Localization of Helicobacter spp. in the fundic mucosa of laboratory Beagle dogs: an ultrastructural study

Abstract

In dogs Helicobacter spp. are found in all gastric regions usually localized in the surface mucus, gastric glands and parietal cells. The aim of this study was to detail the distribution of Helicobacter spp. in the fundic mucosa of asymptomatic Beagle dogs and their intracellular localization within parietal cells, in order to evaluate species-specific pathogenetic effects on gastric cells. The presence of Helicobacter spp. was investigated by immunohistochemistry, TEM, and PCR in the fundic mucosa of six Beagle dogs. Helicobacter spp. were found in all dogs examined, and H. bizzozeronii and H. felis were identified by PCR and confirmed by TEM. In the lumen of the fundic glands, co-localization was common. H. bizzozeronii was present in larger numbers than H. felis in both intraluminal and intraparietal localization. The amounts of H. bizzozeronii were similar in superficial and basal portions of the glands. H. felis was predominantly localized in the superficial portions of gastric glands but almost absent from the base. Within parietal cells, most Helicobacter organisms were intracanalicular, but intact and degenerate Helicobacter organisms were also visualized free in the cytoplasm or in secondary lysosomes. No specific degenerative lesions were found in infected parietal cells. Helicobacter organisms were also observed within macrophages in the lamina propria. In conclusion, there is a differential distribution of H. bizzozeronii and H. felis in the fundic mucosa of Beagle dogs, and their intracellular localization in parietal cells and macrophages suggests novel pathogenic scenarios for the development of immune response and maintenance of chronic gastritis in dogs.

Figure 1

Dog, fundic mucosa (immunoperoxidase staining, anti-H. pylori IHC, 400×). Figure 1A - Large amounts of Helicobacter antigen in the lumen of fundic glands (white arrow) and within parietal cells, where Helicobacter antigen is detectable as well preserved spiral-shaped organisms (black arrow) or brown round dots (asterisk). In scattered parietal cells, a diffuse light cytoplasmic staining is also visible. Figure 1B - Helicobacter antigen is occasionally detectable as diffuse and marked cytoplasmic staining (black arrow).

Figure 2

Dog, ultrastructural investigation of the fundic mucosa. Figure 2A. Intraluminal H. bizzozeronii-like organism. A tightly coiled spiral-shaped organism without periplasmic fibrils is free-floating in the superficial mucus (TEM, original magnification 8900×). Figure 2B. Intraluminal H. felis-like organism. A tightly coiled spiral-shaped bacteria with periplasmic fibrils is visible in the superficial mucus; multifocal contact between H. felis-like organism and parietal cell microvilli is present (TEM, original magnification 8900×). Figure 2C. Parietal cell containing Helicobacter-like organisms. Multiple sections of Helicobacter-like organisms were localized within a canaliculus (black arrow) (TEM, original magnification 4 400×). Inset: other bacterial sections were unclearly delimited by parietal cell membranes and adjacent to secondary lysosomes (TEM, original magnification 28000×). Figure 2D. High magnification of the cytoplasm of a parietal cell containing two electrondense intracytoplasmic Helicobacter-like organisms (H). In the upper left corner a longitudinal section of an Helicobacter-like organism is surrounded by closely apposed cell membranes which are focally interrupted; in the lower left corner a transverse section of Helicobacter-like organism is partially enclosed in a lysosome (black arrow) (TEM, original magnification 28000×). Figure 2E. Parietal cell with degenerate Helicobacter-like organism characterized by fragmentation and condensation of bacterial contents and dispersion into the parietal cell cytosol (TEM, original magnification 14000×). Figure 2F. Lamina propria of the fundic mucosa. Macrophage in the interstitium between a blood vessel (right) and the basal portion of gastric epithelial cells resting on a basal lamina (upper left corner) containing multiple profiles of well preserved intracytoplasmic Helicobacter-like organisms and electrondense remnants of an entire phagocytized cell, most likely a parietal cell (TEM, original magnification 8900×).