Biomarker Discovery in Subclinical Mycobacterial Infections of Cattle

Abstract

BackgroundBovine tuberculosis is a highly prevalent infectious disease of cattle worldwide; however, infection in the United States is limited to 0.01% of dairy herds. Thus detection of bovine TB is confounded by high background infection with M. avium subsp. paratuberculosis. The present study addresses variations in the circulating peptidome based on the pathogenesis of two biologically similar mycobacterial diseases of cattle.Methodology/Principal FindingsWe hypothesized that serum proteomes of animals in response to either M. bovis or M. paratuberculosis infection will display several commonalities and differences. Sera prospectively collected from animals experimentally infected with either M. bovis or M. paratuberculosis were analyzed using high-resolution proteomics approaches. iTRAQ, a liquid chromatography and tandem mass spectrometry approach, was used to simultaneously identify and quantify peptides from multiple infections and contemporaneous uninfected control groups. Four comparisons were performed: 1) M. bovis infection versus uninfected controls, 2) M. bovis versus M. paratuberculosis infection, 3) early, and 4) advanced M. paratuberculosis infection versus uninfected controls. One hundred and ten differentially elevated proteins (P≤0.05) were identified. Vitamin D binding protein precursor (DBP), alpha-1 acid glycoprotein, alpha-1B glycoprotein, fetuin, and serine proteinase inhibitor were identified in both infections. Transthyretin, retinol binding proteins, and cathelicidin were identified exclusively in M. paratuberculosis infection, while the serum levels of alpha-1-microglobulin/bikunin precursor (AMBP) protein, alpha-1 acid glycoprotein, fetuin, and alpha-1B glycoprotein were elevated exclusively in M. bovis infected animals.Conclusions/SignificanceThe discovery of these biomarkers has significant impact on the elucidation of pathogenesis of two mycobacterial diseases at the cellular and the molecular level and can be applied in the development of mycobacterium-specific diagnostic tools for the monitoring progression of disease, response to therapy, and/or vaccine based interventions.

Circulating Mycobacterium bovis Peptides and Host Response Proteins as Biomarkers for Unambiguous Detection of Subclinical Infection

“…paratuberculosis infection. A detailed description of the sample processing and analysis using iTRAQ was described by Seth et al (13). Host-specific peptides and relative abundance (Ͼ95% confidence interval [CI]) were identified using the ProteinPilot software 2.0 and 2.01 (Applied Biosystems, Inc., Foster City, CA) and the nr_bos_CTM_20070802 FASTA database.…”

“…In order to minimize cross-reactivity, we identified 32 host and 16 pathogen circulating peptides that are unique to M. bovis infection and followed up over a 5-month period using a highly sensitive proteomics approach (13). During infection, Mycobacterium-specific proteins, peptides, DNA, and lipids are released into the circulation (14,15) and are likely to provide unprecedented specificity to identify M. bovis infection.…”

Direct Inoculation of Mycobacteriumavium Subspecies Paratuberculosis Into Ileocecal Peyer’s Patches Results in Colonization of the Intestine in a Calf Model

“…The prevalence is unknown in many parts of the world, but in the United States it has been recently estimated to affect as high as 68% of dairy herds. 17 Economic losses result directly from reduced milk yield, premature culling, and reduced slaughter value and indirectly as shedding adults serve as a source of infection for susceptible herd mates and young calves. 12 A considerable amount of research using a variety of experimental model systems has been devoted to this disease in recent years, yet the pathogenesis of intestinal Map infection remains incompletely understood.…”

The identification of tuberculosis biomarkers in human urine samples

“…In addition, SECTM1 has been shown to be a cognate of CD7, a protein found on T-cells, natural killer cells and pre-B cells and, while the function of SECTM1 is not well established [27], the SECTM1 cognate has been reported to stimulate the upregulation of CD25, CD54 and CD69 on human natural killer cells in vitro in TB infection. Furthermore, serum levels of AMBP have also been observed to be elevated levels in cattle with sub-clinical M. bovis infections [25]. Reduced levels of PTGDS have been reported in cerebrospinal fluid samples from patients with bacterial meningitis [28]; however, to our knowledge there has been no previous correlation between PTGDS and TB.…”

“…A number of these have been previously observed to be associated with TB infection. For example, two related studies showed that RBP4 was a marker for TB infection, with elevated levels of RBP4 being found in the serum of cattle infected with Mycobacterium bovis [25] and with differential expression of RBP4 being observed in human whole blood supernatant [26]. In addition, SECTM1 has been shown to be a cognate of CD7, a protein found on T-cells, natural killer cells and pre-B cells and, while the function of SECTM1 is not well established [27], the SECTM1 cognate has been reported to stimulate the upregulation of CD25, CD54 and CD69 on human natural killer cells in vitro in TB infection.…”