JPC SYSTEMIC PATHOLOGY

URINARY SYSTEM

November 2023

U-B03

 

SLIDE A: Signalment (JPC #1619844): Tissue from an 80-pound pig.

 

HISTORY: This pig is one of several among a drove of 40 with clinical signs of fever, lethargy, anorexia, and prostration. Necropsy findings include a swollen, hyperemic liver with petechiation of the kidneys and endocardium. 

 

HISTOPATHOLOGIC DESCRIPTION: Kidney: Multifocally affecting approximately 60% of the kidney, the cortical and medullary interstitium is infiltrated by numerous lymphocytes and plasma cells, fewer neutrophils and macrophages that occasionally contain hemosiderin, as well as fibrin and edema. Tubules are multifocally ectatic and lumina contain variable numbers of viable and degenerate neutrophils, necrotic debris, and eosinophilic proteinaceous material. Multifocally, renal tubular epithelium exhibits one or more of the following changes: attenuation; swollen borders with microvacuolated cytoplasm (degeneration); shrunken and hypereosinophilic with a pyknotic nucleus (necrosis); or increased cytoplasmic basophilia, enlarged vesiculate nuclei, and rare mitotic figures (regeneration). There is mild multifocal dilation of the urinary space of Bowman’s capsule and atrophy of glomeruli. Within the pelvis, the perivascular adventitia is loosely arranged, with increased clear space and ectatic lymphatics (edema), admixed with scattered lymphocytes, plasma cells, and macrophages.  

 

Liver: No significant findings.

 

MORPHOLOGIC DIAGNOSIS: Kidney: Nephritis, tubulointerstitial, lymphoplasmacytic, subacute, multifocal, moderate, with tubular degeneration, necrosis, and regeneration, breed unspecified, porcine.

 

SLIDE B: Signalment (JPC #1255787): Tissue from a 13-year-old male Barbary ape (Macaca sylvanus).

 

HISTORY: This non-human primate died after a one-day illness with lethargy and prostration. Necropsy findings include icterus and widespread visceral ecchymotic hemorrhage and hepatomegaly.

 

HISTOPATHOLOGIC DESCRIPTION: Kidney: Multifocally and variably affecting approximately 80% of the cortex, the cortical interstitium is infiltrated by lymphocytes and plasma cells, fewer neutrophils and rare macrophages, admixed with fibrin and edema. Tubules are often ectatic and lumina contain varying amounts of viable and degenerate neutrophils, sloughed epithelial cells, cellular debris, eosinophilic proteinaceous material, hemorrhage, and fibrin. Multifocally, tubular epithelial cells are either swollen with microvacuolated cytoplasm (degeneration), shrunken and hypereosinophilic with a pyknotic nucleus (necrosis); or have increased cytoplasmic basophilia, enlarged vesiculate nuclei, and rare mitotic figures (regeneration). There is mild multifocal dilation of the urinary space of Bowman’s capsule. 

 

 

MORPHOLOGIC DIAGNOSIS: Kidney: Nephritis, tubulointerstitial, lymphoplasmacytic and neutrophilic, subacute, multifocal, mild, with tubular degeneration, necrosis, and regeneration, Barbary ape (Macaca sylvanus), non-human primate

 

SLIDE C Kidney (Warthin-Starry): There are moderate numbers of 1 – 2 µm x 7 – 10 µm, spiral-shaped, argyrophilic bacteria (spirochetes) within tubular epithelial cells.  

 

ETIOLOGIC DIAGNOSIS: Renal leptospirosis 

 

CAUSE: Leptospira sp. 

 

GENERAL DISCUSSION:  

• Leptospira sp. 

• Generally refers to serovars of Leptospira interrogans
• Slender (0.1 - 0.3 µm), motile, 6 – 20 µm long, weakly gram-negative spirochetes with hooked ends
  • Important cause of abortion and stillbirth in livestock 

• Also causes acute disease in many species (septicemia, hepatitis, nephritis, and meningitis)
• Zoonotic; worldwide distribution; survives months in warm, wet, neutral to alkaline soil, or stagnant water (“flood fever,” “swamp fever,” “mud fever”)
• Each serovar is adapted to one or more “maintenance hosts” in which disease is generally mild or subclinical, and bacteria persist in the kidney or genital tract

• Environmental contamination from urine or abortion discharge

• Disease in “incidental hosts” is generally more severe, with shorter shedding of organisms

 

PATHOGENESIS:  

• Transmission either direct (urine, abortive discharges, venereal, milk, transplacental, venereal, eating infected meat, bite wound) or indirect (urine-contaminated environment) à penetrate mucous membranes (nasal, genital, ocular, intestinal), or water-softened skin à penetrate vessels (blood or lymphatic) à leptospiremia up to 7 days à replication in liver, kidney, lung, placenta, udder, CSF, vitreous humor à next steps depend on production of opsonizing/agglutinating antibodies & serovar (predominantly humoral immunity):

• Adequate antibody titer (vaccinated): organism rapidly eliminated 
• Moderate antibody titer: organism eliminated from most organs, but persists in kidney and may be shed for weeks to months
• Low titer / host-adapted serovar: may or may not cause acute and/or chronic clinical disease à persistence in sites that immunoglobulins penetrate poorly (renal proximal convoluted tubules, CSF, vitreous humor, +/- genital tract) à prolonged (months to years) shedding in urine
• Low antibody titer / virulent serovar:  migration through vascular endothelium à vascular damage à peracute, acute, subacute, and/or chronic disease (thrombocytopenia, renal failure, hepatic necrosis, coagulopathy, hemolysis, pulmonary congestion, meningitis all variable depending on host and serovar) à death, or survival with eventual renal colonization and variable leptospiruria 

• The proximal convoluted tubules of kidney act as natural reservoir for leptospires; renal tubular epithelial cells are likely infected via the basal-lateral cell surface from migration of bacteria through intertubular capillaries à bacteria attach to endothelial cell membranes via adhesins à appear to enter tubular epithelial cell cytoplasm directly via motility (vs. endocytosis and phagosome-lysosome fusion)
• Virulence factors:

• Invasive motility allows penetration of endothelial cells à vasculitis
  • Two flagella with polar insertions are located in the periplasmic space
• Lipopolysaccharide (LPS): stimulates neutrophil adherence and platelet activation, activates macrophages, stimulates IL-1 and IFN secretion 
• The genome of pathogenic Leptospira encodes a group of proteins called Len (Leptospiral endostatin-like) proteins: 
  • Outer membrane leptospiral protein facilitates adhesion to host cell-adhesion molecules and ECM proteins (ie. fibronectin)
  • LigA: attachment & invasion
  • Some proteins bind regulatory proteins such as Plasma factor H
  • LipL32 (another outer membrane protein): Increases the expression of TLR2 (early immune response) *Note: TLR4 plays a role in controlling leptospiral burden during chronic infections
  • Other outer membrane proteins: LipL21, LipL41, OmpL1 (a porin)
• Hemolysins (sphingomyelinase C/H): endothelial damage, tubular necrosis
• Biofilms may also play a role in tubular injury

• In the kidney, the epithelial cells of the cortical proximal convoluted tubules are targeted first; later, medullary epithelial cells of loops of Henle
• Mechanism of injury in renal leptospirosis is cell lysis caused by:

1. Physical properties (penetrating movement) that disrupt endothelial cells
2. Bacterial toxins (sphingomyelinases & lipopolysaccharides) which act directly on tubular epithelial cells
3. Inflammatory cytokines and degradative enzymes  
4. Hypoxia may also play a role in tubule injury

 

TYPICAL CLINICAL FINDINGS:

• Wide variation depending on serovar pathogenicity and host immune status 
• Hyperacute: Puppies (icterohaemorrhagiae); fulminating septicemia > fever, widespread hemorrhages (petechia, melena, hematemesis, epistaxis) > death in hours to days
• Acute (often severe, especially in young animals):

• Fever, muscle tenderness, hemoglobinuria, petechiae, vomiting, diarrhea, intestinal intussusceptions (dog), pneumonia, hepatitis, rare meningitis
• Icterus common in all species due primarily to intravascular hemolysis and toxic and ischemic hepatocellular injury  
• Anemia initially caused by hemolysins, later by autoantibodies (react with leptospiral products coating erythrocytes > intravascular hemolysis); severe in calves and lambs; dogs and humans have milder anemia with hemosiderosis in the spleen and lymph nodes
• Renal failure clinically important classically in L. canicola (and others) in dogs, though all serovars can cause interstitial nephritis (postsepticemic localization in kidneys, with or without overt renal failure) 
• Agalactia and abortion in cattle with incidental serovars (e.g., pomona)

• Subacute: Cattle - less severe with drop in milk production and transient pyrexia ("milk drop syndrome") or "flabby udder mastitis"; Swine - transient mild fever, anorexia, and depression.
• Chronic (most important form in livestock): Abortion, stillbirth, weak newborns, infertility; recurrent uveitis (“periodic ophthalmia”) in horses (pomona)

• Most common manifestation in dogs:  renal insufficiency and chronic interstitial nephritis

• CBC/chem: Initial leukopenia develops into leukocytosis with left shift, thrombocytopenia, moderate anemia (can have hemolysis in horses, calves, lambs, and pigs), hemoglobinemia & hemoglobinuria, hyperbilirubinemia & bilirubinuria, neutrophilia, increased ALT, increased bile acids in cattle

 

TYPICAL GROSS FINDINGS:  

• Acute:  

• Mild icterus, moderate anemia, widespread hemorrhages, petechial and ecchymotic hemorrhages in the mucous membranes, subcutis, visceral surfaces, and occasionally the lungs and heart
• Swollen, hemorrhagic kidneys
• Multifocal to coalescing, discrete, random, white to gray foci (necrosis) with hemorrhage
• Port-wine or bile-colored urine in ruminants and horses

• Chronic:

• Kidneys shrunken and firm, with linear to radiating gray foci +/- uremia
• Abortion:  
  • Ruminants: Autolytic fetus
  • Swine: Mummified fetuses

 

TYPICAL LIGHT MICROSCOPIC FINDINGS:

• Kidney:  

• Neutrophilic (acute) to lymphoplasmacytic and histiocytic (chronic) tubulointerstitial nephritis +/- tubular epithelial injury (degeneration and necrosis)
• Chronic: cortical fibrosis and/or tubular atrophy
• 2^o lesions associated with uremia (mineralization, vasculitis)

• Liver: Portal lymphocytic infiltration with hepatocellular degeneration/necrosis; dissolution of hepatocytes; evidence of cholestasis
• Necrotizing vasculitis, fibrinoid change, thrombosis

 

ULTRASTRUCTURAL FINDINGS:

• Spirochetes with periplasmic flagella associated with microvilli and within phagolysosomes of proximal convoluted tubule
• Interstitium: Capillary endothelial swelling with more surface filopodia, vacuolation, and mitochondrial swelling/cristolysis

 

ADDITIONAL DIAGNOSTIC TESTS:  

• Giemsa or silver stains (Levaditi, Warthin-Starry) and dark-field microscopy are insensitive methods (false-negative & false-positive results are common)
• IHC
• Immunofluorescence of urine & PCR are sensitive diagnostic techniques
• Microscopic agglutination test (MAT): Standard for serologic diagnosis

• May underestimate true prevalence of some host-adapted serovars (low titers are common in maintenance hosts)

 

DIFFERENTIAL DIAGNOSIS:

Interstitial nephritis:

• Pigs:  

• Porcine reproductive and respiratory syndrome (P-V25)
• Porcine circovirus 2 (H-V11)

• Cattle: 

• White-spotted kidney (U-B05)
• Malignant catarrhal fever (U-V02)
• Theileria parva (H-P05)
• Lumpy skin disease

• Dogs:  

• Canine herpesvirus 1 (U-V03)
• Leishmania spp. (H-P07)(I-P15)
• Borrelia burgdorferi (U-B01)
• Hepatozoon canis
• Canine adenovirus 1 (infectious canine hepatitis) (D-V19)

• Horses:  

• Equine infectious anemia (H-V10)

• Sheep:  

• Sheep pox (P-V24)
• Small animal lentivirus (maedi-visna) (P-V17)(N-V13)

 

COMPARATIVE PATHOLOGY 

Leptospirosis sp. in other animals: (Species, [host-adapted serovars])

• Cattle (hardjo):  

• Acute form (infection with non-host adapted pomona) (most severe, uncommon):  
  • Calves: Hemoglobinuria (port-wine urine), fever, anemia, icterus, dyspnea
  • Cows: Agalactia; abortion of decomposed fetus
  • Organisms in liver & kidneys with interstitial nephritis
• Subacute form (most common, less severe) (hardjo type hardjoprajitno):  
  • “Milk drop syndrome”: Drop in milk production, transient pyrexia; thick, yellow, colostrum-like milk
• Chronic form (hardjo and pomona): Fetal infection in pregnant cows > abortion, stillbirth, or birth of premature or weak calves
• Lesions may be mild & nonspecific
  • Periacinar hepatocellular necrosis (anemia) with increased hemosiderin-laden Kupffer cells
  • Multifocal nonsuppurative interstitial nephritis mainly within the cortex
  • Active lesions have atypical regeneration of tubular epithelium with few bizarre syncytia or giant cells

• Sheep & goat (hardjo): less susceptible than cattle

• Lambs: Acute disease (similar to pomona in calves)
• Ewes: Late-term abortion, stillbirth, weak lambs; agalactia 

• Pigs (pomona type kennewicki; tarassovi; bratislava; others):  

• Abortion, small litters of weak piglets most important
• Most often mild fever, anorexia, depression; leptospires persist in kidney and are shed in urine; carrier pigs probably most common route of infection
• Distinct “repeat breeder” infertility syndrome reported in Northern Ireland in sows bred to infected boars for the first time (bratislava, muenchen)
• Most chronic interstitial nephritis lesions in pigs are due to leptospiral infection – usually observed at slaughter

• Horses (bratislava, pomona):  

• Acute (rare): Fever, anorexia, depression, icterus, acute renal failure
• Chronic: Abortion (#3 infectious cause (Canton, J Vet Diagn Invest 2023)), premature foaling, chronic recurrent uveitis (“periodic ophthalmia” especially with pomona infection; S-M01)
  • Leptospiral infection associated with fatal hepatic & renal disease in foals, abortion, premature foaling, and immune-mediated chronic uveitis
  • Giant cell hepatitis (syncytia of hepatocytes) and disruption of hepatic cords in aborted fetuses
• Clinical case horses were more likely to have pomona than other Leptospira serovars (Fagre, J Vet Diagn Invest 2020)

• Dog (canicola): Disease caused primarily by Leptospira interrogans and L. kirschneri

• Increased risk of infection in middle-aged, sexually intact male hounds, herding, working, or mixed breed dogs
• Dogs generally develop clinical signs of renal insufficiency
• Classically: Serovars canicola and icterohaemorrhagiae cause severe disease
• Generally, canicola predominantly affects the kidney, icterohaemorrhagiae the liver, and other serovars affect both (but less severely)
• Characteristic lesions in acute disease: 
  • Widespread hemorrhages; renal cortical and subcapsular hemorrhages are most consistent
  • Dissociation of hepatic cords (not pathognomotic)
• Exudative glomerulonephritis associated with acute leptospirosis in Swiss strain (Hilbe, Vet Pathol 2023)

• Cat: Clinical disease is rare; cats can shed bacteria in urine

• Exposure to leptospires may play a role in the pathogenesis of feline kidney disease
• Baboon: Abortion
• Barbary ape: Convulsions, recumbency, acute death, visceral petechia, hepatomegaly, hepatocellular dissociation

• New world monkeys: Transmitted from rodents 

• Clinical signs: Icterus, anemia, hemorrhagic pneumonia, abortion, renal disease
• Interstitial nephritis +/- necrotizing tubulonephritis 
• Tamarin: Lethargy, icterus, dyspnea, pulmonary hemorrhage, hepatocellular dissociation
• Marmoset: Icterus
• Squirrel monkey: Acute death, fever, icterus, abortion, renal necrosis, pulmonary and visceral hemorrhage, hepatomegaly, exudative glomerulopathy
•  

• NHP: Tubular and/or interstitial nephritis
• Llama & alpaca: Leptospirosis a frequent cause of abortion 
• Hamster: Highly susceptible to ballum (from mice) – severe hemolytic disease. Golden Syrian hamsters get circulating foamy macrophages when infected  (Putz, J Comp Pathol 2021)
• Mouse (ballum) & Rat (icterohemorrhagiae): Fairly resistant to clinical disease; persistent infection with intermittent shedding in urine

• Experimental inoculation of C3H/He mice with icterohaemorrhagiae- pulmonary fibrinoid vasculitis, thrombosis, renal tubular necrosis, nephritis
• T-cell deficiency increases susceptibility; inoculation of C3H-scid mice with Copenhageni- lethal disease with hepatic and renal necrosis

• Gerbil: Very susceptible to experimental infection – icterus, hepatic/renal damage
• Guinea pig: Common model; develop severe acute systemic disease; widespread hemorrhages including perirenal hemorrhage 
• Deer: Fatal nephritis in farmed young red deer (pomona)

• Enlarged kidneys due to severe chronic-active cortical interstitial nephritis

• Sea otter: Rare deaths from leptospirosis; similar gross & microscopic renal findings 

• Gross lesions: None or renal petechiae
• Microscopic findings: Moderate to severe multifocal interstitial nephritis

• Pinnipeds (California sea lion, northern fur seal): 

• Classically pomona, but grippotyphosa common more recently
• Thought to be enzootic in CSLs, with epizootics every 3-5 years
• Clinical findings: Stranding; renal failure
• Gross lesions: Swollen kidneys with loss of distinction between cortex and medulla
• Microscopic findings: Subacute to chronic tubulointerstitial nephritis with lymphoplasmacytic inflammation

• Rhinoceros: Hemolytic anemia in captive black rhinos with leptospirosis 

 

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