The molecular characterization of 4 Hungarian chicken anaemia virus (CAV) strains retrieved from naturally infected broiler chickens was studied, after the recent re-emergence of chicken infectious anaemia (CA). Carcasses from 4 Hungarian broiler flocks experiencing increased mortality were examined by necropsy, histopathology and polymerase chain reaction (PCR) based technique. The chickens presented stunted growth and general anaemia expressed by pallor and haemorrhages in the subcutaneous tissue. Blue wing disease was found in case of one flock (997/99), affecting 70% of the diseased chickens. Secondary bacterial and fungal infections were identified in several chickens from all investigated flocks. Histological examination revealed hypoplasia of the bone marrow and lymphocyte depletion in the spleen, bursa of Fabricius and thymus. The alignment from the deduced VP1 amino acid (aa) partial sequences (114 aas) showed high similarity (99%) among the Hungarian strains except for one that proved to have a higher variation, presenting 97% identity with the rest of the Hungarian CAV strains. Several aas substitution were identified in the Hungarian strains. Blocks of sequence data leading 344 bps were used for the phylogenetic analysis, and the Hungarian nucleotide sequences were compared with 22 other CAV sequences deposited in the GenBank to investigate any geographical relationship. The Hungarian sequences were not positioned in a separate group of the phylogenetic tree, nor did they cluster together only with European CAV strains. This study confirms the presence of CAV infection in Hungary, and presents the sequence and phylogenetic analysis of the circulating CAV strains.

Ten one-day-old goslings were inoculated orally with a Brachyspira alvinipulli strain isolated from the large intestine of geese that had died of intestinal spirochaetosis (Group A), 10 day-old goslings were inoculated orally with a B. hyodysenteriae strain (Group B), and a third group of 10 goslings (Group C) served as uninfected control. The goslings were observed daily for clinical signs. They were sacrificed on days 7, 14, 21 and 35 days postinfection (PI), and necropsied. Segments of the large intestine were subjected to histopathological, immunohistochemical, electron microscopic (TEM, SEM) and microbiological examinations. Mortality did not occur during the experimental period. However, in both groups the caecum of the goslings killed by bleeding was slightly dilated, in its lumen there was a watery, yellowish and frothy content, and the mucous membrane was slightly swollen. By histopathological, immunohistochemical and electron microscopic examination, B. alvinipulli and B. hyodysenteriae could be detected in the caecum or colon, in the lumen of the glands and sometimes among the glandular epithelial cells in goslings of the respective groups, and could be reisolated from these organs by culturing. A mild inflammation of the intestinal mucosa was also noted. In transverse section of the brachyspirae, numerous (16-22) periplasmic flagella could be detected inside the outer sheath, also depending on the plane of section.

Ten one-day-old goslings were inoculated orally with a Brachyspira alvinipulli strain isolated from the large intestine of geese that had died of intestinal spirochaetosis (Group A), 10 day-old goslings were inoculated orally with a B. hyodysenteriae strain (Group B), and a third group of 10 goslings (Group C) served as uninfected control. The goslings were observed daily for clinical signs. They were sacrificed on days 7, 14, 21 and 35 days postinfection (PI), and necropsied. Segments of the large intestine were subjected to histopathological, immunohistochemical, electron microscopic (TEM, SEM) and microbiological examinations. Mortality did not occur during the experimental period. However, in both groups the caecum of the goslings killed by bleeding was slightly dilated, in its lumen there was a watery, yellowish and frothy content, and the mucous membrane was slightly swollen. By histopathological, immunohistochemical and electron microscopic examination, B. alvinipulli and B. hyodysenteriae could be detected in the caecum or colon, in the lumen of the glands and sometimes among the glandular epithelial cells in goslings of the respective groups, and could be reisolated from these organs by culturing. A mild inflammation of the intestinal mucosa was also noted. In transverse section of the brachyspirae, numerous (16-22) periplasmic flagella could be detected inside the outer sheath, also depending on the plane of section.

A Lippizan mare aborted a male fetus a few days before the expected foaling date without showing any clinical sings. Focal lympho-histiocytic hepatitis in the foal and multiplex focal lympho-histiocytic villitis accompanied by villus necroses and marked hypertrophy of chorionic epithelial cells in the arcades were observed. Elongated nucleated organisms were seen in groups in vacuoles or solitarily located in the cytoplasm of the chorionic epithelial cells. The organisms were in large numbers and often extracellularly in areas of villitis and villus necroses. They were Gram-positive, stained with haematoxylin and eosin (HE), periodic acid-Schiff (PAS) and Giemsa, weakly with Warthin-Starry silver stain but not with Gömöri's methenamine-silver stain. By ultrastructural and immunohistochemical examinations, the organisms were identified as microsporidia belonging to the genus Encephalitozoon. No Encephalitozoon organisms were detected in the fetal organs. This is the first reported case of equine abortion induced by Encephalitozoon sp. in Europe. Although abortion induced by Encephalitozoon is rare, microsporidia should be considered a differential diagnosis for intracellular organisms observed in the chorionic epithelial cells of horses.

Histological and electron microscopic examinations of the kidneys of 8 dogs suffering from fatal, naturally acquired Babesia canis infection and nephropathy are presented. Seven animals were treated with imidocarb dipropionate on average 4.5 days prior to death. Severe anaemia was present only in 2 cases. Degenerative histological changes observed mostly in the proximal convoluted tubules included vacuolar-hydropic degeneration, necrosis and detachment of renal tubular epithelial (RTE) cells from the basement membrane. Necrotic debris occasionally formed acidophilic casts within the tubules. In some cases, necrosis of the whole tubule was observed. Haemoglobin casts in the tubules and haemoglobin droplets in RTE cells seldom appeared. No significant histological changes were seen in the glomeruli. Ultrastructural lesions in RTE cells included nuclear membrane hyperchromatosis, karyopyknosis, karyolysis, swelling or collapse of mitochondria with fragmentation of cristae and vacuolar-hydropic degeneration in the endoplasmic reticulum and microvilli. Nuclear oedema was also observed. Many RTE cells exhibiting necrosis collapsed. Vacuolar-hydropic degeneration and necrosis were also observed in the glomerular and interstitial capillary endothelium. The severe acute tubular necrosis described in this study is probably the result of hypoxic renal injury. Systemic hypotension leading to vasoconstriction in the kidneys might be the most important cause of renal hypoxia in B. canis infections, but anaemia may also contribute to inadequate oxygenation. Imidocarb should be applied with caution in patients with possible renal involvement until further data become available on its potential nephrotoxicity in dogs.

Avian nephritis virus (ANV) infection was detected in 4-day-old to 22-day-old chickens collected on Hungarian farms between 2002 and 2005. The animals suffered from diarrhoea, growth retardation, runting-stunting syndrome, and 2 to 6% mortality was reported. Tubulonephrosis, interstitial nephritis and uricosis ( gout) was diagnosed. The presence of ANV RNA was detected in chicken carcasses using reverse transcriptase-polymerase chain reaction. The virus was demonstrated in 69% of the investigated farms. The nucleotide sequence of the amplification products ( corresponding to part of the genome that encodes the GP1 protein) was determined and phylogenetic analysis was performed. The nucleotide sequences showed 76 to 86% identity to the reference strain isolated in 1976 in Japan. The constructed phlyogenetic tree indicates high diversity of the Hungarian ANV sequences, regardless of their origin and year of sample collection. Analysis of the putative amino acid sequences encoded by the partial GP1 sequences also revealed high diversity of the virus. Even samples collected at the same farm, at the same time but from different flocks, differed in nucleotide and putative amino acid sequences. The possible effects of the sequence diversity on the pathogenicity, antigenicity and diagnostics of ANV are discussed.

Kidney samples from chickens diagnosed with acute nephritis and gout were subjected to histological and electron microscopic examination. The investigations revealed cytoplasmic inclusion bodies in the tubular epithelial cells containing round virions of about 30 nm in diameter. Since avian nephritis virus (ANV) is known as a potential causative agent of the so-called baby chick nephropathy, an RT-PCR assay was developed for the molecular detection of ANV-specific nucleic acid in the specimen. The specificity of the assay was confirmed by direct sequencing of the amplicon obtained in the reaction. The nucleotide sequence of the PCR product showed 92% identity with the reference ANV sequence deposited in the GenBank database. After having been validated on some other suspicious cases of avian nephritis, the PCR method described in this study can be a potential tool for routine diagnostic examination of samples submitted from cases of gout and nephropathy in chickens.

Goose embryos were infected with goose haemorrhagic polyomavirus (GHPV) onto the chorioallantoic membrane (CAM) in order to examine the effect of GHPV on the embryos and to obtain data on whether embryos could develop into infected, virus-shedding goslings, as well as to present an accurate biological method for virus titration. The reported method of infection could offer a possibility to express the virus titre as the median embryo infective dose (EID50). As a special pathological feature of the disease, extensive cerebral haemorrhages were observed, which protruded the skullcap in many cases. Some embryos infected with 101.25 or 100.25 EID50/0.2 ml were able to hatch; however, they were in poor physical condition and died by post-hatching day 4 showing haemorrhagic nephritis and enteritis of geese. Virus shedding was revealed by polymerase chain reaction. The ability of some of the infected goose embryos to hatch may indicate the potency of GHPV to spread vertically, although this needs further study for confirmation. Pathologie du polyomavirus hémorragique de l'oie (GHPV) pour les embryons d'oie Des embryons d'oie ont été infectés par le polyomavirus hémorragique de l'oie (GHPV) au niveau de la membrane chorio-allantoïdienne (CAM) dans le but (i) d'examiner l'effet du GHPV sur les embryons et (ii) d'obtenir des données sur le développement éventuel de l'embryon dans les conditions d'infection, la diffusion du virus par les oisons, ainsi que (iii) de présenter une méthode biologique précise pour le titrage du virus. La méthode d'infection qui est présentée permet d'exprimer le titre du virus en DIE50. Une caractéristique de la pathologie de cette maladie est l'importance des hémorragies cérébrales observées qui, dans de nombreux cas, rendent proéminente la calotte crânienne. Quelques embryons infectés avec 101.25 DIE50/0.2 ml ou 100.25 DIE50/0.2 ml ont pu éclore; toutefois, ils étaient dans de mauvaises conditions physiques et sont morts 4 jours après l'éclosion présentant une néphrite hémorragique entérite de l'oie (HNEG). La diffusion du virus a été mise en évidence par PCR. Le fait que quelques embryons d'oie infectés éclosent permet d'indiquer que le virus peut se transmettre verticalement, mais des études complémentaires sont nécessaires pour le confirmer. Pathologie der Infektion mit hämorrhagischen Gänsepolyomavirus in Gänseembryonen Es wurden Gänseembryonen mit dem hämorrhagischen Gänsepolyomavirus (GHPV) mittels Inokulation auf die Chorioallantoismembran infiziert, um (i) den Effekt von GHPV auf Embryonen zu untersuchen, (ii) Erkenntnisse darüber zu gewinnen, ob sich infizierte Embryonen zu Virus ausscheidenden Gänseküken entwickeln können und (iii) eine exakte biologische Methode für die Virustitration präsentieren zu können. Die beschriebene Methode könnte eine Möglichkeit darstellen, den Virustiter als EID50 zu bestimmen. Als eine spezifische pathologische Veränderung der Erkrankung wurden extensive zerebrale Hämorrhagien, die sich in vielen Fällen bis unter die Schädeldecke ausstreckten, beobachtet. Einige Embryonen, die mit 101,25 oder 100,25 EID50/0,2 ml infiziert worden waren, konnten schlüpfen, aber waren in schlechtem Allgemeinzustand und starben 4 Tage nach dem Schlupf, wobei sie das Krankheitsbild der hämorrhagische Nephritis und Enteritis der Gänse (HNEG) zeigten. Die Virusausscheidung wurde mittels PCR nachgewiesen. Die Schlupffähigkeit einiger infizierter Embryonen weist auf eine mögliche vertikale Ausbreitung von GHPV hin, obwohl weitere Untersuchungen nötig sind, um dies zu bestätigen. Patología causada por la infección por Poliomavirus hemorrágico del ganso (GHPV) en embriones de ganso Se infectaron embriones de ganso con Poliomavirus hemorrágico del ganso (GHPV) en la membrana corioalantoidea (CAM) con la finalidad de (i) examinar el efecto de GHPV en los embriones y (ii) obtener datos sobre la posibilidad de que los embriones infectados puedan nacer y dar lugar a gansos que excretan virus, así como también (iii) presentar un método biológico preciso para titular el virus. El método descrito puede ofrecer la posibilidad de expresar el título vírico como EID50. Se observaron, como lesión especial de la enfermedad, hemorragias cerebrales extensas, que afectaban el cráneo en múltiples casos. Algunos embriones infectados con 101.25 EID50/0.2 ml o 100.25 EID50/0.2 ml fueron capaces de nacer, pero presentaban una pobre condición física y murieron a los 4 días tras el nacimiento (PH) mostrando nefritis y enteritis hemorrágica del ganso (HNEG). La excreción vírica se evaluó mediante PCR. La capacidad de algunos de los embriones infectados para nacer puede indicar la potencia de GHPV para transmitirse verticalmente, pero este hecho requerirá estudios confirmatorios.

The authors describe the occurrence of an infectious chicken disease, avian nephritis for the first time in Hungary, which was suspected to be present from the early '70s, but the evidence was provided by a recently developed PCR based diagnostic method. The infectious nephritis is a component of the so called "baby chick nephropathy", referring to the localization and early appearance of the symptoms. The chickens from the third day after hatching develop diarrhea and weakness following per-oral infection: retarded growth is observed from the second week post infection. The mortality is usually low, except those cases where the tubular epithelium of the kidney is damaged due to the virus replication, nephroso-nephritis develops and the regressive changes result in uricosis (gout). The loss of the disease may be decreased or prevented if predisposing factors (such as keeping problems or malnutrition) are recognized and eliminated. In order to detect avian nephritis virus (ANV), kidney samples from chickens suffering acute nephritis and gout diagnosed by necropsy were subjected to histopathologic examination, and subsequently by RNA extraction/purification and polymerase chain reaction following reverse transcription (RT-PCR). A fast and reliable ANV specific RT-PCR based detection method was developed to identify the virus. Using this test the authors proved the presence of the avian nephritis virus in several Hungarian flocks and the results of the screening helped to clarify the occurrence of this viral disease. According to their data the infection is widely distributed in the Hungarian flocks, and avian nephritis virus infection might more often be the reason for the improper breeding results in chicken industry, than diagnosed.

Haemorrhagic nephritis and enteritis of geese as a new disease was first described in Hungary in 1969. The authors identified the causative agent of the outbreaks occurring in 1969 as a polyomavirus by PCR in 2001. In order to study the pathogenesis of the virus, one-day-old goslings were infected with tissue homogenate that tested positive for polyomavirus by PCR. Morphological, light and transmission electron microscopic (TEM) examinations have revealed that goose haemorrhagic polyomavirus replicates in the endothelial cells of the blood vessels and capillaries of diseased birds. Infection causes damage and necrosis of the endothelial cells. The virus was not observed in the parenchymal cells. Oedema and haemorrhages found throughout the body may be due to the dysfunction or functional deficiency of endothelial cells damaged by the virus.

The authors examined 3 English bulldog puppies died on 7-9th day with clinical signs of half-a-day-, one-day-apathy and respiratory distress and finally opistotonus, which puppies originated from a breed of 30 adult dogs, were born with caesarean section, developed properly and seemed to be healthy. In the other litters of the same age or which were born later in the breed, no similar disease was observed. At necropsy, interstitial pneumonia with oedema, accumulation of straw-yellow coagulable serum in the thoracic cavity, hepatitis with necroses of pinhead size and pinprick or lentil sized haemorrhages under the epicardium, serous membrane of the stomach and intestines and renal capsule were found. At pathohistological examination, intralobular-interstitial pneumonia with the development of necrotic foci and catarrhal hepatitis, acute inflammation of the spleen and intestines and lymphocytic encephalitis were found. Acidophylic inclusion bodies were mainly found in the nuclei of damaged hepatic cells. By electron microscope, herpesvirus-particles were demonstrated in the latters. Canine herpesvirus (CHV) was isolated from spleen, liver and kidney samples from 3 died puppies, on canine kidney cell culture in a thermostat of 5% CO2 content, 100% vapour content and 33 degreesC.

An outbreak of the atypical form of myxomatosis struck a rabbit farm in Hungary. The animals had previously been vaccinated with a vaccine containing Shope rabbit fibroma virus strain. The disease appeared in winter when the presence of mosquitoes and fleas is not common. The virus was isolated from an eyelid specimen of a naturally infected rabbit. The surviving animals were observed for four weeks, blood samples were collected and, after euthanasia, organ specimens were also examined by morphological methods including pathology and electron microscopy. Serum samples were examined by virus neutralisation for antibodies. Genetic analysis of the isolated virus was carried out by polymerase chain reaction (PCR) and direct sequencing. The primers were designed on the basis of the major envelope gene (Env) of the Lausanne reference strain in the GenBank. The viral proteins were examined by SDS-PAGE. The isolated virus (ref. no.: BP04/2001) was able to infect the susceptible animals directly, by contact. The disease was characterised by respiratory symptoms of the upper tracheal tract, conjunctivitis and high mortality by the 11th-14th day. Aerogenic infection with strain BP04/2001 resulted in 100% morbidity among the susceptible animals. Sequencing of the amplified 400-bp-long DNA revealed 97% homology with the Env gene of the Lausanne strain, which proves that strain BP04/2001 is a variant of the Lausanne strain having been enzootic throughout Europe. The live vaccine strain used in Hungary against myxomatosis, which is also a Lausanne-derived strain, protected the animals. According to the protein analysis a protein of 200 kDa in size is not expressed in strain BP04/2001. This is the first report on atypical myxomatosis in Central Europe. The virus spreads by airborne transmission and may cause severe losses in the rabbit population.

The pathology, epizootiology and aetiology of a specific disease of young geese, which has been seen in Hungary for more than three decades, were investigated. The disease was characterised by splenitis and hepatitis with miliary necrotic foci during the acute phase, and epicarditis, arthritis and tenosynovitis during the subacute/chronic phase. Clinical signs usually appeared at 2 to 3 weeks of age and persisted for 3 to 6 weeks. From different organs of the affected birds, a reovirus was isolated in embryonated eggs and tissue cultures of Muscovy duck or goose origin, as well as in Vero cells. In experimental infections, the dominant features of the disease were reproduced in day-old and young goslings. The biological and partial molecular characterisation of one of the isolated strains (D15/99) showed that it was related to the reovirus described as the cause of a similar disease of Muscovy ducks. An RT-PCR method suitable for the detection of reoviruses was also elaborated and tested. This is the first report on the involvement of reovirus in arthritis of geese.

The authors present a case of poxvirus caused nodular dermatitis diagnosed in emerald swift (Sceloporus malachiticus) kept in a Hungarian zoo. The disease was described so far only in other reptile species. They call attention to the importance of strict compliance with quarantine measures in case of imported exotic animals.

The reproductive toxicity of lead acetate and of a fungicide formulation (Dithane M-45) containing 80% mancozeb was studied on rats. Lead acetate was applied in the feed in the following dose groups: control, 1,000, 5,000 and 10,000 mg/kg of diet. The three treatment groups received, in addition to the above doses of lead acetate, 4,500 mg/kg Dithane M-45 in the diet. The method was based on the OECD Guideline for Testing of Chemicals No. 415 (1981). Clinical symptoms and mortality were not found in the parent generation. The body weight of female animals decreased significantly before the pregnancy period. This tendency was also seen in males after the combination treatment. At the two high dose levels a remarkable body weight increase was seen in the female animals during the lactation period. As a result of treatment, decreased body weight of offspring was measured during the lactation period. No gross pathological changes were seen. Histological examination showed general tubulonephrosis in the experimental animals. It can be established that the administration of Dithane M-45 did not enhance the reproductive toxicity of lead acetate.