Cytolethal Distending Toxin Is Essential for Helicobacter hepaticus Colonization in Outbred Swiss Webster Mice

Division of Comparative Medicine, Massachusetts Institute of Technology, 16-873, 77 Massachusetts Avenue, Cambridge, MA 02131, USA.
Infection and Immunity (Impact Factor: 3.73). 06/2005; 73(6):3559-67. DOI: 10.1128/IAI.73.6.3559-3567.2005
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Helicobacter hepaticus, which induces chronic hepatitis and typhlocolitis in susceptible mouse strains, produces a cytolethal distending toxin (CDT)
consisting of CdtA, CdtB, and CdtC. A cdtB-deficient H. hepaticus isogenic mutant (HhcdtBm7) was generated and characterized for colonization parameters in four intestinal regions (jejunum,
ileum, cecum, and colon) of outbred Swiss Webster (SW) mice. Inactivation of the cdtB gene abolished the ability of HhcdtBm7 to colonize female mice at both 8 and 16 weeks postinfection (wpi), whereas HhcdtBm7
colonized all of four intestinal regions of three of five males at 8 wpi and then was eliminated by 16 wpi. Wild-type (WT)
H. hepaticus was detected in the corresponding intestinal regions of both male and female mice at 8 and 16 wpi; however, colonization
levels of WT H. hepaticus in the cecum and colon of male mice were approximately 1,000-fold higher than in females (P < 0.0079) at 16 wpi. Infection with WT H. hepaticus, but not HhcdtBm7, at 8 wpi was associated with significantly increased mRNA level of ileal and cecal gamma interferon (IFN-γ)
in females (P < 0.016 and 0.031 between WT H. hepaticus-infected and sham-dosed females, respectively). In contrast, the mRNA levels of IFN-γ were significantly higher in the colon
(P < 0.0079) and trended to be higher in the cecum (P < 0.15) in the HhcdtBm7-colonized male mice versus the sham-dosed controls at 8 wpi. In addition, mRNA levels of ileal IFN-γ
were significantly higher in the control females than males at 8 wpi (P < 0.016). There were significantly higher Th1-associated immunoglobulin G2a (IgG2a), Th2-associated IgG1 and mucosal IgA
(P < 0.002, 0.002, 0.002, respectively) responses in the mice infected with WT H. hepaticus when compared to HhcdtBm7 at 16 wpi. Colonic interleukin-10 (IL-10) expressions at 16 wpi were significantly lower in both
female and male mice colonized by WT H. hepaticus or in males transiently colonized through 8 wpi by HhcdtBm7 versus control mice (P < 0.0159). These lines of evidence indicate that (i) H. hepaticus CDT plays a crucial role in the persistent colonization of H. hepaticus in SW mice; (ii) SW female mice are more resistant to H. hepaticus colonization than male mice; (iii) there was persistent colonization of WT H. hepaticus in cecum, colon, and jejunum but only transient colonization of H. hepaticus in the ileum of female mice; (iv) H. hepaticus colonization was associated with down-regulation of colonic IL-10 production.

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    • "After necropsy, tissue samples from both cecum and colon were weighed, homogenized and plated on selective blood plates for H. hepaticus cfu counts per mg tissue (MHH), or DNA was extracted for quantitative PCR (MIT) [58], [59]. CFU counts from MHH infection experiments and qPCR-derived H. hepaticus genome counts from the corresponding MIT infections yielded similar results. "
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    • "Using NF-κBdeficient mice to study persistent C. jejuni infection, one study indicated a proinflammatory role for Cj-CDT in vivo, as toxinproducing strains were associated with significantly enhanced severity of gastritis and significantly higher occurrence of gastric hyperplasia and dysplasia, which are markers of an early neoplastic process (Fox et al., 2004). For Helicobacter hepaticus , which causes chronic hepatitis and typhlocolitis in some mice, CDT was demonstrated to be critical for persistent infection of Swiss Webster mice, which is characterized with severe inflammation (Ge et al., 2005). In another study, H. hepaticus infection of A/JCr mice demonstrated a significantly higher inflammatory response in strains producing Hh-CDT than isogenic mutant strains. "
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    • "Bacterial transformation was performed by electroporation as previously described with minor modifications [51]. All mutants were passaged five times to ensure the elimination of any WT contaminants. "
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