JPC SYSTEMIC PATHOLOGY

DIGESTIVE SYSTEM

September 2024

D-B13

 

SIGNALMENT (JPC #1782684): 2-day-old piglet

 

HISTORY: This animal had diarrhea. There was 100% morbidity and 90% mortality in affected litters.

 

HISTOPATHOLOGIC DESCRIPTION: Small intestine (multiple sections): Multifocally and frequently, there are aggregates of basophilic, 1x2 µm bacilli carpeting approximately 60% of the mucosal epithelial apical brush border along villi and crypts. Multifocally, intestinal villi are mildly blunted and enterocytes are occasionally sloughed/lost, swollen and vacuolated (degenerate), or shrunken with a scant amount of hypereosinophilic cytoplasm and a pyknotic nucleus (single cell death), and there are occasional neutrophils transmigrating the mucosal epithelium. The lamina propria is multifocally infiltrated by low to moderate numbers of neutrophils, fewer lymphocytes, mild to moderate hemorrhage, fibrin, and edema. Small caliber blood vessels are expanded up to two times normal (congestion). Occasionally, multifocal crypt lumina contain necrotic cellular debris and few neutrophils (crypt abscesses). The Peyer’s patches (GALT) within the submucosa of the ileum contain active germinal centers, up to 2mm diameter, which contain numerous tingible body macrophages (reactive lymphoid hyperplasia and lymphocytolysis). 

 

MORPHOLOGIC DIAGNOSIS: Small intestine, mucosa: Abundant apical surface-associated bacilli, multifocal to coalescing, with mild neutrophilic enteritis, few crypt abscesses, and mild GALT hyperplasia, porcine.

ETIOLOGIC DIAGNOSIS: Enteric colibacillosis

 

CAUSE: Enterotoxigenic Escherichia coli

 

CONDITION: Enterotoxigenic colibacillosis

 

SYNONYMS: ETEC, white scours, neonatal diarrhea, post-weaning diarrhea, young pig diarrhea, coliform gastroenteritis, baby pig diarrhea, coliform scours

 

GENERAL DISCUSSION: 

• Ubiquitous gram-negative, facultative anaerobe, non-spore-forming, oxidase negative, bacillus from the Enterobacteriaceae family; non-virulent strains are considered normal flora
• Major cause of neonatal diarrhea in calves, pigs, and lambs; and weanling pigs
• Risk factors include genetic susceptibility, failure of passive transfer, early weaning, poor nutritional plane, environmental contamination, low ambient temperature, or stress (environmental or physiological) 
• Diagnosis is based on clinical signs, minimal gross lesions, and basophilic bacilli lining the surface of relatively histologically normal small intestines 
• 4 main types of antigens on which E. coli serotypes are based:
  • O antigen (somatic; lipopolysaccharide)-> determines serogroup
  • K antigen (capsular): may enhance ability of bacteria to colonize intestines
  • H antigen (flagellar)
  • F antigen (fimbral): important in enteric colonization; some adhesins are associated with certain serotypes and are relatively host specific

 

PATHOGENESIS:

• Enterotoxigenic E. coli (ETEC): Ingestion of virulent bacteria > adherence to small intestinal enterocytes , mid-jejunum to ileum, (does not invade enterocyte) > colonization of mucus on enterocyte (evades peristalsis) > toxin production causes non-structural alteration of cell membrane and fluid transport system > net secretion of fluid and electrolytes from the systemic circulation into the lumen of the gut (metabolic dysfunction) > secretory diarrhea > dehydration > metabolic acidosis > shock > death 
• Virulence factors:
  • Fimbriae / pili (colonization factor antigens): Made up as a polymer of protein subunits; promote colonization of gut; coded by plasmids; antigenically distinct; over 20 types have been identified
    • Fimbriae commonly found in pigs: F4 (K88), F5 (K99), F6 (987P), F18, F41
    • Genetic resistance is related to age-related susceptibility (the presence or absence of the appropriate receptors in the small intestine; i.e. F18 causes post-weaning diarrhea, but not neonatal) and inherited resistance (i.e. some pigs lack epithelial cell receptors for F4)
  • Enterotoxin: Heat-labile toxin (LT; cAMP) or heat-stable toxin (STa, STb, LT, EAST1; cGMP) coded by plasmids, open chloride channels that irreversibly move chloride, water, and sodium into the lumen (secretory diarrhea) causing death in less than 24 hours from dehydration and electrolyte imbalance if untreated
    • STb also increases intracellular calcium, prostaglandin E2, and 5HT.
    • EAST1: Enteroaggregative E. Coli heat stable toxin
    • ETEC that causes neonatal diarrhea usually only product STa while in post-weaning diarrhea usually produces one or more enterotoxins

 

TYPICAL CLINICAL FINDINGS:

• Neonatal (0-4 days old) and post-weaning (2-4 weeks after) piglets with watery alkaline diarrhea; may lose up to 40% of total body weight due to dehydration, resulting in flaccid and “tucked up” abdominal musculature, enophthalmos, and bluish-gray skin
• Morbidity is extremely variable (average 30-40%; up to 80%); mortality can reach 70% (neonatal) or 25% (post-weaning) in affected litters

 

TYPICAL GROSS FINDINGS: 

 

TYPICAL LIGHT MICROSCOPIC FINDINGS:

• Bacteria adherent to the mucosal epithelial cells of the jejunum and ileum; found on brush borders of villi and/or crypts of Lieberkühn
• Typically no damage to enterocytes with mild to no epithelial cell loss
• Sometimes vascular congestion, mildly increased neutrophils/macrophages in the lamina propria, and villous atrophy

 

ULTRASTRUCTURAL FINDINGS: 

 

ADDITIONAL DIAGNOSTICS:

• Real time PCR for pathogenic gene sequences (virulence plasmids and serogroup identification)
• Indirect fluorescent antibody (IFA) for pilus antigens
• Bacteria highlighted with Giemsa stain or IHC
• Bacterial culture 

 

DIFFERENTIAL DIAGNOSIS:

• Enteropathogenic E. coli (EPEC): 1-4 days old and 2-3 weeks post-weaning; watery yellowish diarrhea, dehydration
• Rotavirus (type A, B, C): 1 day to 7 weeks (most common 2-3 weeks old); watery to pasty diarrhea (may be subclinical); enterocyte sloughing with replacement by cuboidal to squamoid cells and villus blunting/fusion. 
• Clostridium perfringens type C: 1-14 days old; hemorrhagic or watery diarrhea and sudden death; segmental necrohemorrhagic enteritits in proximal 2/3 of small intestine. 
• C. perfringens type A: 3-28 days; mild creamy diarrhea
• C. difficile: 5-21 days; creamy diarrhea and dehydration; necrotizing colitis and mesocolonic edema
• Cryptospirodium spp: 3 days to weaning; mild to moderate yellowish diarrhea
• Coccidiosis (Cystoisospora suis): 5-21 days; watery yellowish diarrhea; fibrinonecrotic pseudomembrane without blood; intracytoplasmic apicomplexan organisms
• Coronavirus (TGE, PED, PDCoV)
  • Transmissible gastroenteritits virus (TGE): All ages (most severe <3 weeks old); severe diarrhea, vomiting, high mortality (100%); severe diffuse atrophic jejunitits and ileitis  
  • Porcine epidemic diarrhea virus (PED): All ages (severe in piglets <1 week); watery diarrhea, vomiting; diffuse atrophic jejunitits and ileitis 
  • Porcine deltacoronavirus (PDCoV): All ages (worst in suckling pigs); watery diarrhea, vomiting; diffuse atrophic jejunitits/ileitis (less severe than TGE/PED)
• Strongyloides sp: 10 days+; diarrhea and ill-thrift; villous atrophy, granulomatous enteritis
   
   

COMPARATIVE PATHOLOGY:

Pathogenic E. coli Subtypes:

• ETEC (enterotoxigenic, F18ac, F4 (K88), East 1): Important common cause of secretory diarrhea in neonatal (and post weaning) piglets, calves, lambs; lesser extent in dogs, horses . 
• EPEC (enteropathogenic, intimin): Rabbits, pigs, dogs, cats, NHPs (rhesus macaques, marmosets, cotton-top tamarins 
  • Attaches to enterocytes; does not invade them; associated with villus atrophy
  • Uncommon in domestic animals
  • Pathogenicity island is LEE (locus for enterocyte effacement)
• EHEC (enterohemorrhagic): Rabbits, dogs, calves, pigs, NHPs; ruminants are reservoirs. Prefers ileum and colon.  
  1. STEC (shigatoxin prod. E. coli): O157:H7 Human pathogen and cause of hemolytic uremic syndrome, hemorrhagic colitis, diarrhea. Carried by cattle and has been isolated from various types of birds (pigeons) and poultry products, however not a significant infectious source. STx 1 and STx 2. Experimentally infects Dutch Belted and New Zealand White rabbits, seen in wild rabbits naturally as well. 
• EDEC (edema disease, O138, O139, O141) : pigs 6-14 weeks (edema disease, gut edema)
  • Usually F18ab; endotoxin (STx2e) à systemic vascular endothelial injury
  • Edema in gastric mucosa, eyelids, spiral colon; encephalomalacia, cerebrospinal angiopathy 
  • Alpha hemolytic 
  • Affects fast growing pigs
  • Gross findings: Edema of stomach wall and mesocolon with stomach full of feed and formed feces in intestine.
• EIEC (enteroinvasive): Lab animals, occasionally cattle/pigs 
  • Internalized by enterocyte, multiplication, and disseminated through body
  • Erosion and ulceration seen at villi
  • Poorly documented in domestic animals
    • Suspected causative agent in histiocytic and ulcerative colitis in Boxers and French Bulldogs; IBD in people
  • A cause of septicemic colibacillosis 
  • Strain 1056, 1126, and 4165 isolated in hamsters 
  • Affects primarily the colon in NHPs
• ExPEC:
  • UPEC: Dogs, cats, pigs 
    • Coliseptemia, polyserositis, urogenital infections
  • APEC: Turkeys, chickens, commercial ducks
    • Typically a localized or systemic secondary disease 
    • Has a virulence plasmid and MDR plasmid that can impact human health
    • Less toxigenic compared to mammalian strains 
    • Local colibacillosis: Enteritis, omphalitis, cellulitis, salpingitis, orchitis, venereal colibacillosis, swollen head syndrome, serositis 
    • O1, O2, O18, O35, O26, O78, K1, K80 common pathogens
    • Colisepticemia: pericarditits, myocarditis, air sacculitis, respiratory collapse, etc
    • Hemorrhagic septicemia
    • Coligranulomas (Hjarre’s disease): liver, ceca, duodenum, mesentery
  • NMEC: Human neonatal meningitis, virulence is attachment and invasion. 

 

REFRENCES:

1. Abdul-Aziz, T. Fletcher OJ. Cardiovascular System. In: Abdul-Aziz T, Fletcher OJ, Barns HJ. Avian Histopathology. 4th Ed. Madison, WI: American Association of Avian Pathologist: 2016: 146, 168, 188
2. Abdul-Aziz, T. Fletcher OJ. Hepatobiliary System. In: Abdul-Aziz T, Fletcher OJ, Barns HJ. Avian Histopathology. 4th Ed. Madison, WI: American Association of Avian Pathologist: 2016: 358, 391, 393.
3. Abee CR, Mansfield K, Tardif S, Morris T. Nonhuman Primates in Biomedical Research: Volume 2: Diseases. 2^nd ed. San Diego, CA: Elsevier; 2012: 126-127, 454. 
4. Barthold SW, Griffey SM, Percy DH. Pathology of Laboratory Rodents and Rabbits. 4th ed. Ames, IA: Wiley Blackwell; 2016: 54-55. 182,226, 277-278 .
5. Constable PD, Hinchcliff KW, Done SH, Grunberg W. Diseases of the Alimentary Tract: Nonruminant. In: Constable PD, Hinchcliff KW, Done SH, Grunberg W. Veterinary Medicine, A Textbook of the Diseases of Cattle, Sheep, Pigs, Goats and Horses. 11th ed. St. Louis, MO: Elsevier; 2017:311-319, 1879-1887.
6. Crespo R, Franca MS, Fenton H, Shivaprasad HL. Galliformes and Colubriformes. In: Terio KA, McAloose D, St. Leger J, eds. Pathology of Wildlife and Zoo Animals. London, UK: Academic Press; 2018:753. 
7. Delaney MA, Treuting PM, Rothenburger JL. Lagomorpha. In: Terio KA, McAloose D, St. Leger J, eds. Pathology of Wildlife and Zoo Animals. London, UK: Academic Press; 2018:494. 
8. Dunn, PA, Boulianne M. Diseases of the Duck (table). In: Boulianne M ed. Avian Disease Manual. 8^th ed. Madison, WI: Omnipress; 2019: 183.
9. Fletcher OJ. Abdul-Aziz, T. Respiratory System. In: Abdul-Aziz T, Fletcher OJ, Barns HJ. Avian Histopathology. 4th Ed. Madison, WI: American Association of Avian Pathologist: 2016: 198, 210, 241-243, 263-265.
10. Fletcher OJ. Abdul-Aziz, T. Alimentary System. In: Abdul-Aziz T, Fletcher OJ, Barns HJ. Avian Histopathology. 4th Ed. Madison, WI: American Association of Avian Pathologist: 2016: 275, 323-324,
11. Fulton RM, Boulianne M. Bacterial Diseases. In: Boulianne M ed. Avian Disease Manual. 8^th ed. Madison, WI: Omnipress; 2019:77-81.
12. Gelberg HB, Spagnoli ST. Alimentary system and the peritoneum, omentum, mesentery, and peritoneal cavity. In: Zachary JF, ed. Pathologic Basis of Veterinary Disease. 7th ed. St. Louis, MO: Elsevier; 2022:448-450.
13. Koenig A. Gram-negative bacterial infections. In: Greene CE, ed. Infectious Diseases of the Dog and Cat. 4th ed. St Louis, MO: W.B. Saunders Company; 2012:351-353
14. Matz-Rensing K, Lowenstine LJ. New World and Old World Monkeys. In: Terio KA, McAloose D, St. Leger J, eds. Pathology of Wildlife and Zoo Animals. London, UK: Academic Press; 2018:357-358,. 
15. Myers, EA. Sander JE. Appendix. In: Boulianne M ed. Avian Disease Manual. 8th ed. Madison, WI: Omnipress; 2019: 193, 197.
16. Schmidt R, Reavill DR, Phalen DN. Pathology of Pet and Aviary Birds. 2nd ed. Ames, IA: John Wiley & Sons, Inc.; 2015: 78-79, 159, 284. 
17. Uzal FA, Plattner BL, Hostetter JM. Alimentary system. In: Maxie MG, ed. Jubb, Kennedy and Palmer’s Pathology of Domestic Animals. Vol 2. 6th ed. Philadelphia, PA: Elsevier Saunders; 2016:112, 158-167.
18. Wallner-Pendleton E. Diseases of the Game Birds (table). In: Boulianne M ed. Avian Disease Manual. 8^th ed. Madison, WI: Omnipress; 2019:187.
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E. coli subtypes	Virulence Factors	Examples
Enteropathogenic (EPEC) / Attaching and Effacing (AEEC) Osmotic diarrhea Acute inflammatory response (IL 18) Structural cellular alterations (attaching and effacing); does NOT invade enterocytes Enterocyte junction leakage Villus atrophy malabsorption Gross: granular and rough mucosa; hemorrhage and fibrin Ultrastructure: pedestal formation beneath attachment at microvillus of enterocytes and gallbladder epithelium. Loss of microvilli architecture. Granulomas.	Locus for enterocyte effacement (LEE): pathogenicity island that controls attaching-effacing trait Fimbriae: P and S Esp A/B/D- bacterial proteins exported via type III secretion system into the host cell; Esp-b & -D create pore in host membrane to introduce Tir (translocated-intimin-receptor protein) Tir “focuses” actinà forming “pedestal” EPEC adherence factor, Intimin (bacterial outer membrane adhesin protein) and bundle-forming pilus (bfp) binding to Tir (in host membrane) = Intimate attachment and effacement. Coded by eae gene NO TOXINS Affects small intestine and colon	Generally uncommon in domestic animals Diarrhea in rabbits, calves, pigs, lambs, dogs and humans (link w/ Crohn’s)  Hemorrhagic colitis 2-5 weeks age in calves Most often seen in calves with other enteropathogens such as Cryptosporidium parvum, ETEC, coronavirus, BVD, coccidian Found in turkeys with PEMS (poult enteritis mortality syndrome) as secondary bacterial agent Infantile diarrhea in NHPs and idiopathic colitis
Enterohemorrhagic (EHEC, VTEC: verotoxin-producing, STEC: Shiga toxin-producing) Acute inflammatory response -> hemolytic uremic syndrome Structural cellular alterations; invades enterocytes Targets colon- hemorrhagic colitis Gross: granular and rough mucosa; hemorrhage and fibrin Ultrastructure: pedestal formation beneath attachment (w/o Tir)	Shiga toxin or Shiga-like toxins (coded by bacteriophages) Bind the host cell receptor glycolip receptor = globotriaosylceramide (Gb3)à inhibition of protein synthesis with subsequent necrosis Primarily affects intestinal epithelium and vascular endothelium, due to the presence of Gb3 receptors in these tissues Mucosal loss à lamina propria exposure à endotoxin (LPS) uptake Acid resistant Intimin Hemorrhagic coli pilus, important in adherence and biofilm formation  Hemolysin (A-B toxin) increases Fe+ availability to microbes	Lab animals ( coliform typhlocolitis ), cattle, pigs, humans (O157:H7) Fibrinohemorrhagic enterocolitis in calves (under 4-weeks old) Major cause of foodborne illness in humans Hemolytic uremic syndrome (humans) Greyhound: cutaneous/renal glomerular vasculopathy – associated with 0157:H7
Enterotoxigenic (ETEC) Secretory diarrhea Non-inflammatory or mild neutrophilic inflammation Non-structural cellular alterations; does NOT invade enterocytes No gross lesions Dysfunction of enterocyte ion pumps	Fimbriae and flagella and adhesion molecule East1 (EtpA) Plasmid regulated enterotoxins Heat-labile toxins: LTIà similar to cholera toxin; inhibits Na+/Cl- absorption LTIIà cAMP pathwayà irreversible secretion of electrolytes (Cl- esp)/H2O in gutà secretory diarrhea Heat-stable toxins: STaà inhibition of Na/Cl co-transport and H2O absorption via ↑ in cGMP; STb (primarily associated with ETEC in pigs)à promotes secretion by stimulating prostaglandin E2 and 5-hydroxytrypamine	<4 days old or post weaning in pigs (up to 3 week old).  Secretory diarrhea in neonatal farm animals (calves, piglets, lambs, kids, foals) and humans (“travelers diarrhea”) Major cause of calf diarrhea <3 days. F5 (K99) common virulence factor. Common in pigs and calves
Enteroinvasive (EIEC, septicemic) Enterocyte internalization à sepsis Mixed bacterial infection common	Obligate intracellular bacteria. Invades intestinal enterocytes (through M cells), replicate in Macrophages, and disseminate throughout body Gains entry through the respiratory system, oral cavity, or umbilicus	Newborn calves, lambs, and occasionally foals that have received insufficient colostrum Only confirmed experimentally in neonatal swine Results in septicemia with fibrinous arthritis, ophthalmitis, serositis, meningitis, white-spotted kidneys (cortical abscesses)
Edema disease, enterotoxemic colibacillosis	Adhesion involved in diffuse adherence (AIDA): an adhesion that works with F18ab Enteric colonization with Shiga-toxin ( or verotoxin 2e; Stx2e) producing E. coli and classic enterotoxemia à targets vascular endothelium	Pigs: few weeks after weaning Endothelial injury of arterioles and arteries -> edema; neurologic signs due to brain edema, edema also in eyelids, gastric sub mucosa, gallbladder, and mesentery of spiral colon Can affect humans
Septicemic colibacillosis Generalized septicemia: enters via navel, upper respiratory tract, tonsil, intestine Enteritis not always present	Colicin V (plasmid): encodes aerobactin: siderophore that helps survive in low iron environments, outer membrane protein to combat bactericidial components of serum, capsule to impede phagocytosis Type 1 and P fimbriae Causes a systemic inflammatory response syndrome (SIRS)	Calves <4 days; less common in others species Neonates most affected with serovar O78 (calves, piglets, lambs) Lambs 1-2 days or 3-8 weeks Necrotoxigenic E coli 2 (NTEC2) affects calves and lambs  Multisystemic disease in poultry
Exraintestinal pathogenic E. coli (ExPEC)	Fimbral adhesins for attachment Non fimbral adhesin Intimin -> surface of enterocytes -> attaching and effacing lesion Cytotoxins and hemolysis -> tissue necrosis Siderophore receptors -> sequesters iron Invasins (endothelium) and protectins (resist complements) Virulent plasmids and MRD plasmids (APEC) Binding w/ fibronectin and lamenectin (BM components) (APEC)	Shed by dogs and cats Endocarditis (tricuspid) UTI in dogs  Aerolized bacteria in poultry-> fulminating necrohemorrhagic pleuropneumonia, septicemia, urogenital infections, meningitis

 

 

 

 

 

 

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