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Serum biomarkers of Burkholderia mallei infection elucidated by proteomic imaging of skin and lung abscesses.

ABSTRACT: BACKGROUND:The bacterium Burkholderia mallei is the etiological agent of glanders, a highly contagious, often fatal zoonotic infectious disease that is also a biodefense concern. Clinical laboratory assays that analyze blood or other biological fluids are the highest priority because these specimens can be collected with minimal risk to the patient. However, progress in developing sensitive assays for monitoring B. mallei infection is hampered by a shortage of useful biomarkers. RESULTS:Reasoning that there should be a strong correlation between the proteomes of infected tissues and circulating serum, we employed imaging mass spectrometry (IMS) of thin-sectioned tissues from Chlorocebus aethiops (African green) monkeys infected with B. mallei to localize host and pathogen proteins that were associated with abscesses. Using laser-capture microdissection of specific regions identified by IMS and histology within the tissue sections, a more extensive proteomic analysis was performed by a technique that combined the physical separation capabilities of liquid chromatography (LC) with the sensitive mass analysis capabilities of mass spectrometry (LC-MS/MS). By examining standard formalin-fixed, paraffin-embedded tissue sections, this strategy resulted in the identification of several proteins that were associated with lung and skin abscesses, including the host protein calprotectin and the pathogen protein GroEL. Elevated levels of calprotectin detected by ELISA and antibody responses to GroEL, measured by a microarray of the bacterial proteome, were subsequently detected in the sera of C. aethiops, Macaca mulatta, and Macaca fascicularis primates infected with B. mallei. CONCLUSIONS:Our results demonstrate that a combination of multidimensional MS analysis of traditional histology specimens with high-content protein microarrays can be used to discover lead pairs of host-pathogen biomarkers of infection that are identifiable in biological fluids.

SUBMITTER: Glaros TG

PROVIDER: S-EPMC4450996 | biostudies-literature | 2015

REPOSITORIES: biostudies-literature

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Serum biomarkers of Burkholderia mallei infection elucidated by proteomic imaging of skin and lung abscesses.

Glaros Trevor G TG Blancett Candace D CD Bell Todd M TM Natesan Mohan M Ulrich Robert G RG

Clinical proteomics 20150310 1

<h4>Background</h4>The bacterium Burkholderia mallei is the etiological agent of glanders, a highly contagious, often fatal zoonotic infectious disease that is also a biodefense concern. Clinical laboratory assays that analyze blood or other biological fluids are the highest priority because these specimens can be collected with minimal risk to the patient. However, progress in developing sensitive assays for monitoring B. mallei infection is hampered by a shortage of useful biomarkers.<h4>Resul ...[more]

PMID: 26034464

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Project description:Burkholderia pseudomallei and Burkholderia mallei cause the diseases melioidosis and glanders, respectively. A well-studied aspect of pathogenesis by these closely-related bacteria is their ability to invade and multiply within eukaryotic cells. In contrast, the means by which B. pseudomallei and B. mallei adhere to cells are poorly defined. The purpose of this study was to identify adherence factors expressed by these organisms.Comparative sequence analyses identified a gene product in the published genome of B. mallei strain ATCC23344 (locus # BMAA0649) that resembles the well-characterized Yersinia enterocolitica autotransporter adhesin YadA. The gene encoding this B. mallei protein, designated boaA, was expressed in Escherichia coli and shown to significantly increase adherence to human epithelial cell lines, specifically HEp2 (laryngeal cells) and A549 (type II pneumocytes), as well as to cultures of normal human bronchial epithelium (NHBE). Consistent with these findings, disruption of the boaA gene in B. mallei ATCC23344 reduced adherence to all three cell types by ~50%. The genomes of the B. pseudomallei strains K96243 and DD503 were also found to contain boaA and inactivation of the gene in DD503 considerably decreased binding to monolayers of HEp2 and A549 cells and to NHBE cultures.A second YadA-like gene product highly similar to BoaA (65% identity) was identified in the published genomic sequence of B. pseudomallei strain K96243 (locus # BPSL1705). The gene specifying this protein, termed boaB, appears to be B. pseudomallei-specific. Quantitative attachment assays demonstrated that recombinant E. coli expressing BoaB displayed greater binding to A549 pneumocytes, HEp2 cells and NHBE cultures. Moreover, a boaB mutant of B. pseudomallei DD503 showed decreased adherence to these respiratory cells. Additionally, a B. pseudomallei strain lacking expression of both boaA and boaB was impaired in its ability to thrive inside J774A.1 murine macrophages, suggesting a possible role for these proteins in survival within professional phagocytic cells.The boaA and boaB genes specify adhesins that mediate adherence to epithelial cells of the human respiratory tract. The boaA gene product is shared by B. pseudomallei and B. mallei whereas BoaB appears to be a B. pseudomallei-specific adherence factor.

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Serum biomarkers of Burkholderia mallei infection elucidated by proteomic imaging of skin and lung abscesses.

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Serum biomarkers of Burkholderia mallei infection elucidated by proteomic imaging of skin and lung abscesses.

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