative region of the prokaryotic 16S-rRNA gene. This rapid ribosequencing method was evaluated with various reference strains and with clinical samples including eye anterior chamber fluid, cerebrospinal fluid (CSF) and blood cultures. RESULTS: All sequences obtained from the reference strains corresponded to the sequences in databases. We correlated severe eye infection with the isolation of Pseudomonas putida, neurological disorder with Tropheryma whippelii and disseminated visceral abscesses in a child with Blastobacter denitrificans. CONCLUSION: We consider the rapid ribosequencing method as a promising new tool for the analysis of infectious agents in primarily sterile body fluids where conventional culturing of microorganisms fails.