Angelika Url

Emergence of Usutu virus, an African Mosquito-Borne Flavivirus of the Japanese Encephalitis Virus Group, Central Europe

During late summer 2001 in Austria, a series of deaths in several species of birds occurred, similar to the beginning of the West Nile virus (WNV) epidemic in the United States. We necropsied the dead birds and examined them by various methods; pathologic and immunohistologic investigations suggested a WNV infection. Subsequently, the virus was isolated, identified, partially sequenced, and subjected to phylogenetic analysis. The isolates exhibited 97% identity to Usutu virus (USUV), a mosquito-borne Flavivirus of the Japanese encephalitis virus group; USUV has never previously been observed outside Africa nor associated with fatal disease in animals or humans. If established in central Europe, this virus may have considerable effects on avian populations; whether USUV has the potential to cause severe human disease is unknown.

Evidence of parvovirus replication in cerebral neurons of cats.

ABSTRACT The correlation between parvovirus infections and lesions in the central nervous system other than cerebellar hypoplasia was studied in 100 cats. The animals were necropsied with a history of various diseases, one third showing typical clinical and pathomorphological signs of panleukopenia. In 18 cats polyclonal antiserum against canine parvovirus consistently labeled neurons mainly in diencephalic regions, whereas the cerebellar cortex remained negative in all cases. In situ hybridization with digoxigenin-labeled minus-sense RNA probes, hybridizing with monomer-replicative form DNA or mRNA, revealed positive signals in nuclei of several neurons of the brain, again excluding the cerebellum. PCR applied to formalin-fixed and paraffin-embedded brain tissue and intestinal tissues of the diseased cats and subsequent DNA sequence analysis yielded canine parvovirus type 2 (CPV-2)-like sequences in the central nervous system. Two aspects of these findings are intriguing: (i) parvoviruses appear to be capable of replicating in neurons, cells that are considered to be terminally differentiated and (ii) CPV-like viruses of the old antigenic type CPV-2 appear to be able to infect cats.

IDIOPATHIC ACUTE ONSET MYELOPATHY IN CHEETAH (ACINONYX JUBATUS) CUBS

Abstract Numerous cases of ataxia, hind limb paresis, and paralysis have occurred in cheetah (Acinonyx jubatus) cubs over the past 10 yr within the European Endangered Species Program population, including 12 in mainland Europe, two in the British Isles, one in Namibia, and one in Dubai. The condition is the most important medical factor limiting European cheetah population growth. Eight cubs at the Salzburg Zoo, Austria, were affected. They demonstrated upper motor neuron lesions when alive and bilateral, symmetrical myelin degeneration of the spinal cord on necropsy. Ballooning of myelin sheaths surrounded mostly preserved axons, and no spheroids, characteristic of acute axonal degeneration, were found. Myelin loss markedly exceeded axonal degeneration. The syndromes etiology is unclear, although viral, bacterial, parasitic, genetic, nutritional–metabolic, toxic, and physical causes have been considered.

Evaluation of a Gene-Directed Enzyme-Product Therapy (GDEPT) in Human Pancreatic Tumor Cells and Their Use as In Vivo Models for Pancreatic Cancer

Background Gene-directed enzyme prodrug therapy (GDEPT) is a two-step treatment protocol for solid tumors that involves the transfer of a gene encoding a prodrug-activating enzyme followed by administration of the inactive prodrug that is subsequently activated by the enzyme to its tumor toxic form. However, the establishment of such novel treatment regimes to combat pancreatic cancer requires defined and robust animal model systems. Methods Here, we comprehensively compared six human pancreatic cancer cell lines (PaCa-44, PANC-1, MIA PaCa-2, Hs-766T, Capan-2, and BxPc-3) in subcutaneous and orthotopical mouse models as well as in their susceptibility to different GDEPTs. Results Tumor uptake was 83% to 100% in the subcutaneous model and 60% to 100% in the orthotopical mouse model, except for Hs-766T cells, which did not grow orthotopically. Pathohistological analyses of the orthotopical models revealed an infiltrative growth of almost all tumors into the pancreas; however, the different cell lines gave rise to tumors with different morphological characteristics. All of the resultant tumors were positive for MUC-1 staining indicating their origin from glandular or ductal epithelium, but revealed scattered pan-cytokeratin staining. Transfer of the cytochrome P450 and cytosine deaminase suicide gene, respectively, into the pancreatic cancer cell lines using retroviral vector technology revealed high level infectibility of these cell lines and allowed the analysis of the sensitivity of these cells to the chemotherapeutic drugs ifosfamide and 5-fluorocytosine, respectively. Conclusion These data qualify the cell lines as part of valuable in vitro and in vivo models for the use in defined preclinical studies for pancreas tumor therapy.

Immunohistochemical screening for viral agents in cheetahs (Acinonyx jubatus) with myelopathy

Numerous cases of acute-onset progressive ataxia, hindlimb paresis and paralysis of unknown aetiology occurred during 1993 to 2003 in cheetahs (Acinonyx jubatus) within the European Endangered Species Programme (eep). This study describes the immunohistochemical investigation of a possible viral aetiology of the ‘cheetah myelopathy’. Antibodies to feline herpesvirus type 1, canine distemper virus, canine parvovirus and Borna disease virus were applied to formalin-fixed and paraffin-embedded brain and spinal cord sections from 25 affected cheetahs aged between three-and-a-half months and 13 years. Using the avidin-biotin complex technique, none of the antibodies gave positive immunosignals in either the brain or the spinal cord tissue.

Construction and Rescue of a Molecular Clone of Deformed Wing Virus (DWV)

European honey bees are highly important in crop pollination, increasing the value of global agricultural production by billions of dollars. Current knowledge about virulence and pathogenicity of Deformed wing virus (DWV), a major factor in honey bee colony mortality, is limited. With this study, we close the gap between field research and laboratory investigations by establishing a complete in vitro model for DWV pathogenesis. Infectious DWV was rescued from a molecular clone of a DWV-A genome that induces DWV symptoms such as crippled wings and discoloration. The expression of DWV proteins, production of infectious virus progeny, and DWV host cell tropism could be confirmed using newly generated anti-DWV monoclonal antibodies. The recombinant RNA fulfills Koch’s postulates circumventing the need of virus isolation and propagation of pure virus cultures. In conclusion, we describe the development and application of a reverse genetics system for the study of DWV pathogenesis.

Evidence of canine distemper and suggestion of preceding parvovirus-myocarditis in a Eurasian badger (Meles meles).

Abstract An approximately 1.5-yr-old free-ranging male Eurasian badger (Meles meles) from the eastern part of Austria had macroscopic and microscopic lesions consistent with canine distemper virus infection, including nonsuppurative meningoencephalitis, interstitial pneumonia with accumulation of macrophages in alveoli that contained intranuclear inclusion bodies, vesicular exanthema of the ventral abdomen, and atrophy of lymphoid tissues. Canine distemper virus-antigen was demonstrable in a variety of organs by using immunohistology. In addition, there were widespread areas of fibrosis in the myocardium that were rich in collagen and paucicellular. Because such changes are comparable with sequelae of the acute cardiac form of canine parvovirus (CPV) infection in dogs, it was speculated that this badger may have experienced CPV myocarditis as a cub but that the corresponding antigen or DNA was not detectable due to resolution of the disease.

Meningoencephalitis in cats in Austria: a study of infectious causes, including Encephalitozoon cuniculi

Objectives Despite comprehensive diagnostics, the aetiology of meningoencephalitis (ME) in cats often remains undetermined. As a result of recently published surveys, Encephalitozoon cuniculi has gained growing importance in cats not only with ocular disorders, but also with central nervous system disease. Therefore, it was hypothesised that E cuniculi may be an underestimated pathogen in the development of feline non-suppurative and/or granulomatous ME. Methods As a first step, histopathological sections of the brain of cats with encephalopathy were retrospectively reviewed to identify cases of granulomatous ME. In a second step, an immunohistochemical screening for detection of E cuniculi was performed in cases with ME of unknown origin. Results In 59/89 (66.3%) cats with ME, an aetiologically relevant pathogen was detected. Forty-three of 89 (48.3%) cats had a diagnosis of feline infectious peritonitis. In 14/89 (15.7%) cats, protozoan cysts were identified and infection with Toxoplasma gondii was confirmed by immunohistochemistry (IHC) in all cases. In 2/89 (2.3%) cats with granulomatous ME, fungal organisms were identified. Thirty of 89 (33.7%) cats with ME of unknown origin that underwent IHC for the detection of E cuniculi remained negative. Conclusions and relevance The results of this study suggest that E cuniculi is unlikely to be directly associated with (non-suppurative and/or granulomatous) ME in cats in Austria.

NEPHROPATHIES IN THE EUROPEAN CAPTIVE CHEETAH (ACINONYX JUBATUS) POPULATION

Abstract According to previous studies in captive cheetah (Acinonyx jubatus) populations, one of the most threatening diseases besides amyloidosis, myelopathy, veno occlusive disease, and gastritis, is renal failure. Contrary to captive cheetahs in North America and South Africa, morphological data concerning renal lesions in the cheetah European Endangered Species Program (EEP) are lacking. This study details the histological characterization as well as immunohistochemical and morphometrical analysis of nephropathies in 35 captive cheetahs from the EEP, which were necropsied between 1985 and 2003. Examination of paraffin- and glycolmethacrylate-methylmethacrylate (GMA-MMA) embedded kidney samples by light microscopy revealed glomerulonephritis in 91%, with a high prevalence for glomerulosclerosis and glomerulonephritis with the histologic pattern of membranous glomerulonephritis (77%). Besides these predominating glomerulopathies, a wide range of other renal lesions, like acute tubular necrosis, interstitial nephritis, calcinosis, and amyloidosis, were present. Pathological expression of collagen type IV, complement C3, fibronectin, and IgG was demonstrated in the glomeruli of the cheetah kidneys with the use of the avidin-biotin complex method. Morphometrical analysis was performed on GMA-MMA embedded kidney samples to obtain glomerulosclerosis index and glomerulosclerosis incidence.