Comparative Gamma Delta T Cell Immunology: A Focus on Mycobacterial Disease in Cattle

A theme among many pathogenic mycobacterial species affecting both humans and animals is a prolonged asymptomatic or latent period that can last years to decades. The mechanisms that favor progression to active disease are not well understood. Pathogen containment is often associated with an effective cell-mediated or T-helper 1 immune profile. With certain pathogenic mycobacteria, such as Mycobacterium avium subspecies paratuberculosis, a shift to active clinical disease is associated with loss of T-helper 1 immunity and development of an ineffective humoral or T-helper 2 immune response. Recently γδ T cells have been shown to play a role early in mycobacterial infections and have been hypothesized to influence disease outcome. The purpose of this paper is to compare recent advancements in our understanding of γδ T cells in humans, cattle, and mice and to discuss roles of γδ T cells in host response to mycobacterial infection

Comparison of Sheep, Goats, and Calves as Infection Models for Mycobacterium avium subsp. paratuberculosis

Animal infection models to study Mycobacterium avium subsp. paratuberculosis (MAP) infection are useful for evaluating the efficacy of vaccines and other therapeutics for the prevention or treatment of infection. The goal of the present study was to compare smaller ruminants, sheep and goats, with calves as infection models. Neonatal sheep, goats, and calves (n = 4) received 109 cfu of a cattle isolate of MAP in milk replacer on days 0, 3 and 6 in a 12-month study and sampled monthly thereafter. Results demonstrated a robust antigen-specific IFN-γ response at 90 days post-inoculation for sheep and goats, with lower responses noted for calves. By 360 days, IFN-γ responses were 50 and 82% higher for calves than for goats and sheep, respectively. Although MAP- specific antibody responses were first observed in sheep at 90 days, calves had higher antibody responses throughout the remainder of the study. Following pass-through shedding on day 7, fecal shedding was fairly negligible across treatments but remained higher for calves throughout the study. Colonization of tissues was variable within treatment group and was higher for calves and sheep for the majority of tissues. Upon antigen stimulation of PBMCs, higher populations of CD4 + T cells cells and lower populations of γδ TCR + and NK cells were observed for goats and calves compared to sheep. Relative gene expression of IL-4, IL-12, and IL-17 in PBMCs was higher in goats, corresponding to lower tissue colonization with MAP. These data suggest that ruminant species are fairly comparable as infection models for MAP, but discrete differences in host responses to MAP infection exist between species

Disseminated \u3ci\u3eLeishmania infantum\u3c/i\u3e infection in two sibling foxhounds due to possible vertical transmission

Two sibling foxhounds born to a Leishmania seropositive bitch were presented after testing seropositive for Leishmania. Leishmania infantum infection was detected via histopathology, culture, and quantitative polymerase chain reaction (q-PCR). This is the first report of natural infection with Leishmania infantum with the possibility for vertical transmission in North America. Infection disséminée à Leishmania infantum chez deux chiots Fox hound d’une même portée reliée possiblement à une transmission verticale. Deux chiots Fox hound d’une même portée nés d’une mère séropositive à Leishmania ont été présentés après un contrôle sérologique positif. Une infection à Leishmania infantum a été détectée par histopathologie, culture et amplification en chaîne par polymérase quantitative (ACP-q). Il s’agit du premier rapport d’infection naturelle par Leishmania infantum possiblement relié à une transmission verticale en Amérique du Nord

Prevention of Mycobacterium avium Subspecies paratuberculosis (MAP) Infection in Balb/c mice by Feeding Lactobacillus acidophilus Strain NP-51®

The immune responses of 390 BALB/c mice fed the probiotic Lactobacillus acidophilus strain NP51 ® and infected with Mycobacterium avium subspecies paratuberculosis (MAP) were evaluated in a 6-month trial. Mice were randomized to nine treatment groups that fed either viable- or heat-killed NP51 and inoculated with either viable- or heatkilled MAP or sterile phosphate-buffered saline. Feeding the NP51 resulted in higher numbers of T lymphocytes in the spleen including the CD8 + cytotoxic T lymphocytes. In addition, feeding the NP51 lowered the number of immune suppressive T regulatory cells CD4 + CD25 + and CD8 + CD25 + cells in the spleen. Additionally, feeding the NP51 resulted in higher concentration of interferon-gamma in the supernatant of splenocytes cultured in vitro. These results suggest that feeding the NP51 to BALB/c mice might prevent the progression of MAP infection in mice

Quantification of Macrophages and Mycobacterium avium Subsp. paratuberculosis in Bovine Intestinal Tissue During Different Stages of Johne’s Disease

Johne’s disease is an enteric disease caused by the intracellular pathogen Mycobacterium avium subsp. paratuberculosis (MAP). Upon ingestion of MAP, it is translocated across the intestinal epithelium and may be killed by intestinal macrophages, or depending on the bacterial burden and immunological status of the animal, MAP may thwart innate defense mechanisms and persist within the macrophage. This study aimed to determine the numbers of macrophages and MAP present in bovine midileal tissue during different stages of infection. Immunofluorescent (IF) labeling was performed on frozen bovine midileal intestinal tissue collected from 28 Holstein dairy cows. The number of macrophages in midileal tissue sections was higher for clinically affected cows, followed by subclinically affected cows and then uninfected control cows. Macrophages were present throughout the tissue sections in clinical cows, including the tunica muscularis, submucosa, and the lamina propria around the crypts and in the villous tips, with progressively fewer macrophages in subclinically affected and control cows. Clinically affected cows also demonstrated significantly higher numbers of MAP and higher numbers of macrophages with intracellular MAP compared to subclinically affected cows. MAP IF labeling was present within the submucosa and lamina propria around the crypts, progressing into the villous tips in some clinically affected cows. Our findings indicate that number of macrophages increases with progression of infection, but a significant number of the macrophages present in the midileum are not associated with MAP

Temporal Dynamics of Chronic Inflammation on the Cecal Microbiota in IL-10-/- Mice

The intestinal microbiota is a critical component of mucosal health as evidenced by the fact that alterations in the taxonomic composition of the gastrointestinal microbiota are associated with inflammatory bowel diseases. To better understand how the progression of inflammation impacts the composition of the gastrointestinal microbiota, we used culture independent taxonomic profiling to identify temporal changes in the cecal microbiota of C3Bir IL-10-/- mice concomitantly with the onset and progression of colitis. This analysis revealed that IL-10-/- mice displayed a biphasic progression in disease severity, as evidenced by histopathological scores and cytokine production. Beginning at 4 weeks of age, pro-inflammatory cytokines including TNF-a, IFN-g, IL-6, G- CSF, and IL-1a as well as chemokines including RANTES and MIP-1a were elevated in the serum of IL-10-/- mice. By 19 weeks of age, the mice developed clinical signs of disease as evidenced by weight loss, which was accompanied by a significant increase in serum levels of KC and IL-17. While the overall diversity of the microbiota of both wild type and IL-10-/- were similar in young mice, the latter failed to increase in complexity as the mice matured and experienced changes in abundance of specific bacterial taxa that are associated with inflammatory bowel disease in humans. Collectively, these results reveal that there is a critical time in young mice between four to six weeks of age when inflammation and the associated immune responses adversely affect maturation of the microbiota

Orally administered extract from \u3ci\u3ePrunella vulgaris\u3c/i\u3e attenuates spontaneous colitis in mdr1a\u3csup\u3e-/-\u3c/sup\u3e mice

AIM: To investigate the ability of a Prunella vulgaris (P. vulgaris) ethanolic extract to attenuate spontaneous typhlocolitis in mdr1a-/- mice. METHODS: Vehicle (5% ethanol) or P. vulgaris ethanolic extract (2.4 mg/d) were administered daily by oral gavage to mdr1a-/- or wild type FVBWT mice from 6 wk of age up to 20 wk of age. Clinical signs of disease were noted by monitoring weight loss. Mice experiencing weight loss in excess of 15% were removed from the study. At the time mice were removed from the study, blood and colon tissue were collected for analyses that included histological evaluation of lesions, inflammatory cytokine levels, and myeloperoxidase activity. RESULTS: Administration of P. vulgaris extracts to mdr1a-/- mice delayed onset of colitis and reduced severity of mucosal inflammation when compared to vehicle-treated mdr1a-/- mice. Oral administration of the P. vulgaris extract resulted in reduced (P \u3c 0.05) serum levels of IL-10 (4.6 ± 2 vs 19.4 ± 4), CXCL9 (1319.0 ± 277 vs 3901.0 ± 858), and TNFα (9.9 ± 3 vs 14.8 ± 1) as well as reduced gene expression by more than two-fold for Ccl2, Ccl20, Cxcl1, Cxcl9, IL-1 α, Mmp10, VCAM-1, ICAM, IL-2, and TNFα in the colonic mucosa of mdr1a-/- mice compared to vehicle-treated mdr1a-/- mice. Histologically, several microscopic parameters were reduced (P \u3c 0.05) in P. vulgaris -treated mdr1a-/- mice, as was myeloperoxidase activity in the colon (2.49 ± 0.16 vs 3.36 ± 0.06, P \u3c 0.05). The numbers of CD4+ T cells (2031.9 ± 412.1 vs 5054.5 ± 809.5) and germinal center B cells (2749.6 ± 473.7 vs 4934.0 ± 645.9) observed in the cecal tonsils of P. vulgaris - treated mdr1a-/- were significantly reduced (P \u3c 0.05) from vehicle-treated mdr1a-/- mice. Vehicle-treated mdr1a-/- mice were found to produce serum antibodies to antigens derived from members of the intestinal microbiota, indicative of severe colitis and a loss of adaptive tolerance to the members of the microbiota. These serum antibodies were greatly reduced or absent in P. vulgaris -treated mdr1a-/- mice. CONCLUSION: The anti-inflammatory activity of P. vulgaris ethanolic extract effectively attenuated the severity of intestinal inflammation in mdr1a-/- mice

A gut pathobiont synergizes with the microbiota to instigate inflammatory disease marked by immunoreactivity against other symbionts but not itself

Inflammatory bowel diseases (IBD) are likely driven by aberrant immune responses directed against the resident microbiota. Although IBD is commonly associated with a dysbiotic microbiota enriched in putative pathobionts, the etiological agents of IBD remain unknown. Using a pathobiont-induced intestinal inflammation model and a defined bacterial community, we provide new insights into the immune-microbiota interactions during disease. In this model system, the pathobiont Helicobacter bilis instigates disease following sub-pathological dextran sulfate sodium treatment. We show that H. bilis causes mild inflammation in mono-associated mice, but severe disease in the presence of a microbiota, demonstrating synergy between the pathobiont and microbiota in exacerbating pathology. Remarkably, inflammation depends on the presence of H. bilis, but is marked by a predominant Th17 response against specific members of the microbiota and not the pathobiont, even upon the removal of the most immune-dominant taxa. Neither increases in pathobiont burden nor unique changes in immune-targeted microbiota member abundances are observed during disease. Collectively, our findings demonstrate that a pathobiont instigates inflammation without being the primary target of a Th17 response or by altering the microbiota community structure. Moreover, our findings point toward monitoring pathobiont-induced changes in microbiota immune targeting as a new concept in IBD diagnotics