Skip to main content
ARS Home » Midwest Area » Ames, Iowa » National Animal Disease Center » Infectious Bacterial Diseases Research » Research » Publications at this Location » Publication #311755

Title: Characterization of the inflammatory phenotype of Mycobacterium avium subspecies paratuberculosis using a novel cell culture passage model

item EVERMAN, JAMIE - Oregon State University
item ECKSTEIN, TORSTEN - Colorado State University
item ROUSSEY, JONATHAN - Michigan State University
item COUSSENS, PAUL - Michigan State University
item Bannantine, John
item BERMUDEZ, LUIZ - Oregon State University

Submitted to: Microbiology
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 5/6/2015
Publication Date: 5/8/2015
Citation: Everman, J.L., Eckstein, T.M., Roussey, J., Coussens, P., Bannantine, J.P., Bermudez, L.E. 2015. Characterization of the inflammatory phenotype of Mycobacterium avium subspecies paratuberculosis using a novel cell culture passage model. Microbiology. 161:1420-1434. doi: 10.1099/mic.0.000106.

Interpretive Summary: Johne’s disease in livestock such as dairy cattle and sheep is chronic in nature and caused by the bacterium Mycobacterium avium subspecies paratuberculosis (MAP). An interesting observation we made previously is the bacterium changes its surface composition when inside the animal during infection. Specifically, the lipid content of the bacterial cell wall undergoes detectable changes. In this study, we showed that, once MAP is passed through the various cow cells that it would see during an infection, those cow cells generate a stronger inflammatory response with each passage. This we termed the MAP inflammatory phenotype. We further observed that this phenotype is linked to the changes in the cell wall lipids of the MAP bacterium. Thus we believe MAP lipid changes may be responsible for the immunopathology of the cow intestine seen during disease. This research is of primary interest to veterinarians, stakeholders and other researchers in the field.

Technical Abstract: Understanding the pathogenic mechanisms and host responses to Johne’s disease, a chronic enteritis of ruminants caused by Mycobacterium avium subspecies paratuberculosis (MAP), is complicated by the multifaceted disease progression, late-onset host reaction, and the lack of ex vivo infection models available to study the disease in its various stages. Here, we describe a novel in vitro cell culture passage model which mimics the course of the infection in vivo, beginning with the interaction of MAP with the intestinal epithelial barrier, followed by infection of macrophages, and return to the intestinal epithelium. Using this model we determined that the initial infection of MDBK epithelial cells with MAP showed no inflammatory response. After passage through RAW macrophages, bacterial re-infection of MDBK epithelial cells was associated with increased levels of the pro-inflammatory signals IL-6, CCL5, IL-8, and IL-18, paired with decreased levels of TGF-beta compared to those infected with non-passaged MAP. Transcriptome analysis of MAP from each stage of epithelial cell infection identified increased expression of genes involved in lipid biosynthesis and lipoprotein modification in the inflammatory phenotype of MAP after re-entry into epithelial cells. Lipid analysis by HPLC-ES/MS identified a differing lipidomic profile between the two phenotypes and a distinct set of lipids specific to the inflammatory MAP phenotype. We validated our newly described model by the identification of these unregulated lipid-modification gene transcripts in ileal tissue of cows with Johne’s disease. By using this novel in vitro cell culture passage model, we have provide evidence that MAP alters its lipids during intracellular infection and acquires a pro-inflammatory phenotype which is very likely associated with the inflammatory phase of Johne’s disease.