Conference 23 - 2015 Case: 03 20160427

Signalment:

2.5 month old, intact female, Yak (Bos grunniens) This yak was bottle-raised from birth and housed in a small pen in the backyard, abutting a building. The yak had an approximately two week history of pica and a one week history of decreased appetite and lethargy, which progressed to loss of a suckle response. The animal developed a fever (104F) and was initially treated with antibiotics and tube feedings of electrolytes and milk at the farm but was referred for suspected laryngeal or pharyngeal trauma based on difficulty tubing and reflux of milk through the nose. On presentation to the referral hospital, the yak was tachypneic (112 rpm) with harsh lung sounds bilaterally and decreased lung sounds ventrally. The yak was markedly azotemic (results below) and was started on intravenous fluids and other supportive care. Thoracic radiographs supported cranioventral pneumonia and a trans-tracheal wash was performed (results below) and antibiotics were initiated. The yak's azotemia did not improve with fluid therapy and the yak was producing minimal urine. The yak developed ascites and a total of 4L was removed via abdominocentesis on the two days preceding euthanasia. The yak did not suckle at any time during hospitalization. The yak was euthanized 7 days after admission to the referral hospital.

Gross Description:

The abdominal cavity contained 2.5 L of translucent, pale pink to yellow, watery fluid and there was marked perirenal edema with mild to moderate hemorrhage, more pronounced surrounding the left kidney. The kidneys were diffusely pale brown and slightly swollen, with small pinpoint foci of hemorrhage in the cortex, reddening of the corticomedullary junction, and hemorrhage and edema in the adipose tissue of the renal pelvis. There was also marked mesocolonic edema and mild to moderate generalized subcutaneous edema. There were no lesions in the esophagus, pharynx or larynx. The rumen contained a moderate amount of gray opaque liquid and small chips of gray paint. The liver was diffusely pale brown. The thoracic cavity contained 0.3 L of fluid and the pericardium contained 0.1 L of fluid, similar to the abdominal fluid. The lungs were dark red caudally and mottled pink and dark red cranially with diffuse interstitial edema and multiple dark red, firm areas of consolidated lung bilaterally in the cranioventral lung fields, consistent with aspiration pneumonia (foreign material present on histopathology).

Histopathologic Description:

Kidney – Diffusely affecting the entire cortex, the proximal tubular epithelium is degenerate or necrotic with marked clear cytoplasmic vacuolization (degeneration) of the proximal convoluted tubule (PCT) segments. The PCT have cuboidal epithelium with a discernible brush border and uniform, basally located nuclei frequently containing aggregates of 2-5 micron diameter, eosinophilic globular material displacing but not marginating the chromatin. Within the PCT lumina, there is eosinophilic material. The plump vacuolated cells of the PCT transition to the proximal straight tubule (PST) segments, which are lined by variably cuboidal to flattened epithelium, often with large nuclei (up to 50 microns in diameter or approximately 3-4 times the size of a normal renal tubular cell) and occasionally multiple nuclei, which contain similar eosinophilic intranuclear material as well as mitoses (interpreted as mixed degeneration and regeneration). There are areas of tubular epithelial necrosis and loss within the PST segment which is characterized by exposure of the basement membrane and numerous cells with pyknotic or karyorrhectic nuclei which are sloughed into the tubular lumen, admixed with small amounts of eosinophilic globular material. There are minimal histologic changes discernible in the medullary components (Loops of Henle, collecting ducts). The distal straight and convoluted tubules (DST, DCT) have uniform cuboidal epithelium with slightly basophilic cytoplasm, central located nuclei which rarely have eosinophilic intranuclear material but moderate anisokaryosis, no discernible brush border and often have homogeneous pale to brightly eosinophilic material in the tubule lumen (high protein-content fluid). Occasionally, there are small aggregates of basophilic granular material within the tubular lumina (mineral). The glomeruli are condensed and occasionally have slightly increased pale eosinophilic material expanding the mesangium. The interstitium of the renal cortex is expanded by clear to amphophilic, occasionally granular material (edema fluid) and mildly increased amounts of immature collagen and fibroblasts (fibrosis). There are multiple nodular aggregates of moderate numbers of lymphocytes, smaller numbers of numbers of plasma cells and macrophages within the cortical interstitium. There are multiple small areas of hemorrhage within the perivascular space, interstitium, fibroadipose tissue subjacent to the renal pelvis, and renal capsule. Occasionally the aggregates of eosinophilic intra-epithelial, intra-nuclear material were strongly acid-fast positive and frequently were moderately PAS positive.

Morphologic Diagnosis:

Kidney, tubules – tubular degeneration and necrosis, diffuse, marked, acute, with intra-epithelial intra-nuclear eosinophilic material (occasionally acid-fast and frequently PAS positive), tubular regeneration, and interstitial edema and hemorrhage Kidney – nephritis, interstitial, lymphocytic, multifocal, mild, chronic

Lab Results:

Clinical pathology:

*Reference intervals were not provided for yak; references ranges in parentheses are for cattle tested at the referral hospital for general comparison only

Significantly abnormal values on presentation:

BUN – 128 mg/dL (10-24)

Creatinine – 17.1 mg/dL (0.6-1.3)

Calcium – 7.6 mg/dL (8.1-10)

Phosphorus – 11.6 mg/dL (3.4-7.7)

Total protein – 4.7 mg/dL (6.2-8.9)

Albumin – 2.8 mg/dL (3.2-4)

HCT – 18% (25-47)

Values on the day of euthanasia (7 days later):

BUN – 119 mg/dL

Creatinine – 20.1 mg/dL

Calcium – 8.2 mg/dL

Phosphorus – 10.2 mg/dL

Total protein – 3.4 mg/dL

Albumin – 1.9 mg/dL

PCV – 12%

Microbiology:

Antemortem aerobic culture of trans-tracheal wash: Pseudomonas aeruginosa

Postmortem aerobic culture of lung: Pseudomonas aeruginosa

Postmortem aerobic culture of kidney, spleen and liver: No growth

Molecular diagnostics:

Kidney was submitted for Leptospira species PCR (16s rRNA): negative

Tissue homogenate of lung and spleen was submitted for BVD PCR: negative

Toxicology:

Liver was submitted for toxic element screen:

Lead detected at 62 ppm (wet tissue basis) (<1 ppm considered normal for cattle, >10 ppm liver lead is consistent with toxicosis)

Condition:

Lead toxicosis, yak

Contributor Comment:

This case is an example of acute tubular necrosis in a milk-fed, domestic yak calf (Bos grunniens) due to acute-to-subacute lead toxicosis. This animal was exposed to lead by ingestion of lead-based paint (gray paint chips in the rumen) chewed from the siding of the adjacent building. The clinical presentation of ruminants with lead toxicosis is often one of neurologic deficits or altered mentation. Histologically, there may be cerebral edema and laminar necrosis;⁹ however, this yak did not have gross or histologic evidence of notable cerebral edema or necrosis and, other than lethargy and an inability to suckle, was not reported to have neurologic deficits. Peripheral neuropathy, including dysphagia and laryngeal paralysis, has been reported to be a common clinical sign in horses with both acute and chronic lead toxicosis¹¹ and may be the underlying reason for this yak's reported loss of suckle, reflux and aspiration pneumonia which resulted in referral. Renal changes attributed to lead toxicosis are usually reported to be mild and occasionally evident only as eosinophilic or poorly-staining intranuclear inclusions which are acid-fast positive but, similar to this case, severe nephrosis has also been described in young calves.⁹ The mechanism of renal toxicity of lead is known to involve numerous pathways. Structural mitochondrial degeneration occurs (mitochondrial swelling and distortion of cristae) as well as decreased activity of mitochondrial heme-pathway enzymes,⁵ which interfere with cellular energy production needed to maintain homeostasis and fuel active transport processes vital to renal tubular cell function. Lead also impacts nuclear function through altered gene expression⁵ and is histologically and ultrastructurally evident as lead-protein inclusion bodies in some cases. In-terestingly, not all of the histologically-evident eosinophilic intranuclear material in this case was acid-fast positive but the majority was variably PAS-positive, sug-gesting that many of the inclusions are high in glycogen or other carbohydrate-rich compounds but did not necessarily contain lead complexes. Electron microscopy was performed in this case and solitary or multiple, variably-sized electrodense in-clusions were present in the nuclei of some renal tubular epithelial cells, however the inclusions incorporated less fibrillar material than expected based on previously published reports of lead inclusions.^3,12 This may be due to the fact that this was a diagnostic case and the tissue was formalin-fixed prior to gluteraldehyde fixation, which may have introduced artifactual changes. Additional information regarding the composition of inclusions could have been obtained from energy-dispersive x-ray spectroscopy (EDS) but this capability is not available within the electron microscope at the submitting institution. The specificity of renal acid-fast positive inclusions for lead intoxication has been reported to be high in cattle¹³ but renal intranuclear acid-fast inclusions should be further investigated. Other general me-chanisms of lead toxicity likely impacting renal function include lead competitively binding in place of calcium, altered calcium regulation, and structural and functional alteration in cellular enzymes.¹⁰

Young animals are known to absorb ingested lead more efficiently than adults⁷ and there are numerous dietary factors known to impact absorption of ingested lead.¹ Many studies on lead absorption are rodent-based; however, studies in cattle identified significant differences in lead absorption from milk-fed vs. grain and hay-fed calves¹⁵ and in absorption based on levels of lactose in the diet.¹⁴ Calves fed exclusively milk and calves fed elevated lactose levels with grain absorbed more lead than calves not receiving milk or without high levels of lactose supplemented to grain. These diet-related factors may be a major factor for susceptibility in young animals, and in this case the affected yak was on a milk-only diet.

The underlying cause and significance of the lymphocytic interstitial nephritis in this case is unknown.

JPC Diagnosis:

1. Kidney: Tubular degeneration, necrosis, regeneration, and proteinosis, diffuse, marked with tubular casts and intranuclear, eosinophilic inclusion bodies within renal tubule epithelial cells.

2. Kidney: Nephritis, cortical, interstitial, chronic, multifocal, mild.

Conference Comment:

In addition to domestic animals, wildlife may also be exposed to lead from various sources. Ingestion of lead is the most common route of exposure, although toxicity due to lead-containing shot is also common (but likely poses a low risk for lead toxicity).⁸

Lead exposure is of particular concern in wild avian species where it may affect a variety of birds ranging from waterfowl to bald eagles. In avian species, lead can inhibit enzymes involved in hemoglobin synthesis and when exposed at high levels, may result in anemia. Lead toxicity is most often a chronic condition and as such, affected birds are often debilitated and in poor body condition.⁶ Gross lesions may include esophageal, ventriculus and pro-ventriculus impaction with food, gall bladder distension, pale streaks in the myocardium and muscle of the ventriculus indicative of necrosis, as well as pallor of internal organs. Eosinophilic lead inclusions in the nuclei of proximal tubule epithelium also occur in the kidney of birds and, while this finding is specific for lead poisoning, it may not be present in all cases. Although not necessarily specific for lead toxicity, other histologic changes in affected birds may include hepatic hemosiderosis, fi-brinoid necrosis of arterioles, en-cephalopathy and peripheral neuropathy. The highest concentration of lead in birds is found in the bone, liver and kidney; lead levels in bone decline much slower than soft tissues and thus bone serves as a much longer term location of lead deposits.⁶ Lead has no biologic function and although lead can exert its toxic effects via a variety of mechanisms, its competition with calcium in various biologic functions is of particular importance. This can result in various pathophysiologic effects such as inhibition of neurotransmitter release, defects in ion pumps and channels as well as alterations in protein kinase function.⁶ Lead is considered neurotropic, although the precise reason is not well understood; neuronal changes are non-specific and laminar cortical necrosis may be seen in some cases.² In addition to the central nervous system, lesions also occur in bone. The ch-aracteristic lesion is referred to as a "lead line", which is a band of sclerosis located at the metaphysis of developing bones. It is seen as an early lesion in both children and animals. The lesion consists of persistent mineralized cartilage trabeculae which cannot be effectively resorbed by osteoclasts in spite of their apparent abundance microscopically. Osteoclasts may also contain acid-fast intranuclear inclusions.⁴ We thank the contributor for providing clinical pathology data and gross images with the submission, which enhance the teaching / learning value of the case.

References:

1. Barltrop D, Khoo HE. The influence of nutritional factors on lead absorption. Postgrad Med J. 1975;51(601):795-800.

2. Cantile C, Youssef S. Nervous system. In: Maxie MG, ed. Jubb, Kennedy, and Palmer's Pathology of Domestic Animals. 6th ed. Vol 1. St. Louis, MO: Elsevier; 2016:316-317.

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9. Maxie MG. Jubb, Kennedy, and Palmer's Pathology of Domestic Animals. 5th ed. Elsevier Saunders; 2007.

10. Needleman H, Needleman. Lead Poisoning. Annu Rev Med. 2004;55(1):209-222.

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12. Richter GW, Kress Y, Cornwall CC. Another look at lead inclusion bodies. Am J Pathol. 1968;53(2):189-217.

13. Thomson RG. Reliability of acid-fast inclusions in the kidneys of cattle as an indication of lead poisoning. Can Vet J. 1972;13(4):88-9.

14. Zmudzki J, Bratton GR, Womac CW, et al. Lactose and milk replacer influence on lead absorption and lead toxicity in calves. Bull Environ Contam Toxicol. 1986;36(1):356-363.

15. Zmudzki J, Bratton GR, Womac C, et al. The influence of milk diet, grain diet, and method of dosing on lead toxicity in young calves. Toxicol Appl Pharmacol. 1984;76(3):490-497.