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Systemic and mucosal immune reactivity upon Mycobacterium avium ssp. paratuberculosis infection in mice.

ABSTRACT: Mycobacterium avium ssp. paratuberculosis (MAP) is the cause of Johne's disease, an inflammatory bowel disorder of ruminants. Due to the similar pathology, MAP was also suggested to cause Crohn's disease (CD). Despite of intensive research, this question is still not settled, possibly due to the lack of versatile mouse models. The aim of this study was to identify basic immunologic mechanisms in response to MAP infection. Immune compromised C57BL/6 Rag2-/- mice were infected with MAP intraperitoneally. Such chronically infected mice were then reconstituted with CD4+ and CD8+ T cells 28 days after infection. A systemic inflammatory response, detected as enlargement of the spleen and granuloma formation in the liver, was observed in mice infected and reconstituted with CD4+ T cells. Whereby inflammation in infected and CD4+CD45RB(hi) T cell reconstituted animals was always higher than in the other groups. Reconstitution of infected animals with CD8+ T cells did not result in any inflammatory signs. Interestingly, various markers of inflammation were strongly up-regulated in the colon of infected mice reconstituted with CD4+CD45RB(lo/int) T cells. We propose, the usual non-colitogenic CD4+CD45RB(lo/int) T cells were converted into inflammatory T cells by the interaction with MAP. However, the power of such cells might be not sufficient for a fully established inflammatory response in the colon. Nevertheless, our model system appears to mirror aspects of an inflammatory bowel disease (IBD) like CD and Johne's diseases. Thus, it will provide an experimental platform on which further knowledge on IBD and the involvement of MAP in the induction of CD could be acquired.

SUBMITTER: Koc A

PROVIDER: S-EPMC3984212 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

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Systemic and mucosal immune reactivity upon Mycobacterium avium ssp. paratuberculosis infection in mice.

Koc Arzu A Bargen Imke I Suwandi Abdulhadi A Roderfeld Martin M Tschuschner Annette A Rath Timo T Gerlach Gerald F GF Hornef Mathias M Goethe Ralph R Weiss Siegfried S Roeb Elke E

PloS one 20140411 4

Mycobacterium avium ssp. paratuberculosis (MAP) is the cause of Johne's disease, an inflammatory bowel disorder of ruminants. Due to the similar pathology, MAP was also suggested to cause Crohn's disease (CD). Despite of intensive research, this question is still not settled, possibly due to the lack of versatile mouse models. The aim of this study was to identify basic immunologic mechanisms in response to MAP infection. Immune compromised C57BL/6 Rag2-/- mice were infected with MAP intraperito ...[more]

PMID: 24728142

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Project description:Chronic enteric Mycobacterium avium ssp. paratuberculosis (MAP) infections are endemic in ruminants globally resulting in significant production losses. The mucosal immune responses occurring at the site of infection, specifically in Peyer's patches (PP), are not well-understood. The ruminant small intestine possesses two functionally distinct PPs. Discrete PPs function as mucosal immune induction sites and a single continuous PP, in the terminal small intestine, functions as a primary lymphoid tissue for B cell repertoire diversification. We investigated whether MAP infection of discrete vs. continuous PPs resulted in the induction of significantly different pathogen-specific immune responses and persistence of MAP infection. Surgically isolated intestinal segments in neonatal calves were used to target MAP infection to individual PPs. At 12 months post-infection, MAP persisted in continuous PP (n = 4), but was significantly reduced (p = 0.046) in discrete PP (n = 5). RNA-seq analysis revealed control of MAP infection in discrete PP was associated with extensive transcriptomic changes (1,707 differentially expressed genes) but MAP persistent in continuous PP elicited few host responses (4 differentially expressed genes). Cytokine gene expression in tissue and MAP-specific recall responses by mucosal immune cells isolated from PP, lamina propria and mesenteric lymph node revealed interleukin (IL)22 and IL27 as unique correlates of protection associated with decreased MAP infection in discrete PP. This study provides the first description of mucosal immune responses occurring in bovine discrete jejunal PPs and reveals that a significant reduction in MAP infection is associated with specific cytokine responses. Conversely, MAP infection persists in the continuous ileal PP with minimal perturbation of host immune responses. These data reveal a marked dichotomy in host-MAP interactions within the two functionally distinct PPs of the small intestine and identifies mucosal immune responses associated with the control of a mycobacterial infection in the natural host.

Project description:Mycobacterium avium subspecies paratuberculosis (MAP) infection is established following the ingestion of bacteria, their penetration of the intestinal mucosa and subsequent events of the host-parasite interaction. These bacteria invade M cells, macrophages and are capable of resisting host defenses and multiply to reach very high numbers intracellular. Mycobactrium avium subsp. avium (MAA) is an antigenically and genetically similar organisms but is relatively nonpathogenic for cattle. MAA organisms appear to infect cattle, but unlike cattle infected with MAP, cattle infected with MAA typically mount an effective systemic immune response, form caseous granuloma, and eliminate the infection. This study was designed to understand the similarities and differences in the host responses during MAP and MAA infection. Neonatal calves were infected (ligated ileal loops) with PBS, MAP, or avirulent MAA following which samples were collected at 30 min, 1h, 2h, 4h, 8h, or 12h. Post-infection, RNA was collected, processed, and then hybridized to custom bovine gene expression arrays, each of which represented 13,258 transcripts spotted in duplicate. Arrays were normalized by scaling against the average reference intensity value (i.e., average across all arrays), normalized by global mean, and then log transformed before statistical analyses were performed. Pairwise comparisons of averaged signal values and Student’s t test were performed using GeneSifter software (VizX Labs, Seattle, WA). A fold-change of at least 1.5-fold and a p value of 0.05 or less was required for a difference in signal to be considered statistically significant. When comparing the transcriptional responses of calves infected with the MAA versus MAP, unique patterns of expression were clearly visible. In general, numbers of transcripts altered in response to infection were much greater for MAA-infected animals than for those infected with the MAP. No genes were significantly up-regulated in MAP-infected animals at the earliest time point tested (30 min), and only modest numbers of genes were increased in expression over the experimental time course. On the other hand, up-regulated transcripts were detected by 30 min in MAA-infected animals and peaked by 2 hr. Differences in the numbers of down-regulated genes between MAP-infected and MAA-infected animals (compared to PBS-infected controls) were less pronounced. At the earliest time points, MAP-infected calves had a greater number of down-regulated genes than did animals infected with MAA. This trend was reversed at 8 and 12 hr post-infection. Keywords: Gene expression profiling by microarray

Project description:Mycobacterium avium subspecies paratuberculosis (MAP) infection is established following the ingestion of bacteria, their penetration of the intestinal mucosa and subsequent events of the host-parasite interaction. These bacteria invade M cells, macrophages and are capable of resisting host defenses and multiply to reach very high numbers intracellular. Mycobactrium avium subsp. avium (MAA) is an antigenically and genetically similar organisms but is relatively nonpathogenic for cattle. MAA organisms appear to infect cattle, but unlike cattle infected with MAP, cattle infected with MAA typically mount an effective systemic immune response, form caseous granuloma, and eliminate the infection. This study was designed to understand the similarities and differences in the host responses during MAP and MAA infection. Neonatal calves were infected (ligated ileal loops) with PBS, MAP, or avirulent MAA following which samples were collected at 30 min, 1h, 2h, 4h, 8h, or 12h. Post-infection, RNA was collected, processed, and then hybridized to custom bovine gene expression arrays, each of which represented 13,258 transcripts spotted in duplicate. Arrays were normalized by scaling against the average reference intensity value (i.e., average across all arrays), normalized by global mean, and then log transformed before statistical analyses were performed. Pairwise comparisons of averaged signal values and StudentM-bM-^@M-^Ys t test were performed using GeneSifter software (VizX Labs, Seattle, WA). A fold-change of at least 1.5-fold and a p value of 0.05 or less was required for a difference in signal to be considered statistically significant. When comparing the transcriptional responses of calves infected with the MAA versus MAP, unique patterns of expression were clearly visible. In general, numbers of transcripts altered in response to infection were much greater for MAA-infected animals than for those infected with the MAP. No genes were significantly up-regulated in MAP-infected animals at the earliest time point tested (30 min), and only modest numbers of genes were increased in expression over the experimental time course. On the other hand, up-regulated transcripts were detected by 30 min in MAA-infected animals and peaked by 2 hr. Differences in the numbers of down-regulated genes between MAP-infected and MAA-infected animals (compared to PBS-infected controls) were less pronounced. At the earliest time points, MAP-infected calves had a greater number of down-regulated genes than did animals infected with MAA. This trend was reversed at 8 and 12 hr post-infection. Keywords: Gene expression profiling by microarray Microarrays were used to examine the transcriptional profiles of bovine intestinal mucosa inoculated with PBS, MAP or MAA and across six time points (30 min, 1, 2, 4, 8, and 12 hours). Experiments were performed in quadruplicate for PBS, MAP, and MAA (bovine ligated ileal loops surgeries were performed with four calves). For one surgery, MAA was used for 4 time ponts only, thus generateing a total of 70 arrays.

Project description:It has long been a quest in ruminants to understand how two very similar mycobacterial species, Mycobacterium avium ssp. paratuberculosis (MAP) and Mycobacterium avium ssp. avium (MAA) lead to either a chronic persistent infection or a rapid-transient infection, respectively. Here, we hypothesized that when the host immune response is activated by MAP or MAA, the outcome of the infection depends on the early activation of signaling molecules and host temporal gene expression. To test our hypothesis, ligated jejuno-ileal loops including Peyer's patches in neonatal calves were inoculated with PBS, MAP, or MAA. A temporal analysis of the host transcriptome profile was conducted at several times post-infection (0.5, 1, 2, 4, 8 and 12 hours). When comparing the transcriptional responses of calves infected with the MAA versus MAP, discordant patterns of mucosal expression were clearly evident, and the numbers of unique transcripts altered were moderately less for MAA-infected tissue than were mucosal tissues infected with the MAP. To interpret these complex data, changes in the gene expression were further analyzed by dynamic Bayesian analysis. Bayesian network modeling identified mechanistic genes, gene-to-gene relationships, pathways and Gene Ontologies (GO) biological processes that are involved in specific cell activation during infection. MAP and MAA had significant different pathway perturbation at 0.5 and 12 hours post inoculation. Inverse processes were observed between MAP and MAA response for epithelial cell proliferation, negative regulation of chemotaxis, cell-cell adhesion mediated by integrin and regulation of cytokine-mediated signaling. MAP inoculated tissue had significantly lower expression of phagocytosis receptors such as mannose receptor and complement receptors. This study reveals that perturbation of genes and cellular pathways during MAP infection resulted in host evasion by mucosal membrane barrier weakening to access entry in the ileum, inhibition of Ca signaling associated with decreased phagosome-lysosome fusion as well as phagocytosis inhibition, bias toward Th2 cell immune response accompanied by cell recruitment, cell proliferation and cell differentiation; leading to persistent infection. Contrarily, MAA infection was related to cellular responses associated with activation of molecular pathways that release chemicals and cytokines involved with containment of infection and a strong bias toward Th1 immune response, resulting in a transient infection.

Project description:BACKGROUND: Mycobacterium avium subsp. paratuberculosis (MAP) causes chronic enteritis in a wide range of animal species. In cattle, MAP causes a chronic disease called Johne's disease, or paratuberculosis, that is not treatable and the efficacy of vaccine control is controversial. The clinical phase of the disease is characterised by diarrhoea, weight loss, drop in milk production and eventually death. Susceptibility to MAP infection is heritable with heritability estimates ranging from 0.06 to 0.10. There have been several studies over the last few years that have identified genetic loci putatively associated with MAP susceptibility, however, with the availability of genome-wide high density SNP maker panels it is now possible to carry out association studies that have higher precision. METHODOLOGY/PRINCIPAL FINDINGS: The objective of the current study was to localize genes having an impact on Johne's disease susceptibility using the latest bovine genome information and a high density SNP panel (Illumina BovineSNP50 BeadChip) to perform a case/control, genome-wide association analysis. Samples from MAP case and negative controls were selected from field samples collected in 2007 and 2008 in the province of Lombardy, Italy. Cases were defined as animals serologically positive for MAP by ELISA. In total 966 samples were genotyped: 483 MAP ELISA positive and 483 ELISA negative. Samples were selected randomly among those collected from 119 farms which had at least one positive animal. CONCLUSION/SIGNIFICANCE: THE ANALYSIS OF THE GENOTYPE DATA IDENTIFIED SEVERAL CHROMOSOMAL REGIONS ASSOCIATED WITH DISEASE STATUS: a region on chromosome 12 with high significance (P<5x10(-6)), while regions on chromosome 9, 11, and 12 had moderate significance (P<5x10(-5)). These results provide evidence for genetic loci involved in the humoral response to MAP. Knowledge of genetic variations related to susceptibility will facilitate the incorporation of this information into breeding programmes for the improvement of health status.

Project description:BackgroundMaintenance of metal homeostasis is crucial in bacterial pathogenicity as metal starvation is the most important mechanism in the nutritional immunity strategy of host cells. Thus, pathogenic bacteria have evolved sensitive metal scavenging systems to overcome this particular host defence mechanism. The ruminant pathogen Mycobacterium avium ssp. paratuberculosis (MAP) displays a unique gut tropism and causes a chronic progressive intestinal inflammation. MAP possesses eight conserved lineage specific large sequence polymorphisms (LSP), which distinguish MAP from its ancestral M. avium ssp. hominissuis or other M. avium subspecies. LSP14 and LSP15 harbour many genes proposed to be involved in metal homeostasis and have been suggested to substitute for a MAP specific, impaired mycobactin synthesis.ResultsIn the present study, we found that a LSP14 located putative IrtAB-like iron transporter encoded by mptABC was induced by zinc but not by iron starvation. Heterologous reporter gene assays with the lacZ gene under control of the mptABC promoter in M. smegmatis (MSMEG) and in a MSMEG∆furB deletion mutant revealed a zinc dependent, metalloregulator FurB mediated expression of mptABC via a conserved mycobacterial FurB recognition site. Deep sequencing of RNA from MAP cultures treated with the zinc chelator TPEN revealed that 70 genes responded to zinc limitation. Remarkably, 45 of these genes were located on a large genomic island of approximately 90 kb which harboured LSP14 and LSP15. Thirty-five of these genes were predicted to be controlled by FurB, due to the presence of putative binding sites. This clustering of zinc responsive genes was exclusively found in MAP and not in other mycobacteria.ConclusionsOur data revealed a particular genomic signature for MAP given by a unique zinc specific locus, thereby suggesting an exceptional relevance of zinc for the metabolism of MAP. MAP seems to be well adapted to maintain zinc homeostasis which might contribute to the peculiarity of MAP pathogenicity.

Dataset Information

Systemic and mucosal immune reactivity upon Mycobacterium avium ssp. paratuberculosis infection in mice.

Publications

Systemic and mucosal immune reactivity upon Mycobacterium avium ssp. paratuberculosis infection in mice.

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