Use of cultivation-dependent and -independent techniques to assess contamination of central venous catheters: A pilot study

Department of Biotechnology, Chemistry, and Environmental Engineering, Aalborg University, Sohngaardsholmsvej 49, DK-9000 Aalborg, Denmark.
BMC Clinical Pathology 11/2008; 8(1):10. DOI: 10.1186/1472-6890-8-10
Source: PubMed


Catheters are the most common cause of nosocomial infections and are associated with increased risk of mortality, length of hospital stay and cost. Prevention of infections and fast and correct diagnosis is highly important.
In this study traditional semiquantitative culture-dependent methods for diagnosis of bacteria involved in central venous catheter-related infections as described by Maki were compared with the following culture-independent molecular biological methods: Clone libraries, denaturant gradient gel electrophoresis, phylogeny and fluorescence in situ hybridization.
In accordance with previous studies, the cultivation of central venous catheters from 18 patients revealed that S. epidermidis and other coagulase-negative staphylococci were most abundant and that a few other microorganisms such as P. aeruginosa and K. pneumoniae occasionally were found on the catheters. The molecular analysis using clone libraries and sequencing, denaturant gradient gel electrophoresis and sequencing provided several important results. The species found by cultivation were confirmed by molecular methods. However, many other bacteria belonging to the phyla Proteobacteria, Firmicutes, Actinobacteria and Bacteroidetes were also found, stressing that only a minor portion of the species present were found by cultivation. Some of these bacteria are known to be pathogens, some have not before been described in relation to human health, and some were not closely related to known pathogens and may represent new pathogenic species. Furthermore, there was a clear difference between the bacterial species found in biofilm on the external (exluminal) and internal (luminal) side of the central venous catheter, which can not be detected by Maki's method. Polymicrobial biofilms were observed on most of the catheters and were much more common than the cultivation-dependent methods indicated.
The results show that diagnosis based on molecular methods improves the detection of microorganisms involved in central catheter-related infections. The importance of these microorganisms needs to be investigated further, also in relation to contamination risk from improper catheter handling, as only in vivo contaminants are of interest. This information can be used for development of fast and more reliable diagnostic tools, which can be used in combination with traditional methods.

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Available from: Trine Rolighed Thomsen,
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    • "Culture-independent identification of bacteria can be carried out by performing a ribosomal RNA (rRNA) gene cycle e.g. (Nielsen et al., 2008; Thomsen et al., 2010). This method is based on the fact that the composition of the gene encoding the bacterial 16S rRNA (16S rDNA) is unique for each bacterial species. "

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    • "Despite the recent appearance of molecular diagnostic methods, namely those based on real-time PCR and quantitative PCR (e.g. Jukes et al. 2010), the method of choice for the detection of S. epidermidis in clinical samples still relies on relatively slow cultivation techniques (Haimi-Cohen et al. 2002; Larsen et al. 2008). It is therefore of the utmost importance to develop novel methods, that are both rapid and robust, to assess the presence of this important pathogen. "
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    • "A combination of cultivation-dependent methods and molecular techniques yielded a more reliable insight into the microbial community. This was also reported by others (Cheng & Foght, 2007; Larsen et al., 2008). "
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