Detection and molecular characterization of Mycobacterium celatum as a cause of splenitis in a domestic ferret (Mustela putorius furo)

Mycobacterium celatum is a slow growing non-tuberculous mycobacterium described mainly as occurring in human patients. Only two cases of infection with this pathogen have been reported previously in animals. A 5-year-old, neutered male ferret was presented with progressive weight loss and muscle atrophy. Pale mucous membranes, slight alopecia of the tail and splenomegaly, confirmed by abdominal ultrasound, were observed. Fine-needle aspirations of the spleen revealed extramedullary haematopoiesis and marked macrophage-dominated inflammation associated with mycobacterial infection. Ziehl-Neelsen staining demonstrated sporadic acid-fast bacilli within macrophages. These organisms were identified as M. celatum by microbiological and molecular?methods. Phylogenetic analysis based on the 16S rDNA gene compared this isolate with previously reported strains and demonstrated close relatedness to the human strains of M. celatum types 1 and 3. The ferret was treated with enrofloxacin, rifampicin and azithromycin, resulting in clinical improvement. After 40 days of treatment, the spleen was re-evaluated. Cytological evaluation revealed only extramedullary haematopoiesis without evidence of infection. Discontinuation of therapy was followed by rapid deterioration and death.     

Morphology of the pituitary gland in ferrets (Mustela putorius furo) with hyperadrenocorticism

Pituitary tumours are the cause of hyperadrenocorticism in a variety of species, but the role of the pituitary gland in hyperadrenocorticism in ferrets is not known. In this species, the disease is mediated by the action of excess gonadotrophins on the adrenal cortex and is characterized by an excessive secretion of sex steroids. In this study, the pituitary gland of four healthy control ferrets, intact or neutered, and 10 neutered ferrets with hyperadrenocorticism was examined histologically following immunohistochemical labelling for adrenocorticotrophic hormone, alpha-melanocyte-stimulating hormone, growth hormone, thyroid-stimulating hormone, luteinizing hormone, follicle-stimulating hormone, and prolactin. Immunohistochemistry revealed that somatotrophs, thyrotrophs and lactotrophs were the most abundant cell types of the pars distalis of the pituitary gland in the healthy ferrets. The distribution of corticotrophs was similar to that in the dog and man. In ferrets, as in dogs, the melanotrophic cell was almost the only cell type of the pars intermedia. Gonadotrophs were found in the pars distalis of neutered, but not intact ferrets. All the ferrets with hyperadrenocorticism had unilateral or bilateral alterations of the adrenal gland. In addition, in the pituitary gland of two of these ferrets a tumour was detected. These tumours were not immunolabelled by antibodies against any of the pituitary hormones, and had characteristics of the clinically non-functional gonadotroph tumours seen in man. In some of the other ferrets low pituitary immunoreactivity for gonadotrophic hormones was detected, which may have been due to the feedback of autonomous steroid secretion by the neoplastic transformation of the adrenal cortex. It is concluded that initially high concentrations of gonadotrophins resulting from castration may initiate hyperactivity of the adrenal cortex. The low incidence of pituitary tumours and the low density of gonadotrophin-positive cells in non-affected pituitary tissue in this study suggest that persistent hyperadrenocorticism is not dependent on persistent gonadotrophic stimulation.     

Contribution of auditory cortex to sound localization by the ferret (Mustela putorius)

Ferrets were tested in a semicircular apparatus to determine the effects of auditory cortical lesions on their ability to localize sounds in space. They were trained to initiate trials while facing forward in the apparatus, and sounds were presented from one of two loudspeakers located in the horizontal plane. Minimum audible angles were obtained for three different positions, viz., the left hemifield, with loudspeakers centered around -60 degrees azimuth; the right hemifield, with loudspeakers centered around +60 degrees azimuth; and the midline with loudspeakers centered around 0 degrees azimuth. Animals with large bilateral lesions had severe impairments in localizing a single click in the midline test. Following complete destruction of the auditory cortex performance was only marginally above the level expected by chance even at large angles of speaker separation. Severe impairments were also found in localization of single clicks in both left and right lateral fields. In contrast, bilateral lesions restricted to the primary auditory cortex resulted in minimal impairments in midline localization. The same lesions, however, produced severe impairments in localization of single clicks in both left and right lateral fields. Large unilateral lesions that destroyed auditory cortex in one hemisphere resulted in an inability to localize single clicks in the contralateral hemifield. In contrast, no impairments were found in the midline test or in the ipsilateral hemifield. Unilateral lesions of the primary auditory cortex resulted in severe contralateral field deficits equivalent to those seen following complete unilateral destruction of auditory cortex. No deficits were seen in either the midline or the ipsilateral tests.     

Retinal projections to the accessory optic system in pigmented and albino ferrets (Mustela putorius furo)

We investigated if a reduced specificity of the retinal projection to the accessory optic system might be responsible for the loss of direction selectivity in the nucleus of the optic tract and dorsal terminal nucleus (NOT-DTN) and, in consequence of this, the optokinetic deficits in albino ferrets. Under electrophysiological control we performed dual tracer injections into the NOT-DTN and the medial terminal nucleus (MTN). Retrogradely labelled ganglion cells were found in the visual streak, the dorsal, and the ventral retina both after injections into the NOT-DTN and the MTN indicating that both nuclei receive input from the same retinal regions. The distribution and spacing of labelled ganglion cells did not differ between pigmented and albino ferrets. However, retinal ganglion cells projecting simultaneously to both the NOT-DTN and the MTN occurred only in albino ferrets. These results suggest that a reduced specificity of the projection pattern of direction specific ganglion cells may contribute to the loss of direction selectivity in the NOT-DTN in albino ferrets.     

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.     

Identification of group 1 coronavirus antigen in multisystemic granulomatous lesions in ferrets (Mustela putorius furo)

Tissues from nine ferrets with granulomatous lesions similar to those seen in feline infectious peritonitis were examined histopathologically and immunohistochemically. Four main types of lesions were observed: diffuse granulomatous inflammation on serosal surfaces; granulomas with areas of necrosis; granulomas without necrosis; and granulomas with neutrophils. Other less commonly seen lesions were granulomatous necrotizing vasculitis and endogenous lipid pneumonia. FCV3-70 monoclonal antibody produced immunolabelling of group 1 coronavirus antigen in tissue samples from eight animals, the antigen being present in the cytoplasm of macrophages in the different types of granulomatous lesions.     

Reproduction in the ferret (Mustela furo) I. Uterine histology and histochemistry during pregnancy and pseudopregnancy

Similar histological and histochemical changes occur in the uteri of pregnant and pseudopregnant ferrets. The pre-implantation period is characterized by general growth of the tissue and moderate secretory activity. At the time of implantation, the luminal epithelium undergoes hypertrophy and hyperplasia and shows much secretory activity. This area of altered activity sweeps progressively toward the gland fundi and the epithelium behind it becomes greatly enlarged and forms symplasmic masses. Symplasma formation occurs in all regions of the uterus, in contrast to other carnivores where it occurs only next to trophoblast and in response to uterine trauma. The principal uterine secretions are mucoproteins and glycoproteins. There is some evidence that pseudopregnant uteri, but not pregnant uteri, secrete acid mucopolysaccharides. Glycogen is present in moderate amounts at estrus, declines during the pre-implantation period, and is absent during gestation. A light to moderate diffuse sudanophilia is seen during pregnancy and pseudopregnancy, becoming more pronounced toward the end of the gestation period. Occasional cholesterol crystals are seen in macrophages in the stroma of pregnant, but not pseudopregnant, uteri. Macrophages containing hemosiderin were seen in all uteri examined, being most abundant in estrous and early pre-implantation uteri. Uterine mast cells were abundant at estrus, declined during the pre-implantation period and decreased markedly at the time of implantation. The results are compared with data from other carnivore species.     

Midline and lateral field sound localization in the ferret (Mustela putorius): contribution of the superior olivary complex.

1. The ability of ferrets to localize sound in space was determined before and after unilateral or bilateral lesions of the superior olivary complex (SOC). Lesions were made by pressure injection of kainic acid into the SOC through a stereotaxically positioned glass micropipette. The lesions destroyed the cell bodies in the superior olive without disrupting fibers of passage in the trapezoid body or other pathways in the auditory brain stem. The integrity of fibers was demonstrated by protargol staining of axonal processes and by the retrograde transport of horseradish peroxidase (HRP) from the inferior colliculus to other auditory brain stem nuclei. Behavioral tests were carried out separately for sound localization at midline and lateral field positions. Minimum audible angles were determined for single 45-ms noise bursts presented through paired loudspeakers positioned symmetrically around 0, -60, and +60 degrees azimuth. 2. Four ferrets received complete lesions of the left SOC, and two received complete lesions of the right SOC. In general, unilateral destruction of the superior olive resulted in impairments in sound localization in both left and right lateral fields. In some cases, deficits were also apparent on midline. Four additional animals received unilateral lesions that spared cells within the SOC. In most cases, deficits were apparent despite incomplete lesions of the SOC. The pattern of deficits was generally consistent with that found in animals with complete lesions. Most animals had difficulty localizing sounds in the lateral fields. 3. Four animals received bilateral lesions of the SOC. Three had complete or near-complete destruction of the superior olive on one side of the brain with relatively minor damage on the other side. Each of these animals exhibited behavioral deficits that were particularly severe ipsilateral to the more extensively damaged superior olive. One animal with complete bilateral destruction of the SOC was incapable of sound localization, even with 2-s noise bursts. This animal, however, suffered severe motor impairments after surgery that might have contributed to the apparent inability to localize sound. 4. Two animals with kainic acid lesions that caused little or no damage to the SOC were still capable of high levels of performance in tests of sound localization and had no elevation in minimum audible angles. These cases served as controls for the possible effects of nonspecific brain damage and demonstrated that kainic acid injections per se resulted in no obvious deficits in our test situation.(ABSTRACT TRUNCATED AT 400 WORDS)     

Variation in basal arteries of the brain in polecat (Mustela putorius putorius L.).

Observations carried out on 106 polecat brains showed that the system of basilar arteries of the brain is similar to those in other cornivorous species. The observed variation was manifested in a differentiation connections of the vessels, in occurence of some variations and in asymmetry. Some cases of such variations were hitherto observed only in some ruminants.     

Hearing in the ferret (Mustela putorius): effects of primary auditory cortical lesions on thresholds for pure tone detection

1. Pure tone thresholds were determined for five adult male ferrets before and after bilateral ablation of primary auditory cortex. Complete audiograms ranging from 0.016 to 48 kHz were obtained for two animals. The remaining three animals were tested at five frequencies selected to assess hearing throughout the audible range (0.125, 0.5, 2.0, 8.0, and 32.0 kHz). 2. Shortly after surgery one animal had elevated thresholds across the entire frequency range with the most pronounced hearing loss above 12.0 kHz. Four other animals had no elevation of thresholds at low and midrange frequencies but suffered a hearing loss at very high frequencies (32 kHz). 3. Repeated testing over a period of several months revealed substantial recovery of sensitivity. There was complete recovery of sensitivity in the low- and middle-frequency range of the audiogram. Some hearing loss persisted at the extreme upper end of the audiogram (32 kHz), but in two cases there was evidence of recovery at this frequency as well. 4. Following determination of absolute thresholds all animals were assessed for their ability to localize sound in space. Minimum audible angles were obtained on midline as well as within both left and right hemifields, i.e., around 0, -60 and +60 degrees azimuth. All animals had severe and persistent deficits in their ability to localize brief sounds within the lateral fields, but were still capable of midline localization.     

Helicobacter pylori infection and chronic gastritis in gastric cancer.

AIMS: To investigate the prevalence of Helicobacter pylori associated chronic gastritis in patients with gastric cancer. METHODS: Serum IgG antibodies for H pylori were determined in 54 consecutive patients with gastric carcinoma. The prevalence of H pylori in gastric mucosa was also examined histologically (modified Giemsa) in 32 patients from whom adequate biopsy specimens of the antrum and corpus were available. Thirty five patients with gastrointestinal tumours outside the stomach and 48 with non-gastrointestinal malignancies served as controls. RESULTS: Of the 54 patients, 38 (70%) had H pylori antibodies (IgG) in their serum (three additional patients had H pylori antibodies IgA, class specific but not IgG specific). This prevalence was significantly higher (p less than 0.05) than that (49%) in the 35 controls. No differences in prevalence of H pylori antibodies were found between gastric cancer cases of intestinal (IGCA) or diffuse (DGCA) type, both these types showing H pylori antibodies (IgG) in 71% of the patients. In the subgroup of 32 subjects, five patients had normal gastric mucosa and four showed corpus limited atrophy ("pernicious anaemia type" atrophy of type A). All of these nine patients had no evidence of current or previous H pylori infection in serum (no IgG antibodies) or in tissue sections (negative Giemsa staining). The remaining 23 patients had antral or pangastritis, and all had evidence of current or previous H pylori infection. CONCLUSIONS: H pylori associated chronic gastritis was the associated disease in 75% of the patients with gastric cancer occurring equally often in both IGCA and DGCA groups. About 25% of cases seem to have a normal stomach or severe corpus limited atrophy, neither of which showed evidence of concomitant H pylori infection.     

Helicobacter pylori gastritis and primary gastric non-Hodgkin's lymphomas.

AIMS--To evaluate further the relation between gastric malignant lymphoma of the mucosa associated lymphoid tissue (MALT) and Helicobacter pylori. METHODS--One hundred and sixty two surgical specimens of MALT lymphoma were retrospectively investigated to determine tumour type and inflammatory patterns. In 121 cases biopsy specimens obtained before surgery were available and stained with haematoxylin and eosin, periodic acid Schiff, Giemsa and Warthin-Starry stains. RESULTS--Residual lymphoid follicles were found less often in high grade malignant than in low grade malignant MALT lymphomas. Chronic active gastritis was shown within the mucosa at some distance from the tumours in 143 of 146 specimens. In all the cases for which biopsy specimens could be evaluated, colonisation of the mucosa by H pylori had occurred. Lymphoid follicles and lymphoid aggregates were detected in 82.7% of the antral, and in 85% of the body mucosa specimens. CONCLUSIONS--These data support the hypothesis that H pylori has an important role in the development of MALT lymphomas. Furthermore, the chronic inflammation preceding malignant transformation might enhance the probability of malignant transformation via chronic stimulation of the lymphoid tissue. This might in part indicate why MALT lymphomas occur most often in the stomach.     

Up-regulation of cathepsin X in Helicobacter pylori gastritis and gastric cancer

Recently, we identified increased cathepsin X expression in H. pylori-infected gastric mucosa. Here, we describe further up-regulation in gastric cancer and report on the role of inflammatory cytokines required for cathepsin X up-regulation in H. pylori-infected gastric mucosa, as well as on consequences for cellular invasion. Biopsy specimens were taken from the antrum, corpus and cardia of H. pylori-infected and non-infected patients. Gastric cancer samples were obtained from patients undergoing gastric surgery. Cathepsin X was detected in gastric mucosa by quantitative real-time RT-PCR, western blotting and immunohistochemistry. Induction of cathepsin X expression in epithelial and inflammatory cells caused by H. pylori infection was tested in in vitro contact and non-contact co-cultures of AGS cells and monocytic cells. Patients with H. pylori gastritis showed significantly higher cathepsin X mRNA (2.5-fold) and protein (1.6-fold) expression than H. pylori-negative patients. Cathepsin X was also up-regulated in gastric cancer (3-12-fold) compared to non-neoplastic mucosa. Cathepsin X was predominantly expressed by macrophages in the mucosal stroma and in glands of the antral mucosa. In addition, tumour cells stained for cathepsin X in 26 (68%) patients with gastric carcinoma. In general, staining was significantly more common (20 vs. 6 patients) and more intense (3.55 vs. 0.83) in intestinal type gastric cancer than in the diffuse type. In vitro cell culture experiments revealed that intercellular signalling between pathogenicity island (PAI)-positive H. pylori-infected epithelial cells and macrophages via soluble factors in the culture medium seems to be responsible for increased expression of cathepsin X in monocytes. Using antisense oligonucleotides, cathepsin X up-regulation was directly associated with higher invasiveness in vitro. Although no correlation of cathepsin X expression and TNM stage was found, our study demonstrates that cathepsin X plays a role not only in the chronic inflammation of gastric mucosa but also in the tumourigenesis of gastric cancer.