B cell phenotypes and maturation states in cows naturally infected with Mycobacterium avium subsp. Paratuberculosis

Authors: Stabel, Judith; Bannantine, John; Humphrey, Samuel

Submitted to: PLOS ONE
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 11/14/2022
Publication Date: 12/7/2022
Citation: Stabel, J.R., Bannantine, J.P., Humphrey, S.B. 2022. B cell phenotypes and maturation states in cows naturally infected with Mycobacterium avium subsp. Paratuberculosis. PLOS ONE. https://doi.org/10.1371/journal.pone.0278313.
DOI: https://doi.org/10.1371/journal.pone.0278313

Interpretive Summary: Johne's disease is a chronic, debilitating intestinal disorder in cattle,sheep and wild ruminants, characterized by diarrhea, reduced feed intake, weight loss and death. Animals usually become infected when they are young by ingesting feces containing the causative bacteria. However, symptoms of disease do not usually present themselves until the animals reach 3 to 5 years of age or even older. During this time the animal is infected and may be shedding the organism in its feces without showing any clinical signs of disease. In addition to reduced production by these animals through reduced milk production, they also present a potential infective threat to the rest of the herd. When cows reach a stage of advanced infection, a switch in the immune response occurs, resulting in the appearance of serum antibodies. These antibodies do not appear to protect the animal from infection or advanced disease. However, the appearance of antibodies is associated with an increase in the number of circulating B cells. It is not known why there is a significant shift in higher B cell numbers in the later stages of disease, especially if they seem to be nonprotective. In the present study, we isolated cells from animals in different stages of infection to assess B cell subpopulations and how this would correlate with disease state. The appearance of specific B cells in clinical cows in response to antigen stimulation indicates decreased functionality of traditional B cells in the last stages of disease and the appearance of novel subsets that may help the host.

Technical Abstract: Little is known about the role that B cells play in immune responses to infection with the intracellular pathogen, Mycobacterium avium subsp. paratuberculosis (MAP). Traditionally, the role of B cells has been constrained to their function as antibody-producing cells, however, antibodies are not thought to play a protective role in mycobacterial infections. The present study was designed to characterize B cell subpopulations as well as activation/maturation states in cattle with paratuberculosis. Peripheral blood mononuclear cells (PBMCs) were isolated from noninfected control cows (n = 8); as well cattle naturally infected with MAP in the subclinical (n = 8) and clinical (n = 7) stage of infection and stimulated with MAP antigen for 6 days. MAP infection resulted in greater numbers of total B cells for clinical cows compared to control noninfected cows. The major subpopulation in freshly isolated PBMCs in clinical cows was B-1a B cells, but this shifted to a composite of both B-1a and B-2 B cells upon stimulation of PBMCs with either MAP antigen or pokeweed mitogen, with higher numbers of B-2 B cells. Early B cells were observed to predominate the population of B cells in PBMCs, with lesser populations of germinal B cells, memory B cells and plasma cells. These subpopulations were elevated in clinical cows upon stimulation of PBMCs with MAP antigen, except for plasma cells which were lower compared to control noninfected cows. Increased numbers of B cells in clinical cows aligned with higher expression of B cell markers such as MAPK1/3, BTG1, Bcl2, CD79A and SWAP70, depending upon in vitro stimulation with either mitogen or antigen. This would indicate that the B cells were capable of activation but were anti-apoptotic in nature. The shift to B-2 B cells in the periphery of clinical cows seems to be indicative of an expansion of memory B cells, rather than plasma cells. This may be a last attempt by the host to control the rampant inflammatory state associated with advanced clinical disease.