Temporal and geographic clustering of polyomavirus-associated olfactory tumors in 10 free-ranging raccoons (Procyon lotor).

Reports of primary nervous system tumors in wild raccoons are extremely rare. Olfactory tumors were diagnosed postmortem in 9 free-ranging raccoons from 4 contiguous counties in California and 1 raccoon from Oregon within a 26-month period between 2010 and 2012. We describe the geographic and temporal features of these 10 cases, including the laboratory diagnostic investigations and the neuropathologic, immunohistochemical, and ultrastructural characteristics of these tumors in the affected animals. All 9 raccoons from California were found within a localized geographic region of the San Francisco Bay Area (within a 44.13-km radius). The tight temporal and geographic clustering and consistent anatomic location in the olfactory system of tumor types not previously described in raccoons (malignant peripheral nerve sheath tumors and undifferentiated sarcomas) strongly suggest either a common cause or a precipitating factor leading to induction or potentiation of neuro-oncogenesis and so prompted an extensive diagnostic investigation to explore possible oncogenic infectious and/or toxic causes. By a consensus polymerase chain reaction strategy, a novel, recently reported polyomavirus called raccoon polyomavirus was identified in all 10 tumors but not in the normal brain tissue from the affected animals, suggesting that the virus might play a role in neuro-oncogenesis. In addition, expression of the viral protein T antigen was detected in all tumors containing the viral sequences. We discuss the potential role of raccoon polyomavirus as an oncogenic virus.

Sudden behavior change in a cat.

A 5-year-old, spayed female, domestic short-haired cat had a 10-day history of sudden behavioral changes followed by seizures. Blood parameters were in the reference ranges, and radiographs failed to detect a mass lesion in the brain. Euthanasia was followed by rabies testing, which was negative. Gross lesions were absent. Histologic changes were present only in the brain and consisted of foci of hippocampal pyramidal cell loss, mild gliosis, pallor of the associated neuropil, and neovascularization.

Diagnostic exercise: Multiple skin nodules in a cat.

A 10-year-old, neutered, male, domestic short-haired cat had numerous, small, firm, round, red, nonpruritic, nonpainful, dermal nodules 5-16 mm in diameter that ruptured within 48 hours of their appearance and subsequently crusted over. The masses were located in all regions of the body. One mass was excised from the dorsal right carpus and examined histologically, and 2 masses from the interscapular region were cultured for bacteria. The excised dermal mass from the carpus effaced normal dermal architecture, pressed tightly against the epidermis, and was composed of tightly packed round to polyhedral cells that extended to the deep margins of the sections. The overlying epidermis was extensively ulcerated and vesiculated with intraepidermal nests of cells identical to those in the dermis. There was marked anisokaryosis in the deeper regions of the mass with numerous multinucleated cells and cells with giant bizarre nuclei. The histological appearance and CD18 immunocytochemical staining of this mass are consistent with a diagnosis of feline progressive epitheliotropic dendritic cell histiocytosis.

Lymphadenopathy associated with a thyroid carcinoma in a dog.

Angiomatoid lesions in a lymph node associated with a thyroid carcinoma of a dog were restricted to the subcapsular and medullary sinuses. Lymphoid atrophy was present, but nodal architecture was not distorted and normal structures were not invaded. Immunohistochemical staining indicated that the vascular spaces formed by spindloid cells were lined by endothelium with a low mitotic index. The spindloid cells were positive for smooth muscle actin, vimentin, and desmin and thus were likely to be fibroblasts, myofibroblasts, smooth muscle cells, and/or pericytes. These features are comparable to vascular transformation of lymph node sinuses in humans (nodal angiomatosis), a nonneoplastic condition often associated with mechanical or functional blockage of efferent lymphatics and veins.

Neurologic disease in a pig.

Three 4-week-old Yorkshire-Hampshire cross piglets from a litter of 9 (7 liveborn) developed convulsions the day of weaning. They were subsequently obtunded, ataxic, and hypermetric and had intention tremors. An affected male pig was presented live for necropsy on day 5 postweaning. This animal was euthanatized and necropsied. No significant grossly visible postmortem lesions were found. Histologic examination of the brain disclosed laminar necrosis of the submeningeal cerebral and cerebellar cortices with replacement by broad sheets of gitter cells. Occasional cerebral and cerebellar leptomeningeal and parenchymal vessels were surrounded by lymphocytes with fewer eosinophils. The morphologic diagnosis was severe multifocal subcortical cerebral and cerebellar laminar necrosis with moderate multifocal lymphocytic and eosinophilic cerebral and cerebellar leptomeningeal and parenchymal perivasculitis. The history and histologic findings are consistent with an etiologic diagnosis of sodium ion intoxication.

Urinary bladder mass in a dog.

A 7-year-old spayed female Schnauzer dog with chronic hematuria had a soft tissue mass within the wall of the bladder. The mass was excised and, when examined histologically, was determined to be a discrete well-organized mural mass consisting of spindloid cells arranged in swirling sheets and palisades punctuated by aggregates of principally eosinophils. The overlying transitional mucosa was extensively ulcerated and focally hyperplastic and nodular with subjacent solid down growths, superficial cysts, and the extension of tubular structures deep into the submucosa. The histologic appearance of this mass is consistent with canine polypoid eosinophilic cystitis, also known as benign inflammatory fibrous polyp.

Purkinje fiber dysplasia (histiocytoid cardiomyopathy) with ventricular noncompaction in a savannah kitten.

In a 2-month-old female savannah kitten that died unexpectedly, the pathologic findings of significance were restricted to the heart and included abnormal Purkinje fibers and biventricular myocardial trabeculation or noncompaction. The Purkinje fibers were large, angular, and tightly packed. They contained few disorganized myofibrils among a rarified cytoplasm. The fibers were distinct from adjacent myocytes and were immunohistochemically positive for desmin, muscle actin, myoglobin, sarcomeric actin, and chromogranin A. These findings are identical to those that occur in children with histiocytoid cardiomyopathy, a fatal genetic mitochondrial disorder of Purkinje fibers. Ventricular noncompaction likely has a multifactoral cause that results from fetal arrest of ventricular organizational development that might occur in conjunction with, or independent of, histiocytoid cardiomyopathy.

Perspectives on academic veterinary administration.

It is important for veterinary administrators to apply knowledge bases from other fields to their own unique administrative needs. For example, although some resources are written for business managers, the discussions of four key management competency areas, guidelines for mastering these skills, organizational assessment tools, and other self-help tools may provide interesting food-for-thought for veterinary administrators.(76) In developing their own administrative styles, administrators should seek to apply those principles that seem to intuitively fit with their personal research styles, work situations, managerial styles, administrative preferences, and unique organizational culture. Through strengthening their liaisons with community and university business programs, counseling agencies, employee assistance programs, and psychology researchers, administrators can continue to be exposed to and benefit from new paradigms for consideration in veterinary medical environments. Through these liaisons, the unique needs of veterinary medical environments are also communicated to individuals within the fields of psychology and business, thus stimulating new research that specifically targets veterinary medical environment leadership issues. Each field has unique contributions to help veterinary administrators work toward creating veterinary medical environments that are creative, energetic, visionary, pragmatic, and highly marketable in order to help administrators recruit and nurture the best and brightest veterinary researchers, teachers, and clinicians.

Endocardial ossifying myxoma of the right atrium in a cat.

Primary cardiac neoplasms are rare in humans and animals. In humans, the most common primary cardiac tumor is the myxoma, which is frequently found in the left atrium. Cardiac myxoma has been reported in the dog but not in the cat. We describe the gross, immunohistochemical, and light microscopic examination of a myxoma in the right atrium of a 6-year-old domestic shorthair cat. Histologically, the tumor consisted predominantly of mesenchymal cells with several foci of bone and cartilage present. The tumor was encapsulated and benign.

Rabbit intestinal xenograft model for human Encephalitozoon infections in mice.

BACKGROUND AND PURPOSE: The gastrointestinal tract is a common portal of entry for Encephalitozoon cuniculi, one of several microsporidial organisms emerging as opportunistic pathogens in immunocompromised humans. Although most human microsporidial pathogens can be propagated in vitro and in a variety of laboratory animals, an experimental animal system to specifically study intestinal uptake and systemic spread of these organisms does not exist. METHODS: Paired segments of near-term fetal rabbit small intestine were implanted subcutaneously into 25 athymic nude or 10 severe combined immune deficient mice. Five weeks after surgery, 65 xenografts were inoculated intraluminally with E. cuniculi (n = 14), E. intestinalis (n = 27), E. hellem (n = 20), or RK-13 cells (n = 2), or were left uninoculated (n = 2). RESULTS: Intestinal xenograft infection with E. cuniculi (n = 11), E. intestinalis (n = 17), and E. hellem (n = 18) was determined by light microscopy; control xenografts remained uninfected. Extraintestinal infection with E. cuniculi developed in host mouse brain, respiratory tract, spleen, salivary glands, and gastrointestinal tract (3 of 3 mice), and infection with E. intestinalis developed in the liver (8 of 15 mice). CONCLUSION: Intestinal xenografts provide a unique, sterile, and biologically relevant animal model system for studying host enterocyte/parasite interactions, mechanisms of microsporidial pathogenicity, antimicrosporidial chemotherapeutic agents, and immune effector mechanisms. This model provides evidence for persistent graft infection with three Encephalitozoon spp., and for intestinal spread of E. cuniculi and E. intestinalis from infected enterocytes in immunoincompetent mice.

Immunohistochemical assay for detecting estrogen receptors in canine mammary tumors.

OBJECTIVE: To determine the estrogen receptor (ER) content of canine mammary gland tumors by use of immunohistochemical (IHC) examination of formalin-fixed sections. SAMPLE POPULATION: 21 mammary gland tumors from 20 adult dogs. PROCEDURE: ER were detected in formalin-fixed tissues, using an avidin-biotin alkaline phosphatase IHC assay and were quantified on fresh-frozen tumor samples, using a modified dextran-coated charcoal (DCC) assay. RESULTS: 7 of 21 tumors had visually detectable nuclear ER by use of IHC staining, whereas 8 of 21 tumors were positive for ER by use of the DCC assay. The ER-positive cells in 5 IHC-positive tumors were epithelial cells with histologic criteria of early malignancy. The remaining 2 ER-positive tumors detected by use of IHC had ER-positive mast cells within areas of connective tissue around the tumor. CONCLUSIONS: Immunohistochemistry is an additional method for detection of ER in canine mammary tumors. The major advantage of this type of assay is that it may be performed on formalin-fixed tissues, and individual ER-positive cells may be identified. Discovery of ER-positive mast cells by use of IHC is of concern, particularly if the ER status of a tumor is based on DCC results alone. CLINICAL RELEVANCE: Because most canine mammary tumors are fixed in formalin prior to histologic evaluation, an IHC assay that identifies ER-positive cells is desirable. Adjunctive antiestrogen therapy could be administered to dogs with ER-positive tumors.

Malnutrition modifies pig small intestinal inflammatory responses to rotavirus.

Infectious diarrheal diseases and malnutrition are major causes of child morbidity and mortality. In this study, malnutrition was superimposed on rotavirus infection in neonatal piglets to simulate the combined intestinal stress of viral enteritis in malnourished infants. Two-day-old piglets were assigned to three treatment groups as follows: 1) noninfected, fully nourished; 2) infected, fully nourished; and 3) infected, malnourished. Intestinal indices of inflammation were monitored over the subsequent 2-wk period. Intestinal damage and diarrhea were observed within 2 d of rotavirus infection and began to subside in nourished piglets by d 9 but persisted through d 16 postinfection in malnourished piglets. Rotavirus upregulated small intestinal expression of major histocompatibility complex (MHC) class I and class II genes; malnutrition intensified MHC class I gene expression and suppressed MHC class II expression. Jejunal CD4(+) and CD8(+) T-lymphocyte numbers were elevated for infected, nourished piglets on d 2, 9 and 16 postinfection. Malnutrition did not significantly affect the local expansion of T cell subsets in response to rotavirus. Intestinal prostaglandin E2 (PGE2) concentrations were elevated early after rotavirus infection independent of nutritional state. By d 9, PGE2 concentrations returned to baseline in infected, nourished piglets but remained elevated in malnourished piglets, corresponding to diarrhea observations. Together, the results identify intestinal indices of inflammation that are modulated by malnutrition and prompt reconsideration of current models of rotavirus pathophysiology.

Polymyalgia, hypersensitivity pneumonitis and other reactions in patients receiving HMG-CoA reductase inhibitors: a report of ten cases.

Ten patients who take hydroxy-methylglutaryl coenzyme A reductase inhibitors, or statin medications, and experience adverse reactions are described. All patients experienced various manifestations of hypersensitivity while receiving the drugs. One patient is described with hypersensitivity pneumonitis, which was graphically demonstrated by both high resolution computerized axial tomography and open lung biopsy.

Sialic acid dependence and independence of group A rotaviruses.

We have found (1), in contrast to previous reports, the human rotavirus Wa strain is sialic acid-dependent for binding to and infectivity of MA-104 cells and (2), a dual carbohydrate binding specificity is associated with both human Wa and Porcine OSU rotaviruses. One carbohydrate binding activity is associated with triple-layered virus particles (TLP) and the other with double-layered virus particles (DLP). In binding and infectivity studies, we found that gangliosides were the most potent inhibitors of both the human and procine rotavirus TLP. Furthermore, glycosylation mutant cells deficient in sialylation or neuraminidase-treated MA104 cells, did not bind rotavirus TLP from either strain. Our results show that human Wa binding and infectivity cannot be distinguished from the porcine OSU strain and appears to be sialic acid-dependent. Direct binding of human or porcine TLP to a variety of intact gangliosides was demonstrated in an thin-layer chromatographic (TLC) overlay assay. Human or porcine rotavirus DLP did not bind to any of the intact gangliosides but surprisingly bound asialogangliosides. This binding was abolished by prior treatment of the glycolipids with ceramide glycanase suggesting the intact asialoglycolipid was required for DLP binding. After treatment of either human or porcine TLP with EDTA to remove the outer shell, virus particles bound only to the immobilized asialogangliosides. These results suggest that rotavirus sugar binding specificity can be interpreted either as sialic acid-dependent or independent based on whether the virus preparation consists primarily of triple-layered or double-layered particles. Of perhaps greater interest is the possibility that sialic acid-independent carbohydrate binding activity plays a role in virus maturation or assembly.

Structure and function of a ganglioside receptor for porcine rotavirus.

A ganglioside fraction isolated from pooled intestines from newborn to 4-week-old piglets, which we previously partially characterized and showed to specifically inhibit the binding of porcine rotavirus (OSU strain) to host cells (M. D. Rolsma, H. B. Gelberg, and M. S. Kuhlenschmidt, J. Virol. 68:258-268, 1994), was further purified and found to contain two major monosialogangliosides. Each ganglioside was purified to apparent homogeneity, and their carbohydrate structure was examined by high-pH anion-exchange chromatography coupled with pulsed amperometric detection and fast atom bombardment mass spectroscopy. Both gangliosides possessed a sialyllactose oligosaccharide moiety characteristic of GM3 gangliosides. Compositional analyses indicated that each ganglioside was composed of sialic acid, galactose, glucose, and sphingosine in approximately a 1:1:1:1 molar ratio. Each ganglioside differed, however, in the type of sialic acid residue it contained. An N-glycolylneuraminic acid (NeuGc) moiety was found in the more polar porcine GM3, whereas the less polar GM3 species contained N-acetylneuraminic acid (NeuAc). Both NeuGcGM3 and NeuAcGM3 displayed dose-dependent inhibition of virus binding to host cells. NeuGcGM3 was approximately two to three times more effective than NeuAcGM3 in blocking virus binding. Inhibition of binding occurred with as little as 400 pmol of NeuGcGM3/50 ng of virus (approximately 2 x 10(7) virions) and 2 x 10(6) cells/ml. Fifty percent inhibition of binding was achieved with 0.64 and 1.5 microM NeuGcGM3 and NeuAcGM3, respectively. The free oligosaccharides 3'- and 6'-sialyllactose inhibited binding 50% at millimolar concentrations, which were nearly 1,000 times the concentration of intact gangliosides required for the same degree of inhibition. Direct binding of infectious, triple-layer rotavirus particles, but not noninfectious, double-layered rotavirus particles, to NeuGcGM3 and NeuAcGM3 was demonstrated by using a thin-layer chromatographic overlay assay. NeuGcGM3 and NeuAcGM3 inhibited virus infectivity of MA-104 cells by 50% at concentrations of 3.97 and 9. 84 microM, respectively. NeuGcGM3 (700 nmol/g [dry weight] of intestine) was found to be the predominant enterocyte ganglioside (comprising 75% of the total lipid-bound sialic acid) in neonatal piglets, followed by NeuAcGM3 (200 nmol/g [dry weight] of intestine). NeuGcGM3 and NeuAcGM3 together comprised nearly 100% of the lipid-bound sialic acid in the neonatal intestine, but their quantities rapidly diminished during the first 5 weeks of life. These data support the hypothesis that porcine NeuGcGM3 and NeuAcGM3 are physiologically relevant receptors for porcine rotavirus (OSU strain). Further support for this hypothesis was obtained from virus binding studies using mutant or neuraminidase-treated cell lines. Lec-2 cells, a mutant clone of CHO cells characterized by a 90% reduction in sialyllation of its glycoconjugates, bound less than 5% of the virus compared to control cell binding. In contrast, Lec-1 cells, a mutant CHO clone characterized by a deficiency in glycosylation of N-linked oligosaccharides, still bound rotavirus. Furthermore, exogenous addition of NeuGcGM3 to the Lec-2 mutant cells restored their ability to bind rotavirus in amounts equivalent to that of their parent (CHO) cell line. In the virus-permissive MA-104 cell line, NeuGcGM3 was also able to partially restore rotavirus infectivity in neuraminidase-treated cells. These data suggest that gangliosides play a major role in recognition of host cells by porcine rotavirus (OSU strain).

Protein-energy malnutrition delays small-intestinal recovery in neonatal pigs infected with rotavirus.

Infectious diarrheal diseases and protein-energy malnutrition (PEM) are major causes of child morbidity and mortality worldwide. In the present study, PEM was superimposed on rotavirus infection in neonatal pigs to simulate chronic small intestinal stress in malnourished infants with viral gastroenteritis. Two-day-old cesarean-derived pigs (n = 39) were allotted to three treatment groups: 1) noninfected, full-fed; 2) infected, full-fed; and 3) infected, malnourished. Two days postinfection, severe diarrhea and weight loss (11%) were accompanied by reductions in villus height (60%) and lactase activity (78%) and increased crypt depth (32%) in infected full-fed compared with noninfected pigs (P < 0.05). Malnutrition blunted (P < 0.05) increases in crypt depth elicited by rotavirus. By 9 d postinfection, body weight was 59% less, villus height and lactase activity remained lower (50%), and crypt depth remained greater (62%) in infected full-fed compared with noninfected pigs (P < 0.05). However, diarrhea began to clear in infected full-fed, but not in infected malnourished pigs. Plasma insulin-like growth factor-I (IGF-I) was reduced 68% and crypt depth was reduced 19% in infected-malnourished compared with infected full-fed pigs (P < 0.05). Sixteen days postinfection, full-fed pigs had recovered from rotaviral infection; however, in infected-malnourished pigs, diarrhea and growth stasis persisted, and plasma IGF-I, villus height and alkaline phosphatase activity remained reduced compared with infected full-fed pigs (P < 0.05). Overall, PEM prolonged diarrhea and delayed small-intestinal recovery, indicating that nutritional status during diarrhea is essential for recovery from rotaviral enteritis.

Application of intestinal xenografts to the study of enteropathogenic infectious disease.

We have developed, characterized and utilized paired segments of fetal intestine subcutaneously transplanted into heterogenic nude or SCID mice as a model system for the study of viral, bacterial and protozoal pathogens. The xenografted intestine matures in the recipient mouse and is biochemically and anatomically comparable to intestine from age-matched, whole-animal controls. The grafted tissue is free of ingesta, intestinal flora, extra-intestinal secretions and host immune functions. The transplanted intestine is long-lived and easily accessible to manipulation and harvest. Tissue from a single fetal donor can be used to create numerous xenografts allowing for tightly controlled experiments. Xenografts enable the study of species-specific intestinal pathogens in the homologous intestinal tissue thus preserving biological applicability of results. Xenografts can be used to study pathogenesis, pathophysiology and therapeutics of enteric disease in situations where such study might otherwise be prohibitively expensive or confounded by intercurrent variables inherent to whole animals. Xenografts have important advantages over in vitro models that may not approximate the in vivo biology of the intestine in the disease process.

Characterization of a porcine enterocyte receptor for group A rotavirus.

We have identified, purified to apparent homogeneity and chemically characterized a biologically-relevant porcine enterocyte receptor for group A porcine rotavirus. Ceramide glycanase digestion followed by acid hydrolysis and monosaccharide compositional analyses indicated the receptor is a family of two GM, gangliosides, one containing N-glycolyl-neuraminic acid and the other N-acetylneuraminic acid. Both gangliosides displayed dose-dependent inhibition of rotavirus binding to, and infectivity of, host cells. Inhibition of infectivity in a focus-forming-unit-reduction assay was achieved with as little as 2 nmols of NeuGcGM3 (50% inhibition with 3.97 nmol) or NeuAcGM3 (50% inhibition with 9.84 nmol) per 10(4) FFU of virus. Preliminary data suggest specific porcine GM3 carbohydrate fine structure or spatial orientation of the sialyloligosaccharide epitopes of the holoGM3 gangliosides may be crucial to enterocyte receptor recognition by rotavirus. We have quantified both NeuGcGM3 and NeuAcGM3 in enterocytes of various-aged pigs from newborn through 16 weeks and have found with increasing age the amount of both GM3 derivatives, especially NeuGcGM3 per gram (dry weight) intestinal brush border decreases rapidly from newborn through 4 weeks of age. These results may help explain the age-sensitivity of piglets to severe rotavirus diarrhea.