Early Immune Markers Associated withMycobacterium aviumsubsp.paratuberculosisInfection in a Neonatal Calf Model

Abstract

The objective of this study was to observe early markers of cell-mediated immunity in naïve calves infected with Mycobacterium avium subsp. paratuberculosis and how expression of these markers evolved over the 12-month period of infection. Groups for experimental infection included control (noninfected), oral (infected orally with M. avium subsp. paratuberculosis strain K-10), oral/DXM (pretreatment with dexamethasone before oral inoculation), intraperitoneal (i.p.) inoculation, and oral/M (oral inoculation with mucosal scrapings from a cow with clinical disease) groups. One of the earliest markers to emerge was antigen-specific gamma interferon (IFN-␥). Only i.p. inoculated calves had detectable antigen-specific IFN-␥ responses at 7 days, with responses of the other infection groups becoming detectable at 90 and 120 days. All infection groups maintained robust IFN-␥ responses for the remainder of the study. At 1 month, calves in the oral and oral/M groups had higher antigen-stimulated interleukin-10 (IL-10) levels than calves in the other treatment groups, but IL-10 secretion declined by 12 months for all calves. T-cell activation markers such as CD25, CD26, CD45RO, and CD5 were significantly upregulated in infected calves compared to noninfected controls. Oral inoculation of calves resulted in significantly increased antigen-specific lymphocyte proliferation at 9 and 12 months, as well as inducible nitric oxide synthase (iNOS) secretion at 6 and 12 months. These results demonstrate that infection of naïve calves with M. avium subsp. paratuberculosis invoked early immunologic responses characterized by robust antigen-specific IFN-␥ responses and induction of CD25 and CD45RO expression on T-cell subsets. These were followed by antigen-specific lymphocyte proliferation, iNOS secretion, and expression of CD26 and CD5 bright markers in the latter part of the 12-month study.Deciphering host immune responses upon exposure to Mycobacterium avium subsp. paratuberculosis, differentiating the responses due to exposure from true infection, and then further characterizing responses at different stages of infection have been daunting and complex tasks that remain elusive. Early measures of host immune responses to infection with mycobacteria, including M. avium subsp. paratuberculosis, have been dominated by a strong bias toward Th1-mediated gamma interferon (IFN-␥) production. Higher levels of antigen-specific IFN-␥ secreted by peripheral blood mononuclear cells (PBMCs) have been reported for animals in the early stages of M. avium subsp. paratuberculosis infection, whether it be natural or experimental infection (11,18,22,24). Since IFN-␥ is a key effector cytokine involved in the activation of T cells and macrophages, maturation of dendritic cells, upregulation of major histocompatibility complex (MHC) class I and II molecules, and production of reactive oxygen and nitrogen species by macrophages, it is purported to be not only an immune response variable but also a correlate of immune protection (7,16). However, it is u...

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

“…Once a high enough intracellular burden has been reached, MAP can trigger necrosis within infected bovine macrophages (Periasamy et al, 2013), suggesting that necrotic macrophages in the early stages of granuloma formation may contribute returning MAP to the host epithelium during the progression of Johne's disease. The model described here, and the inflammatory signals measured during the differing stages of infection, are supported by the scientific literature which has reported the subclinical and clinical stages of Johne's disease and analysed the immunological responses specific to the mucosal intestinal tissue during each stage (Buza et al, 2003;Khare et al, 2012;Lee et al, 2001;Stabel & Robbe-Austerman, 2011;Weiss et al, 2006). Experimental observations support the role of induced immune tolerance in mucosal tissue during early stages of Johne's disease.…”

“…IL-18 is important in immunomodulation during mycobacterial infection, as IL-18-deficient mice are unable to stimulate interferon gamma (IFNc) and develop an excess of granulomatous lesions in response to M. tuberculosis and Mycobacterium bovis BCG challenge (Sugawara et al, 1999). IFNc is a well-known immune signal for combating mycobacterial disease, serves as a marker used to aid in diagnosing MAP infection and can be seen as early as 3 months after oral infection in cattle (Stabel & Robbe-Austerman, 2011). By creating an early anti-inflammatory environment within the epithelium, early MAP phenotypes may be used to create a microenvironment within the mucosal tissue which supports enhanced uptake, intracellular survival and spread of the bacterium.…”

Mycobacterium avium subsp. paratuberculosis Antibody Response, Fecal Shedding, and Antibody Cross-Reactivity to Mycobacterium bovis in M. avium subsp. paratuberculosis-Infected Cattle Herds Vaccinated against Johne's Disease

“…paratuberculosis vaccination of cattle has been shown to result in M. avium subsp. paratuberculosis-specific gamma interferon (IFN-␥) responses in vaccinated calves by 7 days and specific antibody responses in 80% of calves by 3 to 6 months (16,17). The strong antibody levels reported by those authors were sustained throughout the 12-month study period (16,17).…”

Virulence and Immunity Orchestrated by the Global Gene Regulator sigL in Mycobacterium avium subsp. paratuberculosis

“…To evaluate expansion of the cellular immune response following challenge, splenocytes of immunized and challenged mice were analyzed for the production of key cytokines associated with protection against paratuberculosis (37,38). The PPDj-stimulated splenocytes from ⌬sigL mutant-immunized and challenged mice secreted higher levels of IFN-␥ than those from mock-challenged animals at 6 WPC, indicating increased levels of T cell activity (T-helper 1 cells) in the animals that received the ⌬sigL mutant (see Fig.…”