JPC SYSTEMIC PATHOLOGY

DIGESTIVE SYSTEM

September 2024

D-B18

 

SLIDE A:

SIGNALMENT (JPC #2333212): Ferret

 

HISTORY: Bruxism, intermittent vomiting, and dark, tarry stools.

 

HISTOPATHOLOGIC DESCRIPTION: Stomach, fundus: Multifocally affecting 90% of the section, parietal and chief cells are replaced by increased numbers of mucous neck cells with rare mitotic figures (mucous cell metaplasia and hyperplasia). Multifocally, nodular aggregates of lymphocytes and plasma cells expand the lamina propria, surround and separate gastric glands, and extend into the muscularis mucosa and submucosa. Mucous cell metaplasia is most prominent in areas of inflammation. There are lightly basophilic 1um thick, spiral shaped bacilli multifocally adhered to the luminal epithelium or within gastric glands.

 

Liver: Low numbers of lymphocytes and plasma cells infiltrate portal connective tissue. Multifocally, there is scattered yellow-brown globular pigment (hemosiderin) within Kupffer cells which are arranged in small nodules lining sinusoids (micronodular hemosiderosis), and hepatocytes occasionally contain single clear vacuoles that peripherally displace the nucleus (lipid-type vacuolar change).

 

Spleen: Multifocally, the red pulp is expanded by moderate extramedullary hematopoiesis. 

 

SLIDE B: Warthin Starry 4.0 Stomach: Numerous 1x3um, slightly curved, argyrophilic bacilli are present on the mucosal surface and within gastric glands.

 

MORPHOLOGIC DIAGNOSIS: 1. Stomach, fundus: Gastritis, lymphoplasmacytic, multifocal, moderate, with moderate mucous cell metaplasia and hyperplasia and superficial argyrophilic curved bacilli, etiology consistent with Helicobacter sp., ferret (Mustela putorius furo), mustelid.

2. Liver: Hepatitis, portal, lymphoplasmacytic, multifocal, minimal.

3. Spleen: No significant lesions.

 

ETIOLOGIC DIAGNOSIS: Helicobacterial gastritis

 

CAUSE: Helicobacter mustelae

 

SLIDE C:

SIGNALMENT (JPC #2985003): 1-year-old male ICR mouse

 

HISTORY: This mouse was jaundiced and died shortly before a necropsy was performed.

 

HISTOPATHOLOGIC DESCRIPTION: Liver: Multifocally expanding and bridging portal areas and extending into adjacent hepatic parenchyma are moderate numbers of lymphocytes, plasma cells, neutrophils, and fewer histiocytes. There are markedly increased numbers of small caliber bile ducts lined by dysplastic epithelium with moderate anisocytosis and anisokaryosis and vesicular nuclei (bile duct hyperplasia). Rarely, bile ducts are lined by attenuated epithelium and contain luminal sloughed cellular and proteinaceous debris. Multifocally, low numbers of individual hepatocytes are swollen with microvacuolated, lacy cytoplasm (glycogen-type degeneration) or are shrunken with hypereosinophilic cytoplasm and pyknotic or karyolytic nuclei (single cell death). Multifocally, most prominently surrounding areas of portal inflammation, hepatocytes demonstrate cytomegaly, karyomegaly, and occasional binucleation, and nuclei have markedly coarse chromatin and multiple nucleoli. Multifocally portal lymphatics are expanded by pale eosinophilic fluid (edema).

 

MORPHOLOGIC DIAGNOSIS: Liver: Cholangiohepatitis, lymphoplasmacytic and neutrophilic, multifocal, moderate, with marked bile duct hyperplasia and dysplasia, and hepatocyte necrosis, karyomegaly and cytomegaly, ICR mouse, rodent.

 

ETIOLOGIC DIAGNOSIS: Helicobacterial hepatitis

 

CAUSE: Helicobacter hepaticus

 

GENERAL DISCUSSION:

• Helicobacter sp. are gram negative, microaerophilic, curved to spiral, motile bacilli with variable numbers of sheathed flagella and can survive the extremely low pH of the stomach
• Infection with Helicobacter spp. is common in many animals, but disease appears to be rare, restricted to a few species
• The ferret is the only domesticated animal where naturally occurring Helicobacter-associated ulcer disease has been described, and ferrets have the most convincing evidence for clinically significant gastric Helicobacter sp. infections including the fulfillment of Koch’s postulates

• Although virtually all adult ferrets have H. mustelae as part of the normal gastric flora, heavily colonized ferrets develop lymphoplasmacytic gastritis and lymphofollicular hyperplasia; also reports of possible cholangiohepatitis and cholangiocellular carcinoma associated with H. cholecystus-like bacteria
• Progression to neoplasia has not been documented in ferrets according to one source, but has been associated with pyloric adenocarcinoma +/- osseous metaplasia as well as MALT lymphoma according to another source

• In mice, Helicobacter sp. infections have emerged as a major group of gastrointestinal commensals with pathogenic potential, with many named species (H. hepaticus, H. bilis, H. muridarum, H. rodentium, H. typhlonius, H. ganmani, H. rappini, H. mastomyrinus, H. magdeburgensis, H. pullorum, H. muricola); H. hepaticus and H. rodentium appear to be the most prevalent; some isolates are nonpathogenic commensals, but most are capable of inducing proliferative typhlocolitis in immunodeficient mice typically due to mixed Helicobacter spp. infection
• The role of Helicobacter sp. in the development of gastritis or gastric ulceration in other species remains uncertain

 

PATHOGENESIS:

• Ulcers, suggested pathogenesis: Parietal cell trophic effects and increased gastric acid secretion (due to hypergastrinemia) à upper gastrointestinal mucosal damage à ulcer formation

• Hypergastrinemia: Abnormal gastrin hypersecretion due either to direct effect of the bacterium or antral inflammation; may be directly related to ulcerogenesis

• Bacterium has specific ligand interactions with host lipid receptors for mucosal epithelial cell attachment
• Virulence factors: 

• Urease is required for colonization: Urease breaks down urea in gastric juice to produce bicarbonate and ammonia, which neutralizes gastric acidity, and ammonia is toxic to epithelial cells; urease is strongly immunogenic itself à gastritis
• Flagellin molecules FlaA and FlaB: motility
• Superoxide dismutase and catalase: Protection from killing within neutrophil phagosomes

• Mice: Ingestion of contaminated feces (one report of transplacental infection in SCID/NCr mice) à colonization of the cecum, colon, +/- liver (within bile canaliculi) à persistent infection with long-term fecal shedding, decreased overall diversity of cecal microflora, proliferative typhlocolitis, hepatitis

• Pathogenesis of hepatitis and hepatocellular tumors is unknown, but colonization of the liver leads to induction of apoptosis and cellular proliferation
• The pathogenesis of proliferative typhlocolitis is proposed to be due to a strong Th1 mucosal immune response à production if IL-12, IFNγ, and TNF-α in the lamina propria à expression of keratinocyte growth factor (an epithelial mitogen)

 

TYPICAL CLINICAL FINDINGS:

• Ferrets: 

• Gastric or duodenal ulcers leading to vomiting, melena, chronic weight loss, lowered hematocrit, +/- acute gastric bleeding episodes; gastric disease severity increases as animals age
• H. mustelae-associated gastric adenocarcinoma: vomiting, anorexia, weight loss, +/- radiographically notable osseous metaplasia within the neoplasm

• Mice: 

• Immunocompetent mice develop minimal or no lesions 
• Immunodeficient mice develop hyperplastic typhlocolitis and hepatitis: wasting; mortality; unformed sticky, mucoid, or hemorrhagic feces; rectal prolapse (frequent nonspecific sign of colitis in mice)
• Hepatitis-susceptible mice strains include A/JCr, SCID/NCr, BALB/cANCr, C3H/HeNCr, and SJL/NCr, whereas B6 and B6C3F1 are hepatitis-resistant with H. hepaticus infection
• H. hepaticus (and probably other Helicobacter spp.) is associated with an increase in hepatocellular tumors in certain strains of mice, especially A strain mice, and has been shown to promote experimental chemical hepatocarcinogenesis

 

TYPICAL GROSS FINDINGS:

• Ferrets: Varies from no gross lesions to hyperemia and thickening of the gastric mucosa, occasional gastric erosions or ulcerations
• Mice: 

• Proliferative typhlocolitis: Rectal prolapse, segmental thickening of the cecum and colon
• Hepatitis: Variable; if present, randomly scattered white foci up to 4mm diameter

 

TYPICAL LIGHT MICROSCOPIC FINDINGS:

• Ferrets: Numerous spiral bacteria present in the superficial mucus layer, +/- inflammation in the adjacent mucosa, +/- epithelial hyperplasia; histopathologic changes coincide with bacterial location
• Mice: 

• Proliferative typhlocolitis: Marked mucosal crypt hyperplasia, mononuclear infiltrates in the lamina propria; the dysplastic and invasive nature of hyperplastic crypt epithelium, particularly when the mucosa is inflamed or eroded, may mimic neoplasia
• Hepatitis: Focal necrosis and mixed leukocytic infiltrates early in the disease; within a few months, marked hypertrophy and hyperplasia of Kupffer, Ito (stellate), and oval cells, increased hepatocyte mitotic activity, prominent bile ductule formation extending from portal regions, individual hepatocyte apoptosis, hepatocytomegaly, polykarya, lymphoplasmacytic inflammation
• Hepatocellular tumors: Pre-neoplastic findings include foci of cellular alteration including clear cell foci, basophilic cell foci, and eosinophilic cell foci

 

ULTRASTRUCTURAL FINDINGS:

• Ferrets: H. mustelae localizes within the gastric pits in very close association with epithelium alongside microvilli perpendicular to epithelial surfaces and sometimes penetrating epithelial cells within membrane-bound vesicles (endocytic vesicles), with little evidence of bacteria on the external surface or in overlying mucus layer; extensive loss of microvilli; adhesion pedestals are visible
• Helicobacter spp. are curved to spiral bacilli with variable numbers of peripolar flagella

 

ADDITIONAL DIAGNOSTIC TESTS:

• Demonstration of organisms using Warthin Starry 4.0, Genta, Steiner, or other silver stains; organisms located within crypts closely associated with epithelial surface in enteric lesions, and within bile canaliculi in hepatic lesions
• Bacterial culture with provisional diagnosis based on unique ability to produce large quantities of urease
• ELISA: Affected ferrets mount a significant, although nonprotective, IgG response
• PCR
• Cytologic findings:

• Highly prevalent in cytological samples of the gastric mucosa of cats and dogs (though the casual relationship with gastritis in these species is questionable)
• Often more easily identified with cytology relative to histology
• Can frequently be seen embedded within mucous
• Inflammation is often absent, but infection can be associated with nodular lymphocytic inflammation

 

DIFFERENTIAL DIAGNOSIS:

Mouse: 

• Proliferative typhlocolitis: Citrobacter rodentium, E. coli, enterotropic mouse hepatitis virus (coronavirus), nonspecific rectal prolapse
• Hepatitis: Salmonella, Proteus, C. piliforme, mouse hepatitis virus (coronavirus), ectromelia virus (orthopoxvirus), nonspecific hepatic necrosis and inflammation
• Hepatocellular karyomegaly and cytomegaly: These are nonspecific changes that may be associated with age or disease states and are thus not specific to Helicobacter sp. infection

 

COMPARATIVE PATHOLOGY:

• Camels: Recent report (Van de Weyer, J Comp Pathol 2024) of a case of a primary gastric adenoma in a 20 year old camel, helicobacter was found within the gastric pits and necrotic areas of C3 (though the causal relationship between helicobacter and gastric neoplasia was not assessed). 
• Cheetahs: H. acinonychis is hypothesized, but unproven, as a cause for gastritis in captive cheetahs; antibiotic administration improves clinical disease but does not eradicate infection, and wild cheetahs also are infected without gastritis; amyloidosis is a possible sequelae of chronic gastritis. 
  • Recent article (Zingre J Comp Pathol 2023) reported a case of fatal gastritis caused by Helicobacter pylori that occurred concurrently with enterocolitis caused by Campylobacter jejuni
• Dogs and cats: Prevalence of colonization is estimated at least 50% of animals, but the clinical significance is still uncertain, as it is seen in clinically healthy animals and those with gastritis; several studies claim that dogs or cats with Helicobacter infection have increased mucosal mononuclear leukocytes and mucosal lymphoid follicles and faster mucosal turnover
• Pigs: H. heilmannii has been proposed to be associated with gastric ulceration, but gastric ulcers occur in the squamous portion of the stomach in pigs; H. heilmanii has been shown to cause gastric ulceration in the squamous portion of the stomach in gnotobiotic pigs fed a ration high in carbohydrates, but more consistent findings indicate that finely ground rations are ulcerogenic in pigs over other factors
• Humans: H. pylori is a common cause of acute and chronic gastritis, is the leading cause of chronic peptic ulcers that may progress to mucosal atrophy, and significantly increases the risk of gastric carcinoma and MALT lymphoma; infection and inflammation can extend to the duodenum
• Nonhuman primates: H. pylori and a few other Helicobacter spp. can affect NHPs but have never been proven to cause disease, although H. pylori is associated with chronic active gastritis and gastric erosions; infection can persist for years 
  • H. cinaedi (an enterohepatic helicobacter) has been associated with chronic diarrhea in a rhesus macaque with associated inflammation of the lower gastrointestinal tract (colon, liver, mesenteric lymph node)
• Hamsters: A variety of Helicobacter spp. colonize the gastrointestinal tracts of hamsters and are typically subclinical, but disease may arise in aged hamsters
  • H. aurati was isolated from the stomach and cecum of adult hamsters with gastritis; an invasive adenocarcinoma at the pyloric-duodenal junction was found at the site of H. aurati-associated inflammation; serve as an animal model for IBD and IBD-related neoplasia
  • H. cholecystis and H. bilis infection have been associated with bile ductular hyperplasia and fibrosis +/- nodular hepatocellular regeneration (hepatic cirrhosis); hepatic portal fibrosis is a common sequel of enterohepatic Helicobacter infections
  • Spontaneous proliferative and dysplastic typhlocolitis has been associated with Helicobacter spp. infection, typically H. bilis; a round cell sarcoma and histiocytic sarcoma have been identified at the ileocecocolic junction of infected hamsters
• Rats: Proliferative and ulcerative typhlitis, colitis, and proctitis have been observed in athymic nude rats infected with H. bilis, but there is no evidence that any of the several Helicobacter spp. identified in rats produce disease in immunocompetent rats; prevalence estimated at 20% in laboratory colonies
• Gerbils: Mongolian gerbils develop chronic gastritis, gastric ulcers, intestinal metaplasia, and gastric adenocarcinomas following experimental infection with H. pylori and so are an established animal model for human disease; otherwise, enteric infections with several Helicobacter spp. have been identified without any associated lesions
• Rabbits: Rabbits from pet, laboratory, and commercial sources all tested positive for Helicobacter spp., and mild inflammatory lesions were present in the gastric mucosa of some infected rabbits; the prevalence and significance is unknown
• Dolphins: Helicobacter cetorum has been identified multiple species with lymphoplasmacytic gastritis with ulceration but causality is still uncertain 
• Sea otters: H. enhydrae sp. nov has been spatially associated with gastric lesions, but causality is not confirmed

 

REFERENCES:

1. Barthold SW, Griffey SM, Percy DH. Pathology of Laboratory Rodents and Rabbits. 4th ed. Ames, IA: Blackwell Publishing; 2016: 7, 55-58, 101, 113, 139, 182-183, 196, 203, 279.
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4. Hostetter, SJ. Oral Cavity, Gastointestinal Tract, and Associated Structures. In: Raskin RE, Meyer DM. Canine and Feline Cytology, 4^th Ed. St. Louis, MO:Elsevier. 2023:310 - 304.
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12. Van de Weyer Y, Bentley C, Orlowska A, Barbon AR, Chantrey J. Gastric adenocarcinoma with carcinomatosis and degenerative joint disease of the xiphisternum in a Bactrian camel (Camelus bactrianus). J Comp Pathol. 2024;214:7-11. 
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14. Zingre T, Bagatella S, Wenker C, et al. Fatal gastritis and enterocolitis due to concurrent Helicobacter pylori and Campylobacter jejuni infection in a captive cheetah (Acinonyx jubatus). J Comp Pathol. 2023;201:81-86.

 

 

 

 

 

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