Obstrução nasal por granuloma fúngico em eqüino: relato de caso / Nasal obstruction caused by fungal granuloma in a horse: case report

Um eqüino de nove anos de idade apresentou ausência de ar expirado e secreção serossanguinolenta na narina direita, associado a ruído respiratório. Os exames endoscópico e radiológico mostraram uma formação de aproximadamente seis centímetros de diâmetro recoberta por mucosa amarelada, que obstruía a cavidade nasal direita e insinuava-se para a cavidade nasal esquerda. Tal massa foi ressecada por meio de sinusotomia frontal direita. O exame histológico e a cultura revelaram lesão granulomatosa causada por fungos. O tratamento pós-operatório compreendeu associação de antibiótico e antiinflamatório, assim como de lavagens com água destilada e chá de camomila.

A 9-year-old horse presented serosanguineous nasal discharge, absence of breath out through the right nostril, and respiratory noise. Endoscopic and radiographic exams revealed a six centimeter diameter mass, covered by yellowish mucosa, which was obstructing the entire right nasal cavity and part of the left one. The mass was excised through a right frontal sinusotomy. The microscopic exam and the culture revealed a fungic granulomatous rhinitis. Antibiotic and anti-inflammatory drugs were postoperatively administered; moreover, camomile tea and distilled water were flushed in a drain placed above the bone flap.

Morphological changes in mouse embryos cryopreserved by different techniques.

Cryopreservation of mammalian embryos is an important tool for the application of reproductive biotechnologies. Subjective evaluation to determine embryo viability is often used. The determination of the best cryopreservation protocol depends on morphological and molecular analysis of cellular injuries. The main objective of this study was to compare two methods of cryopreservation by assessing morphological alterations of frozen embryos using light, fluorescence, and transmission electron microscope. Fresh (control), slow frozen, and vitrified mouse embryos were composed. To evaluate the viability of the embryos, the cell membrane integrity was assessed using Hoechst33342 and propidium iodide (H/PI) staining. Morphological analyses using hematoxylin and eosin (HE) staining were performed to test different techniques (in situ, paraffin, and historesin) by both light and fluorescence microscopy. Transmission electron microscope was used to detect ultrastructural alterations in Spurr- and Araldite-embedded samples. H/PI staining detected more membrane permeability in the vitrification (69.8%) than in the slow freezing (48.4%) or control (13.8%) groups (P < 0.001). Historesin-embedded samples showed to be more suitable for morphological analyses because cellular structures were better identified. Nuclear evaluation in historesin sections showed the induction of pycnosis in slow freezing and vitrification groups. Cytoplasm evaluation revealed a condensation and an increase in eosinophilic intensity (indicating apoptosis) in the slow freezing group, and weakly eosinophilic structures and degenerated cells (indicating oncosis) in the vitrification group (P < 0.05). Ultrastructural analyses confirmed HE morphological findings. It was concluded that both cryopreservation techniques resulted in oncosis and apoptosis injuries. However, vitrification caused more severe cellular alterations and reduced embryonic viability compared to slow freezing.

Immunopathology of rabies infection in mice selected for high or low acute inflammatory reaction

Rabies is a severe and lethal disease that produces a slight inflammatory response during the infection process. We analyzed the immunopathological mechanisms that occur in the central nervous system (CNS) using mice genetically selected for maximal or minimal acute inflammatory reaction (AIRmax or AIRmin). As viral samples, we adopted the antigenic variant 3 (AgV3) of rabies virus from hematophagous bats and a fixed virus strain (PV1 43/3). Titration of specific antibodies was performed using enzyme-linked immunosorbent assay (ELISA). We observed a slight increase in IgG and IgG1 isotypes in infected AIRmax mice. Incubation period, determined by intracerebral inoculation with 100 LD50, was 6-7 days for PV1 43/4 strain and 9-10 days for AgV3. No difference in viral replication was noticed between AIRmax and AIRmin mice. Mortality was 100 percent with both viral strains. Histopathological analysis of brains and spinal cords showed inflammatory foci in all regions of the CNS. No differences were noticed in the number of neutrophils. Negri bodies were observed in practically all sites analyzed. Results suggested that inflammatory reaction is not a determining factor in the susceptibility to rabies infection.

Histopathological and ultrastructural characteristics of myeloid leukosis in broiler chicken

An ultrastructural and histological study was performed to determine the degree of differentiation of the neoplastic cells. The histological study revealed neoplastic cells with pleomorphism, oval nuclei, prominent nucleoli, irregularly distributed chromatin, atypical mitotic figures and moderate amount of cytoplasm containing spherical eosinophilic granulations, typical features of the myeloid lineage. Ultrastructurally, there were cells with an electron-dense, oval and voluminous nucleus, with predominant euchromatin and cytoplasm containing many spherical, electron-dense and homogeneous granules, indicative of myelocytes with differentiation to eosinophils. Type-C viral particles were also seen in the intercellular space of renal tubules and inside the intracytoplasmic vesicles of immature myelocytes in the bone marrow and ovary. PCR was positive to ALV-J.

Caracterizaram-se a linhagem e o grau de diferenciação das células neoplásicas no estudo histopatológico e ultraestrutural da leucose mielóide. Histologicamente as células neoplásicas apresentaram pleomorfismo, núcleos ovais, nucléolos proeminentes, cromatina distribuída de maneira irregular, figuras de mitose atípicas e moderada quantidade de citoplasma contendo granulações eosinofílicas esféricas. Essas características indicam a linhagem mielóide. Ultraestruturalmente evidenciaram-se células com núcleo oval, volumoso, eletrodenso, com predomínio de eucromatina e citoplasma com numerosos grânulos esféricos, eletrodensos e homogêneos, indicando mielócitos com diferenciação para eosinófilos. Constatou-se também a presença de partículas virais tipo-C no espaço intercelular dos túbulos renais, no interior de vesículas intracitoplasmáticas dos mielócitos imaturos presentes na medula óssea e ovário, e PCR positivo para ALV-J.

Delayed Schwann cell and oligodendrocyte remyelination after ethidium bromide injection in the brainstem of Wistar rats submitted to streptozotocin diabetogenic treatment.

Schwann cell disturbance followed by segmental demyelination in the peripheral nervous system occurs in diabetic patients. Since Schwann cell and oligodendrocyte remyelination in the central nervous system is a well-known event in the ethidium bromide (EB) demyelinating model, the aim of this investigation was to determine the behavior of both cell types after local EB injection into the brainstem of streptozotocin diabetic rats. Adult male Wistar rats received a single intravenous injection of streptozotocin (50 mg/kg) and were submitted 10 days later to a single injection of 10 microL 0.1% (w/v) EB or 0.9% saline solution into the cisterna pontis. Ten microliters of 0.1% EB was also injected into non-diabetic rats. The animals were anesthetized and perfused through the heart 7 to 31 days after EB or saline injection and brainstem sections were collected and processed for light and transmission electron microscopy. The final balance of myelin repair in diabetic and non-diabetic rats at 31 days was compared using a semi-quantitative method. Diabetic rats presented delayed macrophage activity and lesser remyelination compared to non-diabetic rats. Although oligodendrocytes were the major remyelinating cells in the brainstem, Schwann cells invaded EB-induced lesions, first appearing at 11 days in non-diabetic rats and by 15 days in diabetic rats. Results indicate that short-term streptozotocin-induced diabetes hindered both oligodendrocyte and Schwann cell remyelination (mean remyelination scores of 2.57 +/- 0.77 for oligodendrocytes and 0.67 +/- 0.5 for Schwann cells) compared to non-diabetic rats (3.27 +/- 0.85 and 1.38 +/- 0.81, respectively).

Delayed Schwann cell and oligodendrocyte remyelination after ethidium bromide injection in the brainstem of Wistar rats submitted to streptozotocin diabetogenic treatment

Schwann cell disturbance followed by segmental demyelination in the peripheral nervous system occurs in diabetic patients. Since Schwann cell and oligodendrocyte remyelination in the central nervous system is a well-known event in the ethidium bromide (EB) demyelinating model, the aim of this investigation was to determine the behavior of both cell types after local EB injection into the brainstem of streptozotocin diabetic rats. Adult male Wistar rats received a single intravenous injection of streptozotocin (50 mg/kg) and were submitted 10 days later to a single injection of 10 æL 0.1 percent (w/v) EB or 0.9 percent saline solution into the cisterna pontis. Ten microliters of 0.1 percent EB was also injected into non-diabetic rats. The animals were anesthetized and perfused through the heart 7 to 31 days after EB or saline injection and brainstem sections were collected and processed for light and transmission electron microscopy. The final balance of myelin repair in diabetic and non-diabetic rats at 31 days was compared using a semi-quantitative method. Diabetic rats presented delayed macrophage activity and lesser remyelination compared to non-diabetic rats. Although oligodendrocytes were the major remyelinating cells in the brainstem, Schwann cells invaded EB-induced lesions, first appearing at 11 days in non-diabetic rats and by 15 days in diabetic rats. Results indicate that short-term streptozotocin-induced diabetes hindered both oligodendrocyte and Schwann cell remyelination (mean remyelination scores of 2.57 ± 0.77 for oligodendrocytes and 0.67 ± 0.5 for Schwann cells) compared to non-diabetic rats (3.27 ± 0.85 and 1.38 ± 0.81, respectively).

The role of complement in the early phase of Leishmania (Leishmania) amazonensis infection in BALB/c mice.

Complement-depleted and -non-depleted BALB/c mice were inoculated with Leishmania (Leishmania) amazonensis promastigotes into the hind footpad to study the role of the complement system in cutaneous leishmaniasis. Total serum complement activity was measured by hemolytic assay and C3 fragment deposit at the inoculation site was determined by direct immunofluorescence in the early period of infection, i.e., at 3, 24, 48 h and 7 days post-infection. The inflammatory reaction and the parasite burden were evaluated in the skin lesion at 7 and 30 days post-infection. Total serum complement activity decreased in the early phase of infection, from 3 to 24 h, in non-depleted mice compared to non-infected and non-depleted mice. C3 fragment deposit at the site of parasite inoculation was present throughout the period of infection in non-depleted mice. In contrast, no C3 fragment deposit was observed at the inoculation site in complement-depleted mice. Complement-depleted mice showed a significant decrease in the inflammatory response and a significant increase in the number of parasites (70.0 +/- 5.3 vs 5.3 +/- 1.5) at 7 days of infection (P<0.05). A higher number of parasites were also present at 30 days of infection at the inoculation site of complement-depleted mice (78.5 +/- 24.9 vs 6.3 +/- 5.7). These experiments indicate that complement has an important role at the beginning of experimental cutaneous leishmaniasis caused by L. (L.) amazonensis by controlling the number of parasites in the lesion.

The role of complement in the early phase of Leishmania (Leishmania) amazonensis infection in BALB/c mice

Complement-depleted and -non-depleted BALB/c mice were inoculated with Leishmania (Leishmania) amazonensis promastigotes into the hind footpad to study the role of the complement system in cutaneous leishmaniasis. Total serum complement activity was measured by hemolytic assay and C3 fragment deposit at the inoculation site was determined by direct immunofluorescence in the early period of infection, i.e., at 3, 24, 48 h and 7 days post-infection. The inflammatory reaction and the parasite burden were evaluated in the skin lesion at 7 and 30 days post-infection. Total serum complement activity decreased in the early phase of infection, from 3 to 24 h, in non-depleted mice compared to non-infected and non-depleted mice. C3 fragment deposit at the site of parasite inoculation was present throughout the period of infection in non-depleted mice. In contrast, no C3 fragment deposit was observed at the inoculation site in complement-depleted mice. Complement-depleted mice showed a significant decrease in the inflammatory response and a significant increase in the number of parasites (70.0 ± 5.3 vs 5.3 ± 1.5) at 7 days of infection (P < 0.05). A higher number of parasites were also present at 30 days of infection at the inoculation site of complement-depleted mice (78.5 ± 24.9 vs 6.3 ± 5.7). These experiments indicate that complement has an important role at the beginning of experimental cutaneous leishmaniasis caused by L. (L.) amazonensis by controlling the number of parasites in the lesion.

Histopathologic patterns of nephropathy in naturally acquired canine visceral leishmaniasis.

Although the nephropathy of visceral leishmaniasis (VL) is known both in humans and dogs, histopathologic alterations have not been thoroughly studied. We examined renal alterations in 55 dogs with naturally acquired VL compared with five noninfected dogs from an endemic area in northeastern Brazil. Glomerulonephritis was found in 55 dogs, interstitial alterations in 53 dogs, and tubular changes in 43 dogs with VL. The glomerular alterations found were minor glomerular abnormalities (n = 8, 14.5%), focal segmental glomerulosclerosis (n = 10, 18.2%), mesangial proliferative glomerulonephritis (n = 17, 32.7%), membranoproliferative glomerulonephritis, (n = 18, 30.9%), crescentic glomerulonephritis (n = 1, 1.8%), and chronic glomerulonephritis (n = 1, 1.8%). Morphometric and ultrastructural studies complemented the analysis. The five control animals exhibited no glomerular alterations. The glomerular lesions were related to functional alterations. Considering that the alterations of canine and human nephropathy in VL are very similar, the data obtained in this study constitute an important contribution to the understanding of canine and human VL nephropathy.

Pathology of toxoplasmosis in captive new world primates.

Clinical information was available for 32 of 33 New World primates with fatal toxoplasmosis, all of which were subjected to a variable number of pathological observations. Death without apparent clinical signs occurred in 43.7% of cases. The most common clinical findings were malaise (40.6%), dyspnoea (18.7%), hypothermia (15.6%) and a sero-sanguinous or foamy nasal discharge (12.5%). Nutritional status was good in 71.8%, average in 18.7% and poor in 9.4%. The most common post-mortem findings were pulmonary congestion (78.8%), pulmonary oedema (75.8%), splenomegaly (57.6%) and mesenteric lymphadenitis (54.6%). The most common histopathological findings were multifocal necrotic hepatitis (97%), lymphadenitis (95.4%), interstitial pneumonia (90.3%) and necrotic splenitis (71.4%). The gross post-mortem changes in cebids were more variable than those observed in callitrichids, a fact that may complicate the diagnosis of toxoplasmosis in cebids.

Glomus tumour in the digit of a dog.

The gross and microscopical features of a glomus tumour in the digit of a 9-year-old dog are described. The tumour consisted of a red nodule near the nail of the third digit of the right forelimb and appeared painful. The tumour cells, which had round to oval hyperchromatic nuclei and scant cytoplasm, were arranged in sheets around blood vessels, or in nests or duct-like structures. This pattern has not been described previously in canine glomus tumours. Mitotic figures were seen only occasionally. Tumour cells were strongly immunolabelled for vimentin and some expressed smooth-muscle actin and desmin. They were negative for cytokeratins, neuron-specific enolase and CD34. Silver impregnation (reticulin method) stained the reticulum around blood vessels, nests of tumour cells and duct-like structures, and a delicate reticulum was seen around each tumour cell. The morphological, immunohistochemical and histochemical patterns helped in the diagnosis of this glomus tumour.

Susceptibility and features of the ultrastructure of Prototheca zopfii following exposure to copper sulphate, silver nitrate and chlorexidine.

One of the most important forms of the occurrence of protothecosis is bovine mastitis. Studies on the "in vivo" and "in vitro" susceptibility to antimicrobials have shown that the microorganism is resistant to most of them. Looking for alternative treatments this study aimed to study the susceptibility to copper sulphate (which has an important algicide effect) and silver nitrate (used in dairy cattle breeding for the cauterization of mammary glands) and also to chlorexidine (an important post-dipping anti-septic used in dairy practice), and the effect of these antimicrobials in the ultrastructure of Prototheca zopfii before and after the exposure to these drugs. The "in vitro" susceptibility tests to chlorexidine, silver nitrate and copper sulphate of the strains of Prototheca zopfii for the determination of their minimal microbicidal concentrations (MMC), were performed using the tube dilution method in Sabouraud dextrose broth and evaluation of colony growth after plating in Sabouraud dextrose agar. The MMCs of chlorexidine, copper sulphate and silver nitrate of the 50 strains tested were 0.01%, 0.1% and 0.3%, respectively. The tubes containing the material used in the antimicrobial susceptibility tests were prepared for the examination in an electron microscope. The untreated controls of P. zopfii showed a similar ultrastructural appearance with the typical characteristics of the microorganism. Cells exposed to silver nitrate showed changes suggesting thickness of the cell wall. Cells exposed to chlorexidine showed changes suggesting degradation of intra-cellular organelles present in the cytoplasm. P. zopfii treated with copper sulphate showed changes suggesting fibrilation of inner layer of cell wall.

Occurrence of cattle Sarcocystis species in raw kibbe from Arabian food establishments in the city of São Paulo, Brazil, and experimental transmission to humans.

Fifty samples of raw kibbe from 25 Arabian restaurants in the city of São Paulo, Brazil, were examined for the presence of bovine Sarcocystis species, using light and electron microscopy, and for infectivity to humans. Sarcocysts were found in all 50 samples. Based on cyst wall structure, S. hominis (94%), S. hirsuta (70%), and S. cruzi (92%) were identified (mostly as mixed infections). Different raw kibbe samples, positive for S. hominis in fresh preparations, were offered as a meal for 7 human volunteers. Six volunteers (85.7%), 2 of whom developed diarrhea, excreted sporocysts in feces. The prepatent period lasted 10-14 (12 +/- 1.8) days and the patent period lasted 5-12 (8.8 +/- 1.1) days.

Toxicity testing of Senna occidentalis seed in rabbits.

The effect was investigated of administering ground Senna occidentalis seeds to rabbits in different concentrations (1%, 2%, 3% and 4%) in the ration. The experiment lasted 30 days and the toxic effects of the plant were evaluated on the basis of weight gain, histopathological, biochemical and morphometric parameters, as well as histochemistry and electron microscopy. Animals that received the ration containing 4% ground S. occidentalis seeds gained less weight (p < 0.05) and died in the third week. Histopathology revealed that the heart and liver were the main organs affected, with myocardial necrosis and centrolobular degeneration. There was a reduction in cytochrome oxidase activity in the glycogenolytic fibres, together with muscle atrophy, confirmed by the morphometric studies. Electron microscopy of the liver cells revealed dilated mitochondria, with destruction of the internal cristae.

The effect of cyclophosphamide on brainstem remyelination following local ethidium bromide injection in Wistar rats.

Long-term cyclophosphamide (CY) treatment was used in male Wistar rats submitted to ethidium bromide (EB) demyelinating model to investigate ultrastructurally the drug effects on remyelination and on central nervous system (CNS) tissue repair. Demyelination was induced by a single 10 microl intracisternal injection of 0.1% EB solution and the rats anaesthetized and perfused through the heart from the 15th to the 31st day after injection. Brainstem sections were collected and processed for light and transmission electron microscopy studies. At different times after EB injection, it was observed the presence of macrophages in phagocytic activity and non-degraded myelin debris in the extracellular space, as well as remyelinated and demyelinated axons. Remyelination was carried out by both oligodendrocytes and Schwann cells, the latter notably around blood vessels and in areas of expanded extracellular space. It was also noted groups of infiltrating meningeal cells and astrocytes showing hypertrophic processes with numerous bundles of glial filaments. The rats treated with CY showed greater amounts of myelin-derived membranes than non-treated rats, suggesting a delay in the macrophage activity of removing myelin debris. Additionally oligodendrocyte remyelinating activity showed an incipient and restricted pattern, with clear predominance of naked axons. Rare lymphocytes were also found, as well as decreased neovascularization.

Aspectos ultraestruturais da criptosporidiose experimental em suínos / Ultrastructural aspects of experimental cryptosporidiosis in pigs

O presente estudo teve por objetivo investigar as alteraçöes ultraestruturais da mucosa intestinas e do tecido linfóide associado na inoculaçäo experimental de Cryptosporidium sp. Doze leitöes foram inoculados experimentalmente por via oral com 1x10 elevada a sexta potência oocistos e sacrificados 3, 6, 9 e 12 dias depois. Ao exame ultraestrutural de células intestinais observou-se espessamento e irregularidade de microvilosidades, citoplasma vacuolizado e com protrusöes, edema mitocondrial, hipertrofia de organelas citoplasmáticas e do núcleo. Nas placas de Peyer observou-se ocasionalmente mitose de células linfóides, verificando-se maior número de células blásticas

The role of natural killer cells in the early period of infection in murine cutaneous leishmaniasis.

In order to study the role of natural killer (NK) cells during the early period of Leishmania infection, BALB/c mice were selectively and permanently depleted of NK cells by injection with 90Sr and subsequently infected with Leishmania (Leishmania) amazonensis (HSJD-1 strain). 90Sr is known to selectively deplete NK cells, leaving an intact T- and B-cell compartment and preserving the ability to produce both interferon alpha and IL-2. This method of depletion has advantages when compared with depletion using anti-NK cell monoclonal antibodies because the effect is permanent and neither activates complement nor provokes massive cell death. In the present study, after one month of treatment with 90Sr, the depletion of NK cells was shown by a more than ten-fold reduction in the cytotoxic activity of these cells: 2 x 10(6) spleen cells from NK-depleted animals were required to reach the same specific lysis of target cells effected by 0.15 x 10(6) spleen cells from normal control animals. The histopathology of the skin lesion at 7 days after Leishmania infection showed more parasites in the NK cell-depleted group. This observation further strengthens a direct role of NK cells during the early period of Leishmania infection.

The role of natural killer cells in the early period of infection in murine cutaneous leishmaniasis

In order to study the role of natural killer (NK) cells during the early period of Leishmania infection, BALB/c mice were selectively and permanently depleted of NK cells by injection with 90Sr and subsequently infected with Leishmania (Leishmania) amazonensis (HSJD-1 strain). 90Sr is known to selectively deplete NK cells, leaving an intact T- and B-cell compartment and preserving the ability to produce both interferon alpha and IL-2. This method of depletion has advantages when compared with depletion using anti-NK cell monoclonal antibodies because the effect is permanent and neither activates complement nor provokes massive cell death. In the present study, after one month of treatment with 90Sr, the depletion of NK cells was shown by a more than ten-fold reduction in the cytotoxic activity of these cells: 2 x 106 spleen cells from NK-depleted animals were required to reach the same specific lysis of target cells effected by 0.15 x 106 spleen cells from normal control animals. The histopathology of the skin lesion at 7 days after Leishmania infection showed more parasites in the NK cell-depleted group. This observation further strengthens a direct role of NK cells during the early period of Leishmania infection

Influence of low environmental temperature on inflammation in bullfrog (Rana catesbeiana): qualitative and quantitative evaluation

Objectives of this study were to investigate the influence of low environmental temperature on the experimentally induced inflammatory response in post-metamorphic Rana catesbeiana (bullfrogs). To accomplish these goals, 120 specimens of Rana catesbeiana were kept at 6§C and 24§C, and treated by transfixion of thigh muscular tissue with a 5-0 suture or IM carrageenan injection. Results obtained through qualitative and quantitative evaluations showed that the lower environmental temperature significantly modulates the inflammatory process development. The animals in both models that were kept at 6§C showed a significantly lower number of inflammatory cells in the lesion site than the one verified at 24§C, apart from the evolution time. On the other hand, any factor related to the host mechanism of defense ought not to be blocked by the temperature, since the area of reaction to the injury showed to be equivalent in most of the studied time