JPC SYSTEMIC PATHOLOGY

DIGESTIVE SYSTEM

September 2024

D-B12

 

SIGNALMENT (JPC #1946537): Tissue from a 4-year-old yellow-naped Amazon parrot 

 

HISTORY: The bird became progressively docile, passive, and depressed.

 

MICROSCOPIC DESCRIPTION: Liver: Multifocally there are random, irregularly round, variably sized (up to 1 mm diameter) areas of lytic necrosis characterized by loss of hepatic architecture with replacement by eosinophilic cellular and karyorrhectic debris admixed with heterophils and hemorrhage, fibrin, and edema and surrounded by multinucleated giant cells (foreign body and Langhan’s type), epithelioid macrophages, and fewer lymphocytes and plasma cells. Multifocally, there are also random foci of coagulative necrosis characterized by retention of architecture with loss of differential staining. Multifocally, few random hepatocytes and Kupffer cells contain intracytoplasmic, amphophilic to gray, indistinct, granular, coccoid bacteria. Hepatocytes surrounding areas of necrosis often are swollen with vacuolated cytoplasm (degeneration) and are dissociated from hepatic cords. Diffusely, sinusoids are moderately expanded by macrophages, heterophils, and lymphocytes. Multifocally hepatocytes are mildly vacuolated and contain small aggregates of brown, granular pigment (bile, hemosiderin, and/or lipofuscin). There is a mild increase in bile duct profiles; bile ducts are multifocally mildly ectatic and lined by hyperplastic epithelium characterized by piling up, mild anisokaryosis, and increased mitotic activity (regeneration). Portal areas are expanded by increased clear space (edema) and contain dilated lymphatic vessels. 

 

MORPHOLOGIC DIAGNOSIS: Liver: Hepatitis, necrotizing, heterophilic, and granulomatous, multifocal to coalescing, moderate, with mild bile duct hyperplasia and intracellular bacteria, yellow-naped Amazon parrot (Ochrocephala auropalliate), avian.

 

ETIOLOGY: Chlamydia psittaci 

 

ETIOLOGIC DIAGNOSIS: Chlamydial hepatitis

 

CONDITION: Avian chlamydiosis (birds); psittacosis (humans)

 

GENERAL:

• Gram negative obligate intracellular bacteria (related to rickettsia)
• Synonyms: Parrot Fever, Ornithosis (non-psittacines)
• The family Chlamydiaceae contains a single genus: Chlamydia, which includes 11 species (*=have been isolated from birds)

• C. muridarum (mouse, hamster)
• C. suis (swine)
• C. trachomatis (humans)
• C. abortus* (ruminant) 
• C. caviae (Guinea pig)
• C. felis (feline)
• C. pecorum (ruminants, swine, koalas)
• C. pneumoniae (human, marsupial, and amphibious origin)
• C. psittaci* (avian serovar)
• C. avium*
• C. gallinacean*

• C. psittaci has 8 fairly host specific serovars: 6 in birds and 2 in mammals 

• Serovars A and F – Psittaciformes (especially cockatiels & budgerigars)
• Serovar B – Columbiformes (pigeons, doves), galliformes (broiler chickens) *low virulence
• Serovar C – Anseriformes (ducks, geese, swans)
• Serovar D – Galliformes (turkeys, broiler chickens) *high virulence

Serovar E – Columbiformes and Galliformes (pigeons, turkeys) 

• Zoonotic and reportable - Take precautions during necropsy; dead birds should be immersed in disinfectant due to highly infectious nasal/fecal secretions
• Unique morphology and biphasic life cycle with four morphologically distinct forms

• Elementary body (EB): Infectious form that enters the cell
• Reticulate body (RB): Intracellular, metabolically active, replicating form that synthesizes DNA, RNA, and protein, and divides by binary fission
• Intermediate body (IB): Morphologic characteristic between EB and RB
• Persistent aberrant body (PAB): Non-replicating bodies emerge during times of stress (i.e. antibiotics), revert to RB when stressor removed, associated with long-term infection

• Organisms have tropism for coated pits on epithelial cells of the respiratory tract

 

PATHOGENESIS:

• Inhalation or ingestion of contaminated feather dust or feces > multiply in lung, air sacs and pericardial sac within 4 hours > bacteremia within 48 hours > portals of exit (cloaca & nasal turbinates)
• Reproductive cycle: EBs attach to host cell membrane at coated pit > internalization into host cell via invagination of the host cell membrane > inhibition of phagosome - lysosome fusion > differentiation into RB > replication by binary fission > reorganization, through IBs, into new EBs > release from host cell (cell lysis or exocytosis)
• Depend on eukaryotic cells for energy (therefore obligate intracellular)

• Mitochondria are often found in juxtaposition to the C. psittaci inclusion
• RBs may parasitize mitochondrial ATP via a chlamydial ATP-ADP translocase

 

TYPICAL CLINICAL FINDINGS: (in birds)

• Diarrhea and excretion of green to yellow-green urates
• Severely affected birds may become anorectic and produce sparse, dark green droppings, followed by emaciation, dehydration, and death
• Recurrent episodes of illness culminating in chronic debilitation
• Subclinical infections common, especially in cockatiels (Schmidt 201.)

 

TYPICAL GROSS FINDINGS:

• Hepatomegaly and splenomegaly +/- necrotic foci; spleen may be congested or pale (due to increased histiocytes and plasma cells)
• Fibrinous air sacculitis, pericarditis and peritonitis
• Systemic disease: subcutaneous hemorrhage, meningitis, conjunctivitis, pancreatitis, orchitis, epididymitis, enterocolitis with diffuse mucosal necrosis
• Associated with atherosclerosis in psittacines

 

TYPICAL MICROSCOPIC FINDINGS:

• Liver

• Acute: Multifocal coagulation/lytic necrosis 
• Chronic: Granulomas, portal fibrosis, bile duct hyperplasia
• Kupffer cell hyperplasia
• Bacteria within hepatocytes and/or macrophages

• Spleen: Multifocal necrosis, histiocytosis, lymphoid depletion, plasmacytosis, 
• Air sacs: Fibrinous exudate with heterophils and macrophages, some containing bacteria
• Inclusion body-like microcolonies may be seen in affected cells of many organs, particularly in serosal membranes; may be better visualized through impression smear of fresh tissues

 

ULTRASTRUCTURE:

• Elementary body (EB): Spherical, 0.2-0.3 µm body composed of a highly electron dense nucleoid at the periphery, clearly separated from the electron dense cytoplasm
• Reticulate body (RB): 0.5-2.0 µm, binary fission characterized by “hour-glass” profiles
• Intermediate body (IB): 0.3-1.0 µm diameter bodies with a central electron dense core, radially arranged nucleoid fibers surrounding the core and tightly packed cytoplasmic granules at the periphery
• Persistent aberrant body (PAB): Small inhomogeneous reticulate bodies 
• Inside the host cell, organisms are present within endosomes

 

ADDITIONAL DIAGNOSTIC TESTS:

• Nucleic acid amplification tests (real time PCR, DNA microarray-based detection, DNA sequencing) preferred method; PCR of host-specific serovar may help pinpoint source of infection
• Isolation of the organism from tissue, feces, or swabs still most common test, but some serovars difficult to grow and hazardous to laboratory personnel (BSL3)
• Special stains (cytology or histology): Giemsa (dark purple), Gimenez, modified Gimenez (PVK stain) red, Warthin-Starry, or Macchiavello; Gram stain of little value because the chlamydial cell wall lacks peptidoglycan
• Visible on phase contrast and dark-field illumination microscopy
• EM, immunofluorescence, ELISA, immunohistochemistry, strain-specific monoclonal antibodies available
• Serology limited use due to high prevalence in certain populations and long term persistence
• Cytologic findings: In cases of conjunctivitis, can be found as variably sized, discrete basophilic to magenta bodies attached to epithelial cells, that are eventually internalized to form larger intracytoplasmic reticulate bodies and elementary bodies

 

DIFFERENTIAL DIAGNOSIS:

• Hepatic necrosis in psittacine birds:

• Pacheco’s disease (alphaherpesvirus): Intranuclear hepatocellular inclusions
• Polyomavirus (Budgerigar Fledging Disease): Glassy intranuclear inclusions
• Salmonellosis (Salmonella sp.) and colibacillosis (E. coli): Hepatocellular necrosis with necrogranulomas
• Reoviral hepatitis: Hepatocellular necrosis without inclusion bodies
• Lead toxicosis: Eosinophilic, acid-fast, intranuclear inclusions
• Mycobacteriosis (MAIC): Acid-fast, intracellular bacilli

• In poultry, similar signs and lesions may be caused by the following diseases:

• Fowl cholera (Pasteurella multocida)
• Mycoplasmosis: M. gallisepticum (chronic respiratory disease/infectious sinusitis of turkeys); M. meleagridis (air sacculitis)
• Colibacillosis (E. coli)
• Salmonellosis - S. Typhimurium (paratyphoid); S. Pullorum (Pullorum disease); S. Gallinarum (fowl typhoid)
• Adenovirus: Hemorrhagic enteritis of turkeys

 

COMPARATIVE PATHOLOGY:

• Humans: Zoonotic disease; influenza-like symptoms; pneumonia
• Ruminants: C. abortus: sporadic bovine encephalomyelitis, pneumonia, polyarthritis of calves, conjunctivitis, and sporadic abortions, including ovine enzootic abortion (necrotizing placentitis in ewe, hepatic necrosis in fetus), ewes only abort once but can remain as carriers (infected 5-6 weeks abort late gestation, infected after 5-6 weeks abort subsequent pregnancy) 

• Has been reported to cause bronchopneumonia in experimentally challenged calves

• Cats: C. felis, persistent conjunctivitis and rhinitis (not feline pneumonitis like previously overstated as a lung pathogen); one case of fatal C. psittaci infection in kitten
• Guinea pigs: C. caviae, Guinea Pig Inclusion Conjunctivitis (GPIC); mainly in juveniles can cause conjunctivitis, bronchitis, pneumonia and genital tract infections. 

• Histologically: heterophilic keratoconjunctivitis and uveitis, pneumonia has cranioventral pattern and is heterophilic 

• Dogs: Keratoconjunctivitis, bronchopneumonia, and reproductive problems (limited number of case reports)
• Raptors (Hawkes, falcons, eagles, kestrels and owls): Fibrinous pericarditis,   airsacculitis, pleuritis, pneumonia, and sometimes meningoencephalitis in red-tailed hawks. hepatosplenomegaly, fibrinous exudate in the pericardium, air sacs, or capsular surface of the liver 
• Horses: C. psittaci identified in cases of abortion, higher percentage (83%) for which

C. psittaci was confirmed as the etiological agent than in previous reports in equine reproductive loss (14%) (Begg, Vet Pathol, 2022) premature death of foals in Australia, one case in Switzerland (Bauman, JVDI, 2020)

• Chlamydia pecorum: Causes enteritis in calves <10 days old (see N-B05 for encephalitis manifestations of sporadic bovine encephalomyelitis)

• Intestinal tract is the natural habitat and may be the portal of entry for systemic infections resulting in hepatitis, arthritis, encephalitis, and pneumonia
• Enteritis in calves – Even “asymptomatic” animals have a 48% reduction in growth rate; these animals had conjunctival hyperemia, increased serum globulin, and decreased plasma albumin and insulin-like growth factor-1 suggesting that these asymptomatic infections may be the reason why in some calves feed-additive antibiotics aid in growth promotion. Experimental inoculation in newborn calves develop fever and diarrhea within 24 hours and become moribund within 4-5 days. 
• Pathogenesis: Ingestion > adsorbs to brush border of enterocytes at the tips of intestinal villi (these cells are in G1 which is what Chlamydia requires for multiplication) > enters the cell via pinocytosis > multiplication in the supranuclear region > host cell degenerates > released into gut lumen and lamina propria > infects endothelial cells of lacteals, Goblet cells, enterochromaffin cells, and macrophages > systemic spread 
• Gross lesions are most common in the terminal ileum; mucosal edema, congestion, petechiae, +/- ulceration
• Histologically, chlamydial inclusions are demonstrated within host cell cytoplasm using: Giemsa, Jimenez, Macchiavello or immunoperoxidase staining; crypts may be dilated with inflammatory exudate; lymphoid follicles in Peyer’s patches are necrotic

• Chlamydia pecorum: Most common infectious disease in Koalas, associated with urogenital inflammatory disease and conjunctivitis

• Transmitted sexually, during birth, monocytes in milk, or during pap-feeding
• Gross findings: Females-pyometra, metritis, cystitis, and/or ascending pyelonephritis. Males- prostatitis and, rarely, orchitis
• Histologically: Villous epithelial hypertrophy and hyperplasia, fibrosis, and granulation tissue formation, and squamous metaplasia. Lymphocytic +/-histiocytic inflammation most severe in the lamina propria-superficial submucosa. In active cases with greater numbers of inclusions, transepithelial migration of neutrophils and lymphocytes is common, plasma cells predominate in chronic, inactive cases. Chronic cases, extensive fibrosis is seen, the urinary or reproductive tract could be occluded, or marked thickening of the bladder wall.
• Ocular lesions: Mild conjunctival hyperemia and edema to epithelial and lymphoid hyperplasia

• Chlamydia suis: Associated with conjunctivitis, rhinitis, pneumonia, enteritis, reproductive disorders, and asymptomatic infections in swine

• Recognized in the intestinal mucosa of normal and clinically ill swine
• Gnotobiotic pigs experimentally inoculated with C. suis had moderate diarrhea, anorexia, weakness, and body weight loss.
• Microscopically, there was necrosis and exfoliation of enterocytes on the apical half of villi leading to villous atrophy in the distal jejunum and ileum; there was also mild lymphangitis and perilymphangitis
• Chlamydial organisms replicated within small intestinal villus enterocytes, large intestinal enterocytes, lamina propria, submucosa, and mesenteric lymph nodes 

• Crocodilians (farmed Nile, estuarine and Siamese) chlamydiosis: Conjunctivitis and/or systemic infection (hepatitis)
• Anurans and caudates (can act as reservoir hosts): C. pneumonia (less frequently C psittaci), cutaneous hyperemia, excessive shedding of skin (sloughing), subcutaneous edema and hepatosplenomegaly, hepatitis, splenitis, interstitial nephritis, epicarditis, myocarditis
• Pigeons: Chlamydiosis or ornithosis caused by Chlamydia psittaci genotype B. Infects all ages but young birds are more susceptible.
• Chondrichthyes (jawed fish that contain cartilage): Chlamydiales infect gills 
• Mouse/ hamster C. muridarum prevalence in wild type is unknown, and uncommon/ absent in contemporary laboratory mice. Respiratory aerosols or venereal transmission. Silent in naturally infected animals or transient in immunocompromised. Can be infected with C. psittaci resulted in splenomegaly, hepatomegaly, and serofibrinous peritonitis, and intranasal inoculation resulted in pneumonia
• Rabbits: C. psittaci and C. pneumoniae, subclinical but can cause conjunctiva and interstitial pneumonia. 

 

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