Early Phase Morphological Lesions and Transcriptional Responses of Bovine Ileum Infected with Mycobacterium avium subsp paratuberculosis

Department of Veterinary Pathobiology, College of Veterinary Medicine and Biomedical Sciences, Veterinary Research Building, Bldg. 1197, Room 141, Texas A&M University, College Station, TX 77843-4467, USA.
Veterinary Pathology (Impact Factor: 1.87). 04/2009; 46(4):717-28. DOI: 10.1354/vp.08-VP-0187-G-FL
Source: PubMed


Mycobacterium avium subsp. paratuberculosis (MAP) is the causative agent of chronic enteritis in ruminants (Johne's disease) and a possible etiopathologic agent in human Crohn's disease. The host-pathogen interaction in this chronic disease has largely depended on the randomly collected static lesions studied in subclinically or clinically infected animals. We have established and utilized the neonatal calf ligated ileal loop model to study the early temporal host changes during MAP infection. After inoculation of ligated ileal loop with MAP, samples were analyzed for bacterial invasion, histologic and ultrastructural morphologic changes, and gene expression at several times (0.5-12 hours) postinfection. Our results indicate that MAP invades the intestinal mucosa as early as 0.5 hour postinoculation. Distribution and migration of neutrophils, monocytes/macrophages, and goblet cells were confirmed by histopathology, scanning and transmission electron microscopy. Coincident with the morphologic analysis, we measured by real-time polymerase chain reaction gene expression of various cytokines/chemokines that are involved in the recruitment of mononuclear and polymorphonuclear leukocytes to the site of infection. We also detected expression of several other genes, including intestinal-trefoil factor, profilin, lactoferrin, and enteric ss-defensin, which may play significant roles in the early MAP infection. Thus, the calf ligated intestinal loop model may be used as a human disease model to understand the role of MAP in the pathogenesis of Crohn's disease.

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    • "Glycerinaldehyd-3-phosphat-dehydrogenase (GAPDH) and b-actin housekeeping genes were analyzed as described for duodenal tissue [23] [24] [25]. As internal standard, duodenal tissue from an adult mare obtained with biopsy forceps (Olympus FB-36k-1; Olympus, Vienna, Austria) immediately after slaughter was used. "
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    ABSTRACT: A lack of colostrum may delay postnatal intestinal development in foals. Therefore, in this study the effect of colostrum withdrawal on duodenal development of newborn foals was investigated. Shetland foals were fed either colostrum (group COL, n=5) or milk replacer (group MR, n=8) by nasogastric tube over the first 24 h of life and all foals received one liter of hyperimmune plasma. On day 3 of life, duodenal biopsies were taken via gastroduodenoscopy. Expression of mRNA for receptors for IGF1, IGF2 and TGFβ was analyzed by real-time PCR. Biopsies were analysed for insulin-like growth factor (IGF) receptors 1 and 2, transforming growth factor β (TGFβ) receptor 1, immunostaining for Ki-67 proliferation marker, lectin binding and histomorphology. The mRNA expression of receptors for IGF1, IGF2 and TGFß in the duodenum of newborn foals was not altered by withholding colostrum. The number of goblet cells in the epithelium of the duodenum was higher in MR compared to COL foals (1.95% ±1.15 vs. 1.25% ± 0.75), whereas villus length did not differ between groups. Immunohistochemistry of Ki-67 and lectin binding patterns were similar in foals of both groups. Results indicate that a lack of colostrum is without detrimental effects on postnatal duodenal development in foals.
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    • "This early stage infection process takes only a few hours (Clarke, 1997; Khare et al., 2009; Valentin-Weigand and Goethe, 1999). Upon maturation, phagosomes fuse with lysosomes and the mycobacteria undergo digestion by a process of acidification and enzymatic proteolysis. "
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    ABSTRACT: Evasion of host defense mechanisms and survival inside infected host macrophages are features of pathogenic mycobacteria including Mycobacterium avium subspecies paratuberculosis, the causative agent of Johne's disease in ruminants. Protein tyrosine phosphatase A (PtpA) has been identified as a secreted protein critical for survival of mycobacteria within infected macrophages. The host may mount an immune response to such secreted proteins. In this study, the humoral immune response to purified recombinant Mycobacterium avium subsp. paratuberculosis PtpA was investigated using sera from a cohort of sheep infected with Mycobacterium avium subsp. paratuberculosis and compared with uninfected healthy controls. A significantly higher level of reactivity to PtpA was observed in sera collected from Mycobacterium avium subspecies paratuberculosis infected sheep when compared to those from uninfected healthy controls. PtpA could be a potential candidate antigen for detection of humoral immune responses in sheep infected with Mycobacterium avium subspecies paratuberculosis.
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    • "Although intra-uterine transmission of MAP occurs (Sweeney et al., 1992a,b), MAP is typically acquired orally and most frequently in young calves (Whittington and Sergeant, 2001). From the intestinal lumen, MAP enters the host intestine through M cells, epithelial cells, antigen presenting cells and goblet cells of Peyer's patches in the ileum and jejunum (Khare et al., 2009; Momotani et al., 1988). Furthermore, MAP is phagocytized by macrophages located in the submucosal layers via receptor-mediated endocytosis (Zurbrick and Czuprynski, 1987). "
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    ABSTRACT: Mycobacterium avium subsp. paratuberculosis (MAP) causes Johne's disease (JD), a chronic granulomatous intestinal inflammation of ruminants. Current diagnostic tools lack sensitivity to detect JD early in infection; therefore, alternatives are desired. The objective was to identify potential biomarkers in whole blood of high- and low-dose (LD) MAP-challenged Holstein-Friesian calves 3 months after inoculation. Infected calves were designated MAP-positive using the IFN-γ release assay. Differential expression of transcripts in whole blood was compared between non-infected controls and HD, as well as LD calves, using the Affymetrix® GeneChip® Bovine Genome Array. Microarray data were analysed using RMA and PLIER algorithms; 296 transcripts were differentially expressed (17 had ≥ 1.5 fold change). The HD and LD calves had differential gene expression profiles for up to 80% of differentially expressed genes. Pathway analyses using Ingenuity Pathway Analysis (IPA®) indicated inhibition of several defence mechanisms, including apoptosis, leukocyte and lymphocyte trafficking, overall repression of gene expression and potentially hydrogen peroxide production in macrophages. Further validation using qPCR verified increased expression of CD46, ICOS, and CEP350, but decreased expression of CTLA4, YARS, and PARVB in infected calves. Additionally, a comparison of seropositive and seronegative infected calves identified transcripts predictive of seroconversion. We concluded that IL6ST/gp130 and CD22 may have important roles in the induction of antibodies against MAP. Putative biomarkers of early MAP infection with roles in immune responses were identified; in addition, the importance of infective dose on biomarkers was determined
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