Abstract
Antibodies provide a sensitive indicator of proteins displayed by bacteria during sepsis. Because signals produced by infection are naturally amplified during the antibody response, host immunity can be used to identify biomarkers for proteins that are present at levels currently below detectable limits. We developed a microarray comprising ∼70% of the 4066 proteins contained within the Yersinia pestis proteome to identify antibody biomarkers distinguishing plague from infections caused by other bacterial pathogens that may initially present similar clinical symptoms. We first examined rabbit antibodies produced against proteomes extracted from Y. pestis, Burkholderia mallei, Burkholderia cepecia, Burkholderia pseudomallei, Pseudomonas aeruginosa, Salmonella typhimurium, Shigella flexneri, and Escherichia coli, all pathogenic Gram-negative bacteria. These antibodies enabled detection of shared cross-reactive proteins, fingerprint proteins common for two or more bacteria, and signature proteins specific to each pathogen. Recognition by rabbit and non-human primate antibodies involved less than 100 of the thousands of proteins present within the Y. pestis proteome. Further antigen binding patterns were revealed that could distinguish plague from anthrax, caused by the Gram-positive bacterium Bacillus anthracis, using sera from acutely infected or convalescent primates. Thus, our results demonstrate potential biomarkers that are either specific to one strain or common to several species of pathogenic bacteria.
Highlights
Antibodies provide a sensitive indicator of proteins displayed by bacteria during sepsis
Y. pestis Proteome Microarray—Entry ORF clones for Y. pestis (KIM) were subcloned into the pEXP7-DEST expression vector (20) and used to transform E. coli DH10B for preparation of plasmids
We identified several putative antibody biomarkers for Gram-negative pathogens by using a microarray encompassing the majority of proteins from the Y. pestis proteome
Summary
CO92 (14), and the closely related enteric pathogen Yersinia pseudotuberculosis (15, 16) contains ϳ4038 proteins (chromosome plus plasmids). Zhu et al (17) reported the development of a proteome microarray containing the full-length, purified expression products of over 93% of the 6280 proteincoding genes of the yeast Saccharomyces cerevisiae, and Schmid et al (18) described the human antibody repertoire for vaccinia virus recognition by using a viral proteome microarray. This approach opens the possibility of examining the entire bacterial proteome to elucidate proteins or protein pathways that are essential to pathogenicity or host immunity. Based on results from experiments using the Y. pestis proteome microarray, we identified new candidates for antibody biomarkers of bacterial infections and patterns of cross-reactivity that may be useful diagnostic tools
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