First report on natural infection with Leishmania infantum in a domestic ferret (Mustela putorius furo) in Spain.

A pet domestic ferret (Mustela putorius furo) with a papular lesion involving the right pinna was diagnosed with chronic pyogranulomatous dermatitis by histopathologic examination. Intralesional, intracytoplasmic oval microorganisms compatible with Leishmania spp. or Histoplasma spp. were observed in macrophages and multinucleate giant cells. Leishmania infantum (L. infantum) infection was diagnosed by PCR, culture in Novy-MacNeal-Nicolle medium, and immunohistochemistry. Abnormal clinicopathological results included increased alanine transferase, alkaline phosphatase, serum gamma glutamyl transferase and polyclonal gammpathy. Anti-Leishmania antibodies were detected by enzyme-linked immunosorbent assay, immunofluorescence antibody test and western blot using L. infantum antigen. Immunoreactivity against the 16 kDa specific L. infantum antigen fraction was observed by western blot. PCR performed in blood samples obtained from this patient after positive parasite isolation detected L. infantum DNA. To the authors' knowledge, this is the first diagnosis and isolation of L. infantum in a domestic ferret naturally infected in an endemic region (Spain) where canine and feline leishmaniosis is frequently detected. According to these findings, ferrets should be included as potential reservoir hosts of L. infantum. Future investigations should analyze the epidemiological role of ferrets in L. infantum infection including the prevalence of infection.

Encephalitozoon hellem infection in aviary passerine and psittacine birds in Spain.

A European goldfinch (Carduelis carduelis), a canary (Serinus canaria), and a lovebird (Agapornis roseicollis) captive-bred at three different private aviaries in Spain were submitted for necropsy with a history of weakness and ruffled feathers, weight loss associated with glossitis, and respiratory disease, respectively. Microscopically, enterocytes in the jejunum and ileum contained colonies of gram- and Stamp-positive, oval to elliptical microorganisms within parasitophorous vacuoles in the apical cytoplasm. Nested PCR using MSP primers that target microsporidian RNA genes produced amplicons of expected size for Encephalitozoon species, and analysis of forward and reverse DNA sequences confirmed the presence of Encephalitozoon hellem in all cases. The main cause of death of all three birds consisted of concurrent infections. However, intestinal encephalitozoonosis may have contributed to exacerbated catabolism. Encephalitozoonosis (or microsporidiosis) has been rarely described in passerine birds.

Immunohistochemical characterization of nonhuman primate ovarian sex cord-stromal tumors.

This study evaluates the immunoreactivity of 12 sex cord-stromal tumors of nonhuman primates (11 granulosa cell tumors and 1 luteoma). The markers selected are used in the characterization of gonadal tumors in dogs and other species, including cytokeratins AE1/AE3, GATA-4, inhibin-α, neuron-specific enolase, protein gene product 9.5, and vimentin. A normal nonhuman primate ovary was used as a control and to optimize immunolabeling. Staining was graded as follows: 0 (nonstaining), 1+ (< 10% positive cells), 2+ (10%-50% positive cells), and 3+ (> 50% positive cells). Calretinin, GATA-4, neuron-specific enolase, and vimentin were the most consistently expressed markers (12 of 12). Cytokeratins AE1/AE3 were also consistently expressed (11 of 12). Inhibin-α and protein gene product 9.5 were expressed in 8 and 10 sex cord-stromal tumors, respectively. Results indicate that immunoreactivity of nonhuman primate sex cord-stromal tumors is similar to that observed in other species and that calretinin, GATA-4, and neuron-specific enolase are the most consistently expressed markers in nonhuman primate sex cord-stromal tumors.

Pyogranulomatous pleuropneumonia and mediastinitis in ferrets (Mustela putorius furo) associated with Pseudomonas luteola Infection.

Between 2008 and 2009, three pet ferrets from different sources presented with acute episode of dyspnoea. Cytological examination of pleural exudates revealed severe purulent inflammation with abundant clusters of rod-shaped microorganisms with a clear surrounding halo. Treatment was ineffective and the ferrets died 2-5 days later. Two ferrets were subjected to necropsy examination, which revealed pyothorax, mediastinal lymphadenopathy and multiple white nodules (1-2mm) in the lungs. Microscopical examination showed multifocal necrotizing-pyogranulomatous pleuropneumonia and lymphadenitis with aggregates of encapsulated microorganisms, some of which were positively stained by periodic acid-Schiff and alcian blue. In-situ hybridization for Pneumocystis spp., Ziehl-Neelsen staining and immunohistochemistry for distemper, coronavirus and influenza antigen were negative in all cases. Electron microscopically, the bacteria were 2-3 µm long with a thick electron-lucent capsule. Microbiology from one ferret yielded a pure culture of gram-negative bacteria identified phenotypically as Pseudomonas luteola. This speciation was later confirmed by 16S RNA gene amplification.

Outbreak of mortality in psittacine birds in a mixed-species aviary associated with Erysipelothrix rhusiopathiae infection.

Erysipelothrix rhusiopathiae septicemia, associated with an increased mortality of captive psittacines in a mixed-species aviary, was diagnosed by histopathology, Gram staining, bacterial culture and sequencing, immunohistochemistry, and real-time polymerase chain reaction (PCR). Over a period of 23 days with no premonitory signs, 2 rainbow lorikeets and an eclectus parrot died. Of these birds, one lorikeet and the eclectus were submitted for necropsy. The main pathologic findings were thrombosis (2/2), bacterial embolism/thromboembolism (2/2), necrotizing hepatitis (2/2), necrohemorrhagic myocarditis (1/2), fibrinohemorrhagic and heterophilic visceral coelomitis (1/2), submandibular necrosuppurative dermatitis with necrotizing vasculitis and bacterial and fungal thromboembolism (1/2), and locally extensive rhabdomyonecrosis with bacterial embolism (1/2). Intralesional bacteria were positive by Gram staining and immunohistochemistry in both cases. E. rhusiopathiae was isolated by routine bacterial culture from the liver of the lorikeet, which was also positive by real-time PCR. This report is one of the rare descriptions of erysipelas in psittacines, and to the authors' knowledge, it appears to be the first in the described species using immunohistochemistry and real-time PCR on avian paraffin-embedded tissues for the diagnosis.

Congestive heart failure in 6 African grey parrots (Psittacus e erithacus).

Six African grey parrots (Psittacus e erithacus) were diagnosed with cardiomyopathy and congestive heart failure based on gross and microscopic findings. Ages ranged from 15 days to 8 years, and 5 of 6 parrots were either neonates or juveniles at the time of diagnosis. Two neonates and 2 juveniles came from the same breeding aviary; the 2 juveniles were born to the same breeding pair. The 2 other parrots were kept as pets. Clinical signs included distention of the coelomic cavity (4 of 6), rales (3 of 6), weakness (4 of 6), bradyarrhythmia (1 of 6), growth retardation (1 of 6), crop stasis (1 of 6), and regurgitation (1 of 6). Three parrots were euthanized and 3 died. Gross findings included cardiomegaly due to biventricular, right-, or left-sided cardiomyopathy (6 of 6); coelomic effusion (6 of 6); whitish or yellow foci in the liver (6 of 6); atrophy of the liver (particularly, the left lobe; 5 of 6); reddened or grey lungs (5 of 6); subcutaneous edema (2 of 6); hydropericardium (1 of 6); and bilateral thyroid gland enlargement (1 of 6). Relevant microscopic findings included passive hepatic congestion (6 of 6) and pulmonary congestion (2 of 6), lymphocytic thyroiditis (2 of 6), and diffuse thyroid follicular hyperplasia (2 of 6). Microscopically, the heart was unremarkable (2 of 6) or had mild lymphocytic myocarditis (2 of 6), mild multifocal cytoplasmic vacuolation of cardiomyocytes (2 of 6), mild lymphocytic myocardial (Purkinje cell) ganglioneuritis (1 of 6), and mild multifocal interstitial fibrosis and nuclear hypertrophy of cardiomyocytes (1 of 6). One parrot had concurrent proventricular dilatation disease (systemic ganglioneuritis). The cause of cardiomyopathy in these parrots was not determined.

Identification of the source of histoplasmosis infection in two captive maras (Dolichotis patagonum) from the same colony by using molecular and immunologic assays.

Histoplasma capsulatum was isolated from the spleen of a first infected mara (Dolichotis patagonum) and from a second mara's liver and adrenal gland, both in the same colony at the Africam Safari, Puebla, Mexico. Studies of H. capsulatum isolates, using nested-PCR of a 100-kDa protein coding gene (Hcp100) fragment and a two-primer RAPD-PCR method, suggest that these isolates were spreading in the environment of the maras' enclosure. By using a Dot-ELISA method, sera from mice inoculated with three homogenates of soil samples from the maras' enclosed space developed positive brown spot reactions to a purified H. capsulatum antigen, which identified the probable source of the maras' infection.

Second case of histoplasmosis in a captive mara (Dolichotis patagonum): pathological findings.

A second case of histoplasmosis in a captive mara (Dolichotis patagonum) from a colony at the wildlife park Africam Safari, Puebla, Mexico, is described, and the mara died with disseminated clinical form of the disease, affecting mostly the large intestine and adrenal. The pathological findings of this case 2 revealed severe granulomatous typhlocolitis and moderate granulomatous gastrohepatic lymphadenitis with numerous yeast-like cells, 2-4 mum in diameter, with a clear halo surrounding them inside the cytoplasm of macrophages, suggesting the parasitic form of Histoplasma capsulatum. Adrenocortical cells had abundant similar microorganisms in their cytoplasm without any associated lesion. Gomori's methenamine silver and periodic acid Schiff stained positively these microorganisms. Immunohistochemistry, using a rabbit anti-H. capsulatum serum, and transmission electron microscopy supported the diagnosis of H. capsulatum infection.

Pheochromocytoma in six New World primates.

Six New World primates, including 2 golden lion tamarins (Leontopithecus rosalia), 2 cotton-top tamarins (Saguinus o. oedipus), 1 black howler monkey (Alouatta caraya), and 1 black-handed spider monkey (Ateles g. geoffroyi), were diagnosed with unilateral (4/6) or bilateral (1/6) adrenal or extra-adrenal (1/6) pheochromocytoma by light microscopy and immunohistochemical staining for chromogranin A. Overt invasive behavior or metastases were not observed in any primate, and thus these neoplasms were considered benign. All primates either died spontaneously (4/6) or were euthanatized (2/6) as a result of concurrent malignant neoplasia, infection, renal disease, or a combination of several disease processes. Although we did not determine whether these pheochromocytomas were functional, all 6 primates had myocardial fibrosis, and some had arteriosclerosis.

Clinicopathologic features of a systemic coronavirus-associated disease resembling feline infectious peritonitis in the domestic ferret (Mustela putorius).

From 2002 to 2007, 23 ferrets from Europe and the United States were diagnosed with systemic pyogranulomatous inflammation resembling feline infectious peritonitis (FIP). The average age at the time of diagnosis was 11 months. The disease was progressive in all cases, and average duration of clinical illness was 67 days. Common clinical findings were anorexia, weight loss, diarrhea, and large, palpable intra-abdominal masses; less frequent findings included hind limb paresis, central nervous system signs, vomiting, and dyspnea. Frequent hematologic findings were mild anemia, thrombocytopenia, and hypergammaglobulinemia. Grossly, whitish nodules were found in numerous tissues, most frequently the mesenteric adipose tissue and lymph nodes, visceral peritoneum, liver, kidneys, spleen, and lungs. One ferret had a serous abdominal effusion. Microscopically, pyogranulomatous inflammation involved especially the visceral peritoneum, mesenteric adipose tissue, liver, lungs, kidneys, lymph nodes, spleen, pancreas, adrenal glands, and/or blood vessels. Immunohistochemically, all cases were positive for coronavirus antigen using monoclonal antibody FIPV3-70. Electron microscopic examination of inflammatory lesions identified particles with coronavirus morphology in the cytoplasm of macrophages. Partial sequencing of the coronavirus spike gene obtained from frozen tissue indicates that the virus is related to ferret enteric coronavirus.

Disseminated encephalitozoonosis in captive, juvenile, cotton-top (Saguinus oedipus) and neonatal emperor (Saguinus imperator) tamarins in North America.

Disseminated encephalitozoonosis was diagnosed in 2 sibling, juvenile, cotton-top tamarins (Saguinus oedipus) and 3 sibling, neonatal, emperor tamarins (S. imperator) by use of histologic examination, histochemical analysis, electron microscopy, and polymerase chain reaction (PCR) analysis with nucleotide sequencing. All tamarins were captive born at zoos in North America and died with no premonitory signs of disease. The main pathologic findings were myocarditis (4/5), hepatitis (3/5), interstitial pneumonia (3/5), skeletal myositis (3/5), meningoencephalitis (2/5), adrenalitis (2/5), tubulointerstitial nephritis (1/5), myelitis (1/5), sympathetic ganglioneuritis (1/5), and retinitis (1/5). Central nervous system lesions were the most prominent findings in cotton-top tamarins. The inflammation was predominantly lymphocytic and suppurative in cotton-top tamarins, whereas emperor tamarins had granulomatous or lymphoplasmacytic lesions. Intralesional periodic acid-Schiff-, gram-, or acid-fast (or all 3)-positive, oval-to-elliptical shaped organisms were found in 1 cotton-top and the 3 emperor tamarins. By electron microscopy, these organisms were consistent with microsporidia of the genus Encephalitozoon. E. cuniculi genotype III was detected by PCR analysis and sequencing in paraffin-embedded brain, lung, and bone marrow specimens from the cotton-top tamarins. Although PCR results were negative for one of the emperor tamarins, their dam was seropositive for E. cuniculi by ELISA and Western blot immunodetection. These findings and recent reports of encephalitozoonosis in tamarins in Europe suggest that E. cuniculi infection may be an emerging disease in callitrichids, causing high neonatal and juvenile mortality in some colonies. The death of 2 less than 1-day-old emperor tamarins from a seropositive dam supports the likelihood of vertical transmission in some of the cases reported here.

Pathological findings in a captive colony of maras (Dolichotis patagonum).

This paper describes the causes of death of 54 maras (Dolichotis patagonum) in a captive colony in Mexico over a period of seven years. There were 35 adults, 11 juveniles, five neonates, two fetuses and one stillbirth--27 males, 21 females and six whose sex was not determined. Trauma was the cause of 25 deaths, and there were eight cases of fatal bacterial infection. Besnoitiosis was the only parasitic disease found frequently (13 cases), and was associated with fatal interstitial pneumonia in three juveniles. Right-sided hypertrophic cardiomyopathy attributed to high altitude was observed in 26 maras, and in three cases death was attributed to acute cardiac dysfunction. Two maras died of disseminated histoplasmosis and two of hyperthermia. Additional causes of death included one case each of uterine torsion, intestinal intussusception, aspiration pneumonia and hydranencephaly. Gastric erosions with luminal haemorrhage were found in 27 of the maras and splenic lymphoid depletion in 20, changes that were attributed to stress.

Fatal placental subinvolution in a captive capybara (Hydrochaeris hydrochaeris, Order Rodentia).

An adult, captive-born female capybara died of systemic thrombosis and hemoperitoneum associated with placental subinvolution. Grossly, the uterus was enlarged, segmentally thickened, and associated with a large blood clot in the abdominal cavity. There was hemometra and a large ovoid mass in each uterine horn weakly adhered to the endometrium, and the right uterine horn wall had a small perforation over the mass. The mesometrial veins were markedly dilated due to thrombosis and occasionally perforated. Histologically, the uterine masses consisted of partly necrotic placental and subplacental tissue. The uterine wall surrounding the masses had full-thickness coagulative necrosis of the myometrium and diffuse endometrial ulceration with abundant syncytiotrophoblast-like cells within capillaries. Vascular lesions in the uterus and mesometrium consisted of mural invasion by cytotrophoblast and syncytiotrophoblast-like cells, thrombosis, fibrinoid necrosis, and/or heterophilic vasculitis. This is the first report of placental subinvolution in capybaras or any rodent species, to the authors' knowledge.

Pulmonary besnoitiasis in captive maras (Dolichotis patagonum) associated with interstitial pneumonia.

Six captive maras (Dolichotis patagonum) were found to have cysts consistent with Besnoitia in the lungs by light microscopy and electron microscopy. Three were juveniles that died with no premonitory signs during a 17-month period. The most prominent finding consisted of severe, subacute, diffuse interstitial pneumonia with syncytia and Besnoitia cysts. The severity of pneumonia correlated with the number of cysts and the presence of lytic cysts, often surrounded by chronic granulomatous inflammation. Disseminated infection was observed in one of these maras. The three other maras died of unrelated conditions and had a few Besnoitia cysts in their lungs with mild or no inflammation associated. This is the first report of besnoitiasis in maras and of its association with fatal interstitial pneumonia in any species. Although other agents may have caused the interstitial pneumonia described here, it is possible that zoites released from lytic cysts were involved in the development of this lesion.

Anemia, myopathy, and pansteatitis in vitamin E-deficient captive marmosets (Callithrix spp.).

Five young adult pet marmosets (Callithrix spp.) were presented with weight loss (5/5); fecal retention (3/5); diarrhea (2/5); impaired locomotion (3/5); anemia (4/4); hypoproteinemia or hypoalbuminemia (3/4); elevations of creatine phosphokinase, lactic dehydrogenase, and alanine aminotransferase (3/4); and renal failure with hypercholesterolemia (2/4). All anemic marmosets had low serum vitamin E levels. The anemia responded to vitamin E and selenium therapy in two marmosets. One of the five marmosets died before presentation, and two others died despite therapy. The two marmosets necropsied had degenerative myopathy, pyogranulomatous pansteatitis, and increased erythrophagocytosis and hemosiderosis. The striated muscle and adipose tissue of both marmosets were negative for coxsackievirus ribonucleic acid by in situ hybridization. These findings suggest that vitamin E deficiency may be involved in the development of anemia, myopathy, and steatitis in callitrichids; however, in some marmosets, underlying diseases such as chronic colitis may have influenced the development of anemia and impaired vitamin E status.

Outbreak of salmonellosis in a zoologic collection of lorikeets and lories (Trichoglossus, Lorius, and Eos spp.).

During August 2001, a syndrome characterized by acute lethargy and dyspnea was observed in a population of 45 lorikeets and lories in an open-air zoologic exhibit. The first death occurred on August 10, and within the next 12 days, nine more birds died (22% mortality rate). Hepatomegaly, reddening and congestion of the lungs, and injection of the serosal surface of the intestines were the common gross findings. Histologic changes, including fibrinonecrotic hepatitis and splenitis, bacterial emboli (liver, spleen, lung, kidney, proventriculus), pulmonary congestion and hemorrhage, and enteritis, were indicative of an acute, overwhelming bacterial septicemia. Salmonella typhimurium, with the same antibiogram, was isolated from four birds. Several birds had attacked and killed a snake on July 24, and Salmonella serogroup B (untypeable) was isolated from intestine and kidney samples of a garter snake caught in the open-air exhibit on August 28. Salmonella was also isolated from environmental samples of the exhibit but not from food preparation areas. After antimicrobial therapy, Salmonella spp. was not isolated from the surviving birds. The source of Salmonella in this outbreak remains unknown, but infection either directly or indirectly from snakes in the exhibit is possible. Contact between captive psittacine populations and reptiles should be avoided to prevent the risk of salmonellosis.

Disseminated toxoplasmosis in a Mediterranean pregnant Risso's dolphin (Grampus griseus) with transplacental fetal infection.

Fatal disseminated toxoplasmosis was diagnosed in a Risso's dolphin (Grampus griseus) dam and its fetus on the basis of pathologic findings, immunohistochemistry, and structure of the parasite. The dolphin was stranded alive on the Spanish Mediterranean coast and died a few hours later. At necropsy the dam was in good condition. From the standpoint of pathology, however, it had generalized lymphadenomegaly and splenomegaly, enlargement of and multifocal hemorrhage in the adrenal glands, diffuse mucosal hemorrhage of the glandular and pyloric stomach, ulcerative glossitis and stomatitis, focal erosions and reddening of the laryngeal appendix, and severe paraotic sinusitis with intralesional nematodes Crassicauda grampicola. The dolphin was pregnant, most probably in the first gestational trimester. The most prominent microscopic lesions were multifocal granulomatous encephalomyelitis, diffuse subacute interstitial pneumonia, mild multifocal necrotizing hepatitis and nonsuppurative cholangiohepatitis, gastritis and adrenalitis, mild lymphoid depletion, medullary sinus and follicular histyocitosis, and systemic hemosiderosis. The fetus had foci of coagulative and lytic necrosis in the kidneys, the lung, and the heart. Most lesions were associated with tachyzoites and tissue cysts of Toxoplasma gondii. The diagnosis was confirmed immunohistochemically. This is the first report on toxoplasmosis in a Risso's dolphin (G. griseus) and on transplacental transmission to an early-stage fetus in any cetaceans.