JPC SYSTEMIC PATHOLOGY
REPRODUCTIVE SYSTEM
JANUARY 2022
R-B08

Signalment (JPC #2015774):  A cross-bred ewe

HISTORY:  A flock of cross-bred ewes experienced an abortion outbreak with 40% of the ewes affected.  The aborted fetuses were near-term and in a fresh state. 

HISTOPATHOLOGIC DESCRIPTION:  Chorioallantois, cotyledon and intercotyledonary area: Within the cotyledon, there is multifocal to coalescing loss of chorionic villar architecture with replacement by abundant eosinophilic cellular and karyorrhectic debris (lytic necrosis), hemorrhage, fibrin, edema, and viable and degenerate neutrophils with fewer macrophages and concretions of intensely basophilic granular material (mineral) that multifocally expands the intervillous space.  Scattered throughout the necrotic debris, attached and sloughed trophoblasts are often markedly expanded by intracytoplasmic, amphophilic to pale basophilic, homogenous inclusions composed of aggregates of <1 um diameter cocci occasionally admixed with scant mineral.  Within the chorioallantoic connective tissue, the tunica intima and tunica media of multifocal blood vessels are infiltrated and expanded by neutrophils admixed with eosinophilic karyorrhectic debris and scant fibrin (necrotizing vasculitis).  Multifocal vascular lumens are variably occluded by fibrin thrombi.  There are perivascular accumulations of viable and degenerate neutrophils with fewer macrophages, lymphocytes, and plasma cells.  Within the chorioallantoic stroma there are low numbers of previously described inflammatory cells as well as increased clear space (edema) and ectatic lymphatics. The intercotyledonary chorionic epithelium is either ulcerated or lined by degenerate or necrotic trophoblasts that occasionally contain previously described intracytoplasmic inclusions of <1um cocci.  The surface and underling stroma contain variable amounts of necrotic debris and mineral admixed with neutrophils, lymphocytes, and plasma cells.

MORPHOLOGIC DIAGNOSIS:  Chorioallantois, cotyledon and intercotyledonary area:  Placentitis, necrosuppurative, diffuse, severe, with vasculitis and intratrophoblastic inclusions of cocci, consistent with Chlamydia abortus, cross-bred sheep, ovine.

ETIOLOGIC DIAGNOSIS:  Placental chlamydophilosis

CAUSE:  Chlamydia abortus (formerly Chlamydophila abortus)

CONDITION:  Ovine enzootic abortion or Enzootic abortion of ewes (EAE)

GENERAL DISCUSSION:

• Contagious disease with worldwide distribution that causes fever, abortion, stillbirths, and birth of weak lambs
• Gram-negative, obligate intracellular bacterium; forms a cell wall; multiplies in membrane-bound vacuoles; larger than viruses; possess DNA and RNA
• Does not synthesize ATP like bacteria; susceptible to antibiotics
• Zoonotic and reportable - may cause abortion in women in close contact with aborting ruminants
• Unique morphology and biphasic life cycle with four morphologically distinct forms
• Elementary body (EB): extracellular and infectious form of bacteria that enters the cell; (0.2 µm)
• Reticulate body (RB): intracellular, metabolically active, noninfectious, replicating form that synthesizes DNA, RNA, and protein, and divides by binary fission (0.8 µm)
• Intermediate body (IB): morphologic characteristics 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

PATHOGENESIS:

• Ingestion of infective placenta or uterine discharge (also vaginal discharge and semen) from aborting ewes > oral, conjunctival, or reproductive mucosae exposure > chlamydophilemia > latent infection at an unknown site in body maintained under the control of IFN-gamma (induces production of indoleamine 2,3-dioxygenase which makes tryptophan unavailable to the organism and induces production of nitric oxide that directly inhibits the organism) > release from latency during pregnancy due to immune modulation > chlamydophilemia > hematogenous spread to the placenta > organisms transverse maternal capillaries and enter the extravasated blood in the lacunae > infect trophoblasts and replicate within chlamydial inclusions > host cell bursts and infects neighboring cells > organism enters the fetus at the site of placental damage > abortion occurs secondary to fetal death, resulting from damage to the pregnant uterus, placenta and/or fetus (production of TNF-alpha by fetal macrophages expressing MHC II molecules)
• Chlamydiae use major outer membrane proteins, heat shock protein 70, OmcB, and heparin sulfate-like glycosaminoglycans as adhesins > concentrate on specialized membrane microdomains rich in cholesterol, glycosphingolipid, and caveolin on host cell surfaces > polymorphic membrane proteins involved in early entry > host cell cytoskeleton rearrangement leads to bacterial uptake in a chlamydial inclusion, the membrane of which has clatharin coated pits > organism replicates within the inclusion > Type III secretion system to secrete proteins in the inclusion membrane

TYPICAL CLINICAL FINDINGS:

• Abortions occur in outbreaks (up to 75%) when newly introduced to a flock; once enzootic in a flock, yearly abortion rates of 5%; incubation is 50-90 days
• Infected ewes: Little to no clinical signs of disease after infection until they abort
• Ewes infected before 5-6 weeks of gestation -> abort in late gestation (or have still birth or weak lamb)
• Ewes infected after 5-6 weeks of gestation -> abort in subsequent pregnancy
• Abortion occurs during the final trimester (late term)
• Ewes that abort will not abort again but will carry the organism for several years
• Lambs may be still born or weak at birth
• Ewes may be sick, retain fetal membranes, or develop metritis

TYPICAL GROSS FINDINGS:

• Placenta (similar to bovine brucellosis (R-B03)): affects cotyledons and intercotyledonary areas
  • Cotyledons dull-clay or dark-red color, firm, and matted with a dirty-red exudate;
  • Intercotyledonary areas are red to brown, irregular patches of edema amidst a dry, leathery thickening of the chorioallantois
• Fetus: usually well preserved, few gross lesions; hemorrhage of the subcutis, thymus, lymph nodes, muscle, thoracic and abdominal cavities; hepatomegaly

TYPICAL LIGHT MICROSCOPIC FINDINGS:

• Placenta: Necrotizing placentitis, organisms within trophoblasts (inclusion bodies)
  • Cotyledon and intercotyledonary areas: Infiltrate of neutrophils and macrophages, fewer lymphocytes, with inclusions (aggregates of gram-negative Chlamydia cells within cytoplasmic vacuole) within trophoblasts
  • Endometrium: High numbers of lymphocytes
  • Blood vessels: Marked vasculitis; fibrinoid necrosis of tunica media, infiltrate of neutrophils and mononuclear cells
• Fetus:
  • Liver/spleen: Foci of coagulative necrosis surrounded by mononuclear cells
  • Lung: Alveolar septa thickened by mononuclear cells
  • Brain: Mild meningoencephalitis with vasculitis and hemorrhage

ULTRASTRUCTURAL FINDINGS:

• Elementary body (EB): Spherical, 0.2-0.3 um 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 um, binary fission characterized by “hour-glass” profiles
• Intermediate body (IB): 0.3-1.0 um 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
• Chlamydial cell wall has inner and outer membranes separated by a periplasmic space but no peptidoglycan layer
• Host cell glycogen and mitochondria are closely associated with the bacteria

ADDITIONAL DIAGNOSTIC TESTS:

• Staining of cotyledonary scrapings or histologic sections with modified Ziehl-Neelsen, Gimenez, Machiavello, or Giemsa
• Fluorescent antibody test, ELISA, PCR, immunoperoxidase procedure
• Demonstration of antibodies in the dam is of little use as a large proportion of cattle and sheep have been exposed and test positive

DIFFERENTIAL DIAGNOSIS:

• Chlamydia pecorum: Similar lesions to abortus; differentiate by cell culture isolation and PCR
• Brucella ovis: Edematous, opaque, leathery intercotyledonary areas and necrotic cotyledons (grossly indistinguishable from mycotic abortion or rare cases of campylobacteriosis with late term abortion); light brown exudate on chorionic surface; trophoblasts with gram-negative coccobacilli; vasculitis of larger chorionic vessels; nonspecific fetal gross lesions (calcified plaques on hooves are found but not specific); pneumonia, lymphadenitis, interstitial nephritis and pericholangitis
• Campylobacter fetus fetus and C. jejuni: Edematous intercotyledonary areas and friable yellow cotyledons; necrotizing and suppurative placentitis most severe in chorionic villi with large dense gram-negative bacterial emboli within chorionic capillaries; yellow hepatic foci with targetoid depressed red centers (necrotizing hepatitis) and fibrinous peritonitis in fetus
• Flexispira rappini: Similar lesions as Campylobacter
• Coxiella burnetii (R-B07): Gross and histologic lesions similar; placenta thick, leathery and covered by a thick exudate; necrotizing and suppurative placentitis primarily involving intercotyledonary areas (cotyledons more affected with abortus); fetus with mononuclear inflammation in the lung, liver, and kidneys; hypertrophic trophoblasts filled with organisms visible with modified Ziehl Neelsen or Macchiavello stains; often lacks vasculitis
• Listeria monocytogenes: Necrotizing and suppurative placentitis affecting the cotyledons and intercotyledonary areas, with Gram-positive bacilli filling trophoblasts; hepatomegaly and dissociation of hepatic cords in fetus; possible sepsis and metritis in aborting ewe
• Toxoplasma gondii: Gross edema of the intercotyledonary area and 1-2 mm yellow-white sometimes gritty foci in cotyledons; trophoblast hypertrophy, hyperplasia, necrosis and mineralization with toxoplasma organisms; nonsuppurative encephalitis and necrosis with tachyzoites in heart, liver, lungs and skeletal muscle of the fetus

COMPARATIVE PATHOLOGY:

• Chlamydial nomenclature has undergone multiple revisions over recent years, including a controversial split of genus Chlamydia into 2 genera, Chlamydia and Chlamydophila, based on analysis of the 16 S and 23 S rRNA genes; this scheme has since been abandoned with reversion use of a single genus, Chlamydia (Borel et al, Vet Pathol. 2018)

Chlamydia sp. in other species:

• Ruminants
  • C. abortus: sporadic bovine encephalomyelitis, pneumonia, polyarthritis of calves, conjunctivitis, and sporadic abortions; reported to cause bronchopneumonia in experimentally challenged calves
  • C. pecorum: causes:
    • Enteritis in calves <10 days old
    • Meningoencephalitis, vasculitis, and abortion in cattle (Struthers, Vet Pathol. 2021) (see N-B05 for encephalitis manifestations of sporadic bovine encephalomyelitis)
    • Arthritis in sheep (Ostfeld, Vet Pathol. 2021)
    • Aporadic ovine abortions (Westermann, Vet Pathol. 2021)
  • C. psittaci
  • C. suis
• Camelids: abortus:  possible cause of ovarian hydrobursitis; 86% camels have serologic evidence of infection, suggesting organism involved in pathogenesis
• Horses
  • C. abortus, C. pneumonia: equine pneumonia or abortion
  • C. psittaci: identified in cases of abortion, premature death of foals in Australia, one case in Switzerland (Bauman, J Vet Diagn Invest. 2020)
• Avians
  • C. psittaci (D-B12, psittacosis): more than 465 avian species affected; pigeons and psittacines most commonly affected; multifocal hepatic necrosis, splenomegaly and fibrinous airsacculitis, pericarditis, and peritonitis; zoonotic infections range from asymptomatic to severe systemic disease with pneumonia, myocarditis, and encephalitis
  • C. gallinacea: detected in domestic poultry, guinea fowl, turkeys, ducks, and wild birds; no overt disease in chickens but associated with significant body weight reduction; thought to be endemic in chickens with ability to persist over time; zoonotic potential
  • C. avium: found in pigeons and psittacines in European countries; limited data linking to respiratory disease in these species
• Koalas
  • C. pecorum: keratoconjunctivitis, vaginitis, ovarian cysts, infertility; urogenital disease ("wet tail" or "dirty tail"); 3 infection categories: subclinical (no signs of disease), overt disease (presence of obvious external clinical signs such as conjunctivitis and “wet tail”) or inapparent overt disease (lesions are not clinically obvious but can be only detected using ultrasound, postmortem examination or histopathology) (Palmieri, Vet Pathol. 2019)
  • C. pneumoniae: rhinitis, pneumonia, conjunctivitis
• Pigs ( suis, C. abortus, C. pecorum, and C. psittaci)
  • C. abortus: abortion, vaginitis, endometritis, seminal vesiculitis, mastitis (latent)
  • C. pecorum: polyarthritis, serositis, enteritis, pneumonia
  • C. suis: Conjunctivitis, pneumonia, enteritis, polyarthritis
• Cats
  • C. felis: persistent conjunctivitis
  • C. psittaci: one case of fatal infection in kitten (Sanderson, J Vet Diagn Invest. 2021)
• Guinea pigs: caviae: Guinea Pig Inclusion Conjunctivitis (GPIC); can also cause rhinitis and genital tract infections
• Mice and hamsters: muridarum: pneumonitis, ileitis
• Reptiles: psittaci and C. pneumoniae (more common): free-ranging and captive reptilian hosts including puff adders, boas, chameleons, crocodiles, turtles and tortoises; granulomatous inflammation in inner organs such as heart, liver, spleen and lung
• Humans:
• C. pneumoniae: respiratory infections; nearly ubiquitous in humans, with seropositivity rates of 70–80% in older populations
• C. trachomatis: ocular (inclusion conjunctivitis in newborns) and genital infections; infantile pneumonia
• C. abortus: abortion in pregnant women following contact with aborting/lambing sheep and goats; if untreated, may progress to life-threatening illness
• C. psittaci: influenza-like symptoms; pneumonia
• C. muridarum

REFERENCES:

1. Agnew D. Camelidae. In: Terio K, McAloose D, Leger J, eds. Pathology of Wildlife and Zoo Animals, San Diego, CA: Elsevier 2018:190.
2. Barthold SW, Griffey SM, Percy DH. Pathology of Laboratory Rodents and Rabbits. 4th ed. Ames, IA: Blackwell Publishing; 2016: 60-61, 224.
3. Baumann S, Gurtner C, Marti H, Borel Nc. Detection of Chlamydia species in 2 cases of equine abortion in Switzerland:  a retrospective study from 2000 to 2018. J Vet Diagn Invest. 2020; 34(4):542-548.
4. Borel N, Polkinghorne A, Pospischil A. A review on Chlamydial diseases in animals: still a challenge for Pathologists?. Vet Pathol. 2018; 55(3):374-390.
5. Foster RA. Female reproductive system and mammae. In: McGavin MD, Zachary JF, eds. Pathologic Basis of Veterinary Disease. 6th ed. St. Louis, MO: Elsevier; 2017:1182.
6. Fratzke A, Howard LL, Tocidlowski ME, et al. Chlamydia pneumoniae Polioencephalomyelitis and Ganglionitis in Captive Houston Toads (Anaxyrus houstonensis). Vet Pathol. 2019;56(5):789-793.
7. Giannitti F, Anderson M, Miller M, et al. Chlamydia pecorum: fetal and placental lesions in sporadic caprine abortion. J Vet Diagn Invest. 2016; 28(2): 184-9.
8. Longbottom D, Livingstone M, Maley S, van der Zon A, Rocchi M. Intranasal infection with Chlamydia abortus induces dose-dependent latency and abortion in sheep. PLoS ONE. 2013; 8(2) e57950.
9. Lopez A, Martinson SA. Respiratory system, mediastinum, and pleurae. In: McGavin MD, Zachary JF, eds. Pathologic Basis of Veterinary Disease. 6th ed. St. Louis, MO: Elsevier; 2017:552.
10. Ostfeld N, Islam MM, Jelocnik M, et al. Chlamydia pecorum-Induced Arthritis in Experimentally and Naturally Infected Sheep. Vet Pathol. 2021;58(2):346-360.
11. Palmieri C, Hulse L, Pagliarani S, et al. Chlamydia pecorum Infection in the Male Reproductive System of Koalas ( Phascolarctos cinereus). Vet Pathol. 2019;56(2):300-306.
12. Sammin DJ, Markey BK, Quinn PJ, McElroy MC, Bassett HF. Comparison of fetal and maternal inflammatory responses in the ovine placenta after experimental infection with Chlamydophila abortus. J Comp Pathol. 2006;135:83-92.
13. Sanderson H, Vasquez M, Killion H, Vance M, Sondgeroth K, Fox J. Fatal Chlamydia psittaci infection in a domestic kitten. J Vet Diagn Invest. 2021;33(1):101-103.
14. Schlafer DH, Foster RA. The female genital system. In: Maxie MG, ed. Jubb, Kennedy, and Palmer’s Pathology of Domestic Animals. Vol 3. 6th ed. St Louis, MO; 2016:414-416.
15. Struthers JD, Lim A, Ferguson S, et al. Meningoencephalitis, Vasculitis, and Abortions Caused by Chlamydia pecorum in a Herd of Cattle. Vet Pathol. 2021;58(3):549-557.
16. Thongkamkoon P, Tohmee N, Morris EK, Inamnuay L, Lombardini ED. Combined Fatal Systemic Chlamydia sp. and Aeromonas sobria Infection in Juvenile Siamese Crocodiles ( Crocodylus siamensis). Vet Pathol. 2018;55(5):736-740.
17. Westermann T, Jenkins C, Onizawa E, et al. Chlamydia pecorum-Associated Sporadic Ovine Abortion. Vet Pathol. 2021;58(1):114-122.

Click the slide to view.

---