Abstract
BackgroundThe 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.ResultsReasoning 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.ConclusionsOur 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.Electronic supplementary materialThe online version of this article (doi:10.1186/s12014-015-9079-4) contains supplementary material, which is available to authorized users.
Highlights
The bacterium Burkholderia mallei is the etiological agent of glanders, a highly contagious, often fatal zoonotic infectious disease that is a biodefense concern
Lung and skin tissues procured during necropsy of Chlorocebus aethiops monkeys that succumbed to aerosol infection by B. mallei, as well as non-infected controls, were examined for inflammation and bacterial burden associated with glanders
Because the respiratory tract was the primary route of infection, we first examined lung tissue that contained an abscess with B. mallei observable by IHC (Figure 2B)
Summary
The bacterium Burkholderia mallei is the etiological agent of glanders, a highly contagious, often fatal zoonotic infectious disease that is 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. Progress in developing sensitive assays for monitoring B. mallei infection is hampered by a shortage of useful biomarkers. Elevated levels of pathogen-specific antibodies and antigens are indicators of current or recent infection, while perturbations of other serum proteins can illuminate disease progression and recovery. The identification of these classes of critical biomarkers in biological fluids obtainable by non-invasive means is generally performed by trial and error. Limited use of PCR based tests [6], complement fixation and agglutination assays [7] were reported
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