Microbial Proteomics in Pathogen Detection

Though in its infancy, proteomic technology has the potential to play a key role in the future of clinical microbiology diagnostics as techniques become more rapid, affordable, and the list of applicable biomarkers expands. Mass spectroscopy (MS) and 2-D gel electrophoresis are the 2 common techniques in microbial proteomics (Douglas, 2004). In 2-D gel electrophoresis, proteins are first separated by their isoelectric point (pI) in glass tubes (Bjellqvist et al., 1982). Gels are then removed from glass and placed horizontally on top of polyacrylamide slab gels, and polyacrylamide gel electrophoresis (PAGE) further separates proteins with similar charges by their size (molecular weight). Gel electrophoresis is a simple method to catalogue microbial proteins grown under different conditions and disease states. A mass spectrometer can take proteins from PAGE and further separate them by producing charged particles (ions) (Shevchenko et al., 1996). The mass spectrometer differentially moves ionized molecules, separated by their mass-to-charge (m /z) ratio, through a vacuum by means of an electromagnetic field. For the sake of discussion, if one assumes that each component of the mixture has a different molecular weight, then the mass spectrum contains unique "peaks" for each compound that is present. For more information about the different types of mass spectroscopy, refer to Douglas (2004).

A few reports have begun to surface in the clinical microbiology literature and describe how proteomic methods may impact laboratories in the future. In one report, matrix-assisted laser desorption/ionization time-of-flight mass spectroscopy (MALDI-TOF-MS) was used to rapidly identify fungal proteins that evoked a specific human immune response, which may prove to be linked to active infection and outcome (Pitarch et al., 2004). In another study, MALDI-TOF-MS, gelelec-trophoresis, and tandem mass spectrometry were used to identify intra-amniotic proteins, which could lead to discovery of novel human biomarkers for human intra-amniotic infection (Gravett et al., 2004). Ultimately, these tools will help to elucidate the interaction of proteins with protein precursors, DNA, and mRNA to add to the understanding of pathogenesis and disease. Out of this understanding, novel biomarkers for early detection of disease or disease outcomes are expected to occur.

One emerging technology, the Luminex xMap System, can identify multiple immune proteins, like serotype-specific antibodies, in a single well or tube multiplex format. It has been used to identify multiple immune proteins (Jones et al., 2002) and bacterial DNA (Dunbar et al., 2003), but routine applications in the clinical laboratory will require further translational.

Although mass spectrometry is typically used to identify proteins, highperformance mass spectrometry has recently been adapted and developed for use in conjunction with PCR for rapid identification and strain typing of emerging pathogens, such as Bacillus anthracis and coronavirus, among others (Van Ert et al., 2004; Ecker et al., 2005; Sampath et al., 2005)

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