Proteomic analysis of plasma from Holstein cows testing positive for Mycobacterium avium subsp. paratuberculosis (MAP).
Johne's disease (JD) is a widespread and economically important chronic inflammatory disease of the small intestine of ruminants caused by Mycobacterium avium subsp. paratuberculosis (MAP). Although there are several techniques available for diagnosis of JD, their sensitivity is questionable. New proteome profiling methods, such as serum/plasma protein fingerprinting by 2-Dimensional Fluorescence Difference Gel Electrophoresis (2D-DIGE), may therefore be useful for identifying novel protein biomarkers of MAP infection. In this study, plasma samples were collected from 380 Holstein cows and screened for the presence of MAP infection using the M.pt. Johne's antibody Kit (IDEXX). Five negative (MAP-), and 5 strongly positive (MAP+) cows were selected for proteomic analysis. Highly abundant proteins were depleted from the plasma samples using the ProteoMiner technology (Bio-Rad) to enhance the resolution of low abundance proteins. Plasma samples from MAP-, MAP+, and a pooled internal control were labelled with different fluorescent dyes and separated based on their isoelectrical point (IP) and then their molecular weight. Gel images of the fluorescent plasma protein maps were acquired using a Typhoon scanner and analyzed using the DeCyder software. Proteins that were differentially expressed were excised from the gels, trypsin digested, and subjected to MS/MS analysis for identification. Six proteins were identified as being up-regulated at least 2-fold in MAP+ cows including: transferrin, gelsolin isoforms α & β (actin binding protein - ABP), complement subcomponent C1r, complement component C3, amine oxidase - copper containing 3 (AOC3), and coagulation factor II (thrombin) (p<0.05). Two proteins that were down-regulated approximately 2-fold in the MAP+ cows included coagulation factor XIII -B polypeptide (COAFXIII), and fibrinogen γ chain (FGG) and its precursor.
Department of Animal and Poultry Science, University of Guelph, Guelph, Ontario, Canada. qyou@uoguelph.ca
Qiumei
Q
You
Chris P
CP
Verschoor
Sameer D
SD
Pant
Joseph
J
Macri
Gordon M
GM
Kirby
Niel A
NA
Karrow
D006631:Amine Oxidase (Copper-Containing) / Q000097:blood; D000818:Animals; D015415:Biomarkers / Q000097:blood; D002417:Cattle; D002418:Cattle Diseases / Q000097:blood* / Q000175:diagnosis / Q000382:microbiology; D015923:Complement C1r / Q000032:analysis; D003176:Complement C3 / Q000032:analysis; D005260:Female; D005456:Fluorescent Dyes; D018260:Gelsolin / Q000097:blood; D016927:Mycobacterium avium subsp. paratuberculosis; D010283:Paratuberculosis / Q000097:blood* / Q000175:diagnosis; D040901:Proteomics / Q000379:methods; D011516:Prothrombin / Q000032:analysis; D014168:Transferrin / Q000032:analysis; D057825:Two-Dimensional Difference Gel Electrophoresis / Q000662:veterinary
D015415:Biomarkers; D003176:Complement C3; D005456:Fluorescent Dyes; D018260:Gelsolin; D014168:Transferrin; D011516:Prothrombin; D006631:Amine Oxidase (Copper-Containing); D015923:Complement C1r