Neuronal ceroid lipofuscinosis in a Vietnamese pot-bellied pig (Sus scrofa).

The neuronal ceroid ipofuscinoses (NCL) are a group of heritable, neurodegenerative, storage diseases, typically with an autosomal recessive mode of inheritance. Cytoplasmic accumulation of storage material in cells of the nervous system and, variably in other tissues, characterizes NCL. NCL has been reported in many animal species, but to the authors' knowledge, this is the first report of the disease in a pig. Blindness and seizures are common clinical signs of disease, neither of which was a feature in this pig. The lesions were restricted to the central nervous system, which was diffusely affected, with the most severe lesions in the hippocampus, cerebral cortex, and cerebellum. The histologic lesions included neuronal loss and gliosis, which contributed to mild cerebrocortical and cerebellar atrophy and accumulation of autofluorescent storage material in neurons and glial cells. The storage material had morphologic, histologic, and ultrastructural properties typical of NCL.

Immunocytochemical localization of adrenocorticotropic hormone-immunoreactive cells of the pars intermedia in thoroughbreds.

OBJECTIVE: To analyze and characterize adenoma and hyperplasia of the pars intermedia (PI) of Thoroughbred mares. ANIMALS: 165 Thoroughbred mares, without clinical signs of hyperadrenocorticism that had been euthanatized or had died, of causes such as sudden death, colic, pneumonia, or trauma, and were necropsied. Five of those horses, 17 to 25 years old, had a large pituitary gland at necropsy. Eight mares, 5 to 15 years old with normal-size pituitary gland, were selected at random for comparison. PROCEDURE: A morphologic comparison of the pituitary gland between horses with and without tumors of the PI was conducted by use of immunocytochemistry and morphometry. RESULTS: In horses with normal pituitary gland, the PI was supplied by a vast capillary or sinusoidal plexus, which connected that in the pars distalis (PD) with that in the pars nervosa (PN). Cells of the PI stained slightly with ACTH antiserum, but some cells in the border region, which is contiguous to the PD, were strongly ACTH immunoreactive. At necrospy, horses with an enlarged pituitary gland also had adenoma of the pituitary gland involving the PI. Cells of the border region were hypertrophied and stained strongly with ACTH antiserum. The area and number of individual tumor cells of the border region of the PI of horses with adenoma were significantly increased, compared with those in horses with normal pituitary gland. CONCLUSIONS: Cells of the PI-PD border region may secrete substantial quantities of ACTH, owing to stimulation by corticotropin-releasing factor. Adenoma and hyperplasia of the PI in Thoroughbred mares may be associated with hyperadrenocorticism.

Degenerative leukoencephalopathy and myelopathy in dasyurids.

From June 1985 to November 1992 eight eastern quolls (Dasyurus viverrinus) and six Tasmanian devils (Sarcophilus harrisii), held at institutions in Australia and Canada, developed progressive blindness and posterior paresis progressing to paralysis. All were aged individuals. At necropsy white matter tracts, particularly in the spinal cord, optic tracts, and cerebellum, contained evidence of severe axon degeneration with vacuolation and macrophage infiltration. The surrounding tissue had evidence of gliosis. The etiology of the condition is unknown but may be a form of age-related degeneration.

An immunodominant epitope on 40 kilodalton outer membrane protein is conserved among different strains of Haemophilus (Histophilus) somnus.

Four murine monoclonal antibodies (mAb) were constructed against Haemophilus (Histophilus) somnus, an important bovine pathogen, and used to analyze immunologically significant antigenic determinants on these organisms. These mAbs specifically recognized immunodominant epitopes present on the 40 kilodalton (kD) fraction of the major outer membrane protein (OMP) of H. somnus. The 9D3 IgG1 kappa mAb recognized an immunodominant epitope on the 40 kD major outer membrane protein that is conserved or shared among all the three strains (septicemic, respiratory and uro-genital) of H. somnus. Two IgM kappa mAbs (4D6 and 10C2) recognized epitope(s) on the 40 kD OMP from septicemic and respiratory strains of H. somnus but none from the whole bacterial cell preparations. Another IgM kappa mAb 9D2 recognized an antigenic determinant on the 40 kD protein from the OMP as well as a whole bacterial cell preparation of a septicemic strain of H. somnus. These data demonstrate that, at least, three immunologically significant antigenic determinants on H. somnus are defined by mAbs against this bovine pathogen. Importantly, these studies suggest that the epitope present on the 40 kD major OMP, recognized by the 9D3 mAb, is immunodominant and conserved among septicemic, respiratory and urogenital strains, and is, therefore, suitable for further investigating its use in the development of an immunodiagnostic assay and also as a recombinant vaccine.

Adherence of Haemophilus somnus to tumor necrosis factor-alpha-stimulated bovine endothelial cells in culture.

Vascular thrombosis and tissue infarction is a principal lesion in Haemophilus somnus septicemia known also as thrombotic meningoencephalitis. This study was undertaken to examine whether tumor necrosis factor-alpha (TNF-alpha) can influence the adherence of H. somnus to cultured bovine aortic endothelial cells (BAEC). Confluent BAEC were exposed to 0-100 nM of human recombinant TNF-alpha for 12-48 h. Suspensions of different strains of H. somnus (approximately 1.5-3 x 10(8) labelled with [methyl-3H]-thymidine, were added to BAEC and incubated for 1.5 h. Initial studies with one pathogenic (P) strain and one non-pathogenic (NP) strain revealed that both strains adhered to normal endothelial cells but minimally to subendothelial matrix remaining after removal of BAEC. Adherence to BAEC was reduced by an excess of unlabelled H. somnus of the same strain. Adherence was enhanced for both strains by exposure of BAEC to TNF-alpha in a manner that increased with TNF-alpha concentration and with duration of exposure to TNF-alpha prior to addition of bacteria. A survey of adherence of six live P strains and six NP strains demonstrated considerable variation but no difference in adherence between P and NP strains to normal or to TNF-alpha-stimulated BAEC. However, TNF-alpha consistently increased adhesion of each strain to BAEC. Both P and NP strains caused more severe cytotoxic changes in TNF-alpha-treated BAEC. Tumor necrosis factor-alpha also increased adhesion of formalin-killed bacteria of P and NP strains.(ABSTRACT TRUNCATED AT 250 WORDS)

Differences in protein expression of Haemophilus somnus grown under conditions of iron-restriction.

Outer membrane protein profiles were compared in 14 H. somnus strains isolated from brain and lung lesions as well as from the genital tract of asymptomatic carriers during in vitro growth under iron-restricted conditions. Ethylenediamine-di-O-hydroxyphenyl acetic acid (EDDA) was used to obtain iron-restricted conditions in media used for this study. The outer membrane protein profiles were studied by the discontinuous sodium dodecyl sulfate-polyacrylamide gel electrophoretic system (SDS-PAGE), and the proteins were stained with silver or transferred to nitrocellulose sheets and western blots conducted. Growth under iron-restricted conditions resulted in the induction of outer membrane proteins in most H. somnus strains examined. Studies also indicated differences among H. somnus strains in the number of induced proteins and their molecular weights but the results did not indicate a specific relationship between these strain-dependent differences and tissue trophism. Western blot analysis revealed a high degree of immunological relatedness among strains of H. somnus in their iron-regulated proteins. However, hyperimmune serum used in these assays failed to recognize certain iron-regulated proteins expressed by some H. somnus strains, a finding which may have important implications for the induction of protective immunity in cattle against this bovine pathogen.

Genomic fingerprinting of "Haemophilus somnus" isolates by using a random-amplified polymorphic DNA assay.

The random-amplified polymorphic DNA (RAPD) assay was used to generate DNA fingerprints for 16 isolates of "Haemophilus somnus," and one isolate each of "Haemophilus agni," "Histophilus ovis," "Actinobacillus seminis," Pasteurella haemolytica, and Escherichia coli. The RAPD assay differentiated among "H. somnus" isolates, which shared similarity coefficients of 0.46 to 1.00 on the basis of pairwise comparisons of RAPD markers produced with nine random decamer primers. Three virulent encephalitic "H. somnus" isolates exhibited identical banding patterns, suggesting a common clonal ancestry. The RAPD assay clearly distinguished between the "H. somnus"-"H. agni"-"H. ovis" group and the other bacterial species tested. The results of the present study suggest that DNA fingerprinting of "H. somnus" isolates by the RAPD assay could be valuable in revealing subspecific divisions within this largely unexplored species.

Comparison of magnetic resonance images and the histopathological findings of lesions induced by interstitial laser photocoagulation in the brain.

Interest has developed in using magnetic resonance (MR) imaging to monitor the volume of tissue destroyed by interstitial laser photocoagulation (ILP). In these experiments, ILP was induced in the normal brains of 9 anesthetized cats by delivering 1.5 W of continuous-wave Nd:YAG laser energy (1,064 nm) from a single 400-microns core optical fiber for 1,000 s. The irradiations were monitored using proton spin-echo MR imaging during and immediately after ILP and at postirradiation survival times of 2, 5, and 14 days. At 2 days postirradiation, the necrotic thermal lesion consisted of a central cavity surrounded by 2 concentric zones of coagulative necrosis, one dense and the other dispersed. The lesion shrank and the zonal appearance became less obvious over the 14 day survival period. An enhancing halo on contrast-enhanced T1-weighted images acquired immediately postirradiation best approximated the total lesion diameter at 2 days. These images also indicated that the volume of tissue destroyed during ILP corresponded better to the necrotic volume determined at 2 days than at 5 days and 14 days postirradiation. T2-weighted images acquired during and immediately after ILP consistently underestimated the total lesion diameter at 2 days.

Neuropathologic findings in young male rats in a subchronic oral toxicity study using triethyl lead.

This study was undertaken to ascertain the neuropathologic effects of low level exposure of triethyl lead (3EL) to young male rats. Groups of 20 male Sprague-Dawley weanling rats were given 3EL at 0, 0.05, 0.10, 0.20, 0.50, and 1.00 mg/kg body wt for 91 days, 5 days/week by oral gavage. Lead acetate (PbHOAC) was given at 200 mg/kg body wt/day as a positive control. Animals (five or six) were perfused with glutaraldehyde following barbiturate anesthesia at the termination of the experiment. These animals and the remaining members of the group received a thorough gross and microscopic postmortem examination. Sections of the central, peripheral, and autonomic nervous systems were examined and lesions scored. No lesions were noted in the brain, but randomly distributed light microscopic changes of spinal cord Wallerian degeneration were noted to increase in a dose responsive manner (rho = 0.48; p < 0.01), with 3EL administration. Ultrastructural examination of selected sections of the lumbosacral nerves, revealed lesions characterized by reduced neurofilaments and neurotubules, and irregular lamellated axoplasmic dense bodies in all animals receiving lead. Organolead was only detected in animals receiving 3EL, but lead cations were detected in all lead-treated animals. The brain lead levels of 1.00 mg/kg/day and 200 mg Pb acetate positive control animals were equivalent. As distinctive ultrastructural lesions were seen in all rats treated with 3EL, we suggest that the no observed adverse effect level (NOAEL) for 3EL be lowered to less than 0.05 mg/kg/day.(ABSTRACT TRUNCATED AT 250 WORDS)

Isolation of pathogenic strains of Haemophilus somnus from the female bovine reproductive tract.

The prevalence of Haemophilus somnus in the genital tract of slaughtered and live cows in southern Ontario was investigated. The vagina and uterus of slaughtered cows were swabbed separately. Live cows were examined and sampled in two field surveys: Centre A and Centre B. In the former, aspirated mucus secretions and in the latter, specimens obtained by guarded swabbing were examined bacteriologically. Haemophilus somnus was isolated from 28 genital tracts of 461 slaughtered (6.1%), and seven of 199 live (3.5%) cows during the centre B survey. The isolates were recovered from both normal and diseased reproductive tracts. Fourteen strains isolated from genital organs were examined for pathogenicity in vivo to test the occurrence of pathogenic isolates. In the initial stage of the in vivo study on pathogenicity, each of the fourteen isolates was examined on one calf using an intracisternal inoculation. Subsequently, one pathogenic and one nonpathogenic strain were inoculated into five calves each to statistically confirm their pathogenic potential. Of 14 genital isolates of H. somnus examined in an intracisternal calf assay, six (43%) caused a fatal peracute neurological disease, while eight were nonpathogenic. A comparative pathological study of pathogenic and nonpathogenic isolates showed that the former caused a severe fatal suppurative meningoencephalitis whereas the latter caused no lesions whatsoever or a mild leukocytic leptomeningitis. The salient data obtained in this study indicate that there are pathogenic strains of H. somnus in the genital tract of apparently normal cows as well as of those with inflammatory disease.

Magnetic resonance imaging of interstitial laser photocoagulation in brain.

Magnetic resonance (MR) imaging can be used to monitor the development of thermal lesions induced in tissue using interstitial laser photocoagulation (ILP). A potential application for ILP is the treatment of surgically inaccessible brain tumors. For the successful clinical application of MR-monitored ILP, it is necessary to relate MR images of ILP lesions to the actual induced lesions. In this preliminary study we performed ILP in the normal brains of anesthetized cats by delivering interstitially 1.0, 1.5, and 2.0 W of continuous-wave Nd:YAG laser energy (1,064 nm) for 1,000 s via a plane-cut 400 microns core optical fiber. At 48 h post-irradiation the lesions consisted of four sharply demarcated concentric zones of thermal damage. Lesion diameter increased linearly with delivered power. T2-weighted proton spin-echo images acquired during ILP showed a region of complete or near signal loss that underestimated the actual lesion at 48 h. Gadolinium-enhanced T1-weighted spin-echo images acquired immediately post-irradiation showed the actual lesion precisely.

Haemophilus somnus and reproductive disease in the cow: A review.

Haemophilus somnus causes inflammatory disease in the genital tract of cows as reported in several field surveys and experimental trials. This organism can also innocuously colonize the healthy genital mucosa of the cow, which indicates its dual relationship with the host, that of pathogen and commensal. Experimental data indicate embryocidal capability of this pathogen suggesting a possible role in early embryonic death. Haemophilus somnus also causes sporadic abortions after a bacteremia in the dam. Retrograde infection of the pregnant uterus from the lower genital tract appears unlikely; however, this process can account for post-parturient endometritis. Detection of high homologous IgG(2) serum antibody titers using an ELISA test may be useful for the diagnosis of current or recent genital inflammation. Experimental laboratory data indicate that a proportion of genital strains of H. somnus are pathogenic and capable of causing thrombotic meningoencephalitis and perhaps pneumonia. In vivo testing of the pathogenicity of genital strains remains to be conducted.

The development of protein A-gold electron microscopy for immunological studies of Haemophilus somnus.

Two separate experiments were conducted using modifications of protein A-gold immunoelectron microscopy (PAGIEM) to evaluate the ability of sera from calves vaccinated against Haemophilus somnus to bind virulent organisms (experiment I) and to detect differences in the antibody accessible antigenic sites on the outer membrane of selected strains of H. somnus having different virulence attributes using a high IgG2 titre specific bovine hyperimmune serum (experiment II). The results of experiment I demonstrated that the direct opsonisation of H. somnus by specific antisera was related to its IgG2 titre. In experiment II, strain-dependent differences in the labelling of antigenic sites by specific IgG2 antibodies were observed. The virulent strains of both septicaemic and genital isolates of H. somnus showed higher protein A-gold labelling than their non-virulent counterparts. The results from a comparison of pathogenic and non-pathogenic respiratory isolates did not reveal the same difference in labelling intensity. The studies demonstrated the PAGIEM technique to be a sensitive, versatile and a reliable laboratory method to analyse antigen-antibody interactions of H. somnus.

Endothelial cytotoxicity of Actinobacillus pleuropneumoniae.

The cytotoxicity of Actinobacillus pleuropneumoniae serotype 1 strain CM5 for porcine and bovine endothelial cells in vitro, was dose-dependent. This strain and its attenuated and avirulent substrain CM5A were equally cytotoxic. The cytotoxicity observed during five hours of exposure of endothelial cells to bacterial products was abolished if the bacteria were inactivated by heat or sonication. Exposure of the endothelial cells for five hours to 100 and 200 micrograms of purified lipopolysaccharide resulted in a partial cytotoxicity only, which was not enhanced in the presence of fresh guinea pig serum. The cytotoxicity of viable bacteria could be neutralised by a polyclonal rabbit antiserum to the purified 104kD haemolysin. A bacteria-free supernate of a culture of strain CM5 had both haemolytic and cytotoxic activity. The haemolytic activity could be neutralised completely by the anti-serum to the 104kD haemolysin, whereas the cytotoxic activity was only partially neutralisable. Hence A pleuropneumoniae is cytotoxic for endothelial cells and this cytotoxicity is possibly mediated by the 104kD haemolysin.

Four bovine meningeal tumors.

Four bovine meningeal tumors from a total of 14 brain tumors were recorded in the files of the Ontario Veterinary College and the Ontario Veterinary Laboratory Services Branch from the years 1966 to 1984. The affected cattle ranged in age from 10 months to adult. They exhibited various nervous signs and, in each case, the animals became recumbent. Tumors were located in various areas of the central nervous system. Neoplastic cells had oval vesicular nuclei containing numerous cytoplasmic invaginations. Virus particles were not observed. A fibroblastic appearance, with the presence of loose whorls or bundles, was present in all. Three of the tumors were invasive and were classified as a fibroblastic meningioma.

Subchronic oral toxicity of triethyl lead in the male weanling rat. Clinical, biochemical, hematological, and histopathological effects.

This study was designed to ascertain the effects of low level exposure of triethyl lead (3EL) to the male weanling rat. Groups of 20 animals were administered by gavage 3EL at 0.05, 0.10, 0.20, 0.50, and 1.00 mg/kg body wt for 91 days, 5 days/week. Lead acetate (PbHOAC) at 200 mg/kg body wt/day was given as a positive control. Weight gain was reduced in those animals receiving 1.0 3EL. Spleen and kidney weights were elevated in the PbHOAC group. Residues of 3EL and its metabolites diethyl lead (2EL) and lead (Pb) accumulated in a dose-dependent manner in blood, liver, kidney, and brain; 3EL accumulated preferentially in the liver while inorganic lead accumulated in the kidney. Dose-dependent changes occurred in serum calcium which was decreased and in phosphorus which was elevated for all dose groups. Serum cholesterol was elevated in the three highest 3EL groups as was alkaline phosphatase. LDH was lowered in the PbHOAC-treated group but microsomal aniline hydroxylase was elevated. Hematological changes consisted of elevated platelet counts in the 1.0 3EL group and decreased mean corpuscular hemoglobin content and mean corpuscular volume in the PbHOAC-treated group. Treatment related histopathological changes were seen in thyroid, liver, kidney, and bone marrow. Based on these data a no observed adverse effect level for 3EL was set at 0.10 mg/kg/body wt.

The protective effect of vaccination against experimental pneumonia in cattle with Haemophilus somnus outer membrane antigens and interference by lipopolysaccharide.

A semi-purified outer membrane anionic antigen (AA) fraction was isolated from Haemophilus somnus by a modified procedure of anion exchange chromatography to yield a protein fraction free of lipopolysaccharides (LPS). The AA fraction (1 mg) was administered with or without the homologous lipopolysaccharide (10 micrograms/kg body weight) as vaccines to groups of cattle twice, three weeks apart. A control group which did not receive any antigen was included in the trial. Six weeks after the first vaccination, the animals were challenged intratracheally with a virulent pneumonic strain of H. somnus (70986) and observed for clinical signs of respiratory disease. The cattle were euthanized six days later and the lungs were evaluated for the severity of lesions macroscopically as well as histopathologically. Vaccination with AA alone provided the best protection against pneumonia as indicated by significantly lower clinical scores, less extensive gross lung lesions and mild histopathological lesions with immune cell infiltration. However, when AA was combined with LPS in the vaccination, this protective effect was negated and the animals showed more detrimental histopathological lesions than the controls.