Acta Veterinaria Hungarica 53 (1) (2005)

Acta Veterinaria Hungarica 53 (1) (2005)

CONTENTS AND ABSTRACTS

Anatomy and histology

Morphology of the lymph nodes in bottlenose dolphin (Tursiops truncatus) and striped dolphin (Stenella coeruleoalba) from the Adriatic Sea. Snježana Vuković, H. Lucić, H. Gomerčić, Martina Đuras Gomerčić, T. Gomerčić, Darinka Škrtić and Snježana Ćurković. pp. 1-11.
Bacteriology

Identification of Borrelia burgdorferi genospecies inducing Lyme disease in dogs from Western Poland. Bogumiła Skotarczak and Beata Wodecka. pp. 13-21.
Clinical veterinary medicine

Comparison of two-dimensional echocardiographic measurements of the left atrium in healthy dogs. Cs. Hetyey, K. Vörös and J. Reiczigel. pp. 23-33.
Mycotoxin research

Ochratoxin A content of urine samples of healthy humans in Hungary. B. Fazekas, Andrea Tar and Melinda Kovács. pp. 35-44.
Parasitology

Histopathological changes caused by the metacestodes of Neogryporhynchus cheilancristrotus (Wedl, 1855) in the gut of the gibel carp, Carassius gibelio. K. Molnár. pp. 45-52.

Investigation of lectin activity in Theileria annulata piroplasms. Ö. Kaynar, T. Güldür and T. Karapinar. pp. 53-63.

Evaluation of efficacy of entomopathogenic nematodes against larvae of Lucilia sericata (Meigen, 1826) (Diptera: Calliphoridae). Erika M. Tóth, K. Márialigeti, A. Fodor, A. Lucskai and R. Farkas. pp. 65-71.
Pathology

Comparative pathological studies on domestic geese (Anser anser domestica) and Muscovy ducks (Cairina moschata) experimentally infected with parvovirus strains of goose and Muscovy duck origin. R. Glávits, Anna Zolnai, Éva Szabó, Éva Ivanics, P. Zarka, T. Mató and V. Palya. pp. 73-89.
Reproduction

The effect of nerve growth factor on nuclear progression of porcine oocytes during in vitro maturation and embryo development. Ágnes Bali Papp, T. Somfai, Mabel Tartaglione, Erika Varga and J. C. Gardon. pp. 91-101.

Vitrification of biopsied mouse embryos. B. Baranyai, Sz. Bodó, A. Dinnyés and Elen Gócza. pp. 103-112.
Virology

The molecular diagnosis of porcine viral diseases: A review. S. Belák. pp. 113-124.

Cytopathogenicity markers in the genome of Hungarian cytopathic isolates of bovine viral diarrhoea virus. Á. Bálint, Claudia Baule, S. Kecskeméti, I. Kiss and S. Belák. pp. 125-136.

Detection of respiratory and enteric shedding of bovine coronaviruses in cattle in Northwestern Turkey. M. Hasoksuz, A. Kayar, T. Dodurka and A. Ilgaz. pp. 137-146.

The replication of canine herpesvirus (CHV) induces apoptosis in canine kidney cell line: Short communication. O. Kim and S. J. Yi. pp. 147-151.

Acta Veterinaria Hungarica 53 (1), pp. 1–11 (2005)

MORPHOLOGY OF THE LYMPH NODES IN BOTTLENOSE DOLPHIN (Tursiops truncatus) AND STRIPED DOLPHIN (Stenella coeruleoalba) FROM THE ADRIATIC SEA

Snježana Vuković^1*, H. Lucić¹, H. Gomerčić¹, Martina Đuras Gomerčić¹, T. Gomerčić², Darinka Škrtić¹ and Snježana Ćurković¹

1Department of Anatomy, Histology and Embryology, Faculty of Veterinary Medicine, University of Zagreb, Heinzelova 55, 10 000 Zagreb, Croatia; ²Department of Biology, Faculty of Veterinary Medicine, University of Zagreb, Zagreb, Croatia

(Received February 5, 2004; accepted September 1, 2004)

Morphology of the lymph nodes was examined in six bottlenose dolphins (Tursiops truncatus) and three striped dolphins (Stenella coeruleoalba) from the Adriatic Sea. All animals had been found dead in nature. One group of the nodes was taken from the tracheal branching area and was marked as bifurcational lymph node, and the other group was taken from the mesenteric root and was marked as mesenteric lymph node. Microscopic analysis showed that the lymph nodes in both dolphin species were surrounded by a connective tissue capsule comprising smooth muscle cells. The parenchyma of the mesenteric and bifurcational lymph nodes in bottlenose dolphin was divided into the peripherally situated cortex with the lymphatic nodules and diffuse lymphatic tissue, and the centrally situated medulla structured of the medullary cords separated by the medullary sinuses. These lymph nodes structurally correspond to the lymph nodes in the majority of terrestrial mammals. The mesenteric lymph node of striped dolphin also had a peripherally situated cortex and a centrally positioned medulla as the majority of terrestrial mammals. In the bifurcational lymph nodes of striped dolphin, there was a central dense lymphatic tissue with the lymphatic nodules and a peripheral less dense lymphatic tissue structured of the cell cords and sinuses. The bifurcational lymph node in striped dolphin resembled porcine lymph nodes and belonged to the inverse lymph nodes.

Key words: Lymph node, bottlenose dolphin, striped dolphin, terrestrial mammals, pig

*Corresponding author: Assoc. Prof. Snježana Vuković; E-mail: svukovic@vef.hr; Phone: +385 1 23 90 249; Fax: +385 1 24 41 390

Acta Veterinaria Hungarica 53 (1), pp. 13–21 (2005)

IDENTIFICATION OF BORRELIA BURGDORFERI GENOSPECIES INDUCING LYME DISEASE IN DOGS FROM WESTERN POLAND

Bogumiła Skotarczak^* and Beata Wodecka

Department of Genetics, Faculty of Biology, Szczecin University, 71-215 Szczecin, Piastow 40B, Poland

(Received March 8, 2004; accepted May 24, 2004)

Canine Lyme borreliosis may be caused by three Borrelia burgdorferi sensu lato genospecies. The prevalence of infection by Borrelia species was determined by nested polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) with the enzyme Fsp4H I in the blood of dogs naturally infested by ticks in an endemic region of Poland. Blood samples were collected from 98 dogs of various breeds, delivered to the Veterinary Clinic in Szczecin (northwestern Poland) for various reasons. Nested PCR revealed the presence of DNA characteristic of only 1 genospecies, i.e. B. burgdorferi sensu stricto (s.s.), in all PCR-positive samples. Digestion of PCR products from a fragment of the fla gene amplified with primers FLA1 and FLA2 gave only one band pattern consistent with the pattern obtained from sequence analysis of the fla gene from a reference isolate of B. burgdorferi s.s. GeHo (X15660) from GenBank.

Key words: Borrelia burgdorferi genospecies, canine borreliosis

*Corresponding author; E-mail: Bogumila_Skotarczak@sus.univ.szczecin.pl; Phone/Fax: 48 91 4442782

Acta Veterinaria Hungarica 53 (1), pp. 23–33 (2005)

COMPARISON OF TWO-DIMENSIONAL ECHOCARDIOGRAPHIC MEASUREMENTS OF THE LEFT ATRIUM IN HEALTHY DOGS

Cs. Hetyey^1*, K. Vörös¹ and J. Reiczigel²

1Department and Clinic of Internal Medicine, ²Department of Biomathematics and Informatics, Faculty of Veterinary Science, Szent István University, H-1078 Budapest, István u. 2, Hungary

(Received April 15, 2004; accepted May 24, 2004)

The aim of the study was to establish normal echocardiographic values of the left atrium just above the mitral annulus (LA_ama) in healthy dogs. In the first part of the study 20 dogs of various breeds were examined. The diameter of the left atrium just above the mitral annulus (LA_ama) was compared to the linear (left to right) dimension of the left atrium (LA_r–l) as published earlier. There was a linear relationship between LA_r–l and the body weight (LA_r–l = 0.5061 BW (kg) + 22.206; R² = 0.81), but the proportion of LA_r–l/LA_ama was independent of the body weight (LA_r–l/LA_ama = 0.0004 BW (kg) + 1.0833; R² = 0.01). In the second part of the study the left atrial diameter just above the mitral annulus (LA_ama) was measured in 121 dogs of various breeds. There was a positive linear relationship (R² = 0.697; p < 0.001) between body weight and LA_ama (LA_ama = 0.54 BW (kg) + 18.502 ± 4.76), but there were no significant correlations between the age of animals and LA_ama (p = 0.45) as well as between the gender of animals and LA_ama (p = 0.78). Two-dimensional echocardiographic (2DE) determination of LA_ama as described in the present study can be recommended for use in those dogs where measurement of LA_r–l encounters technical difficulties. In these cases LA_r–l can be calculated from LA_ama using the formula LA_r–l/LA_ama = 0.0004 BW (kg) + 1.0833, worked out in the first experiment.

Key words: Two-dimensional echocardiography, measurement, dog, normal values, left atrium

*Corresponding author; E-mail: cshetyey@univet.hu; Fax: +36 (1) 478 4137

Acta Veterinaria Hungarica 53 (1), pp. 35–44 (2005)

OCHRATOXIN A CONTENT OF URINE SAMPLES OF HEALTHY HUMANS IN HUNGARY

B. Fazekas^1*, Andrea Tar¹ and Melinda Kovács²

1Veterinary Institute of Debrecen, H-4031 Debrecen, Bornemissza u. 3–5, Hungary; ²Animal Breeding and Animal Hygiene Research Group of the Hungarian Academy of Sciences and the University of Kaposvár, Faculty of Animal Science, University of Kaposvár, Kaposvár, Hungary

(Received April 7, 2004; accepted September 1, 2004)

The ochratoxin A (OTA) content of urine samples from 88 healthy humans living at five settlements in three counties of Hungary was determined by immunoaffinity column cleanup and high-performance liquid chromatography (HPLC). OTA was detected in 61% of the samples in an average concentration of 0.013 ng/ml (range: 0.006–0.065 ng/ml). OTA concentrations measured in urine samples from men and women were not significantly different. The OTA concentration of samples from Heves county was significantly (t-test; p < 0.003) higher than that of samples from Hajdú-Bihar and Somogy counties. The regional differences in OTA concentration of urine samples indicate regional differences in the OTA exposure of the human population. Further studies are necessary to determine the cause of the regional differences in the OTA intake. The studies allow us to conclude that the OTA intake of the majority of the Hungarian population is low (< 1 ng/kg of body weight per day) but a certain part of the rural population may take up higher levels of OTA.

Key words: Ochratoxin A, human urine

*Corresponding author; E-mail: fazekasb@oai.hu; Fax: +36 (52) 310 823

Acta Veterinaria Hungarica 53 (1), pp. 45–52 (2005)

HISTOPATHOLOGICAL CHANGES CAUSED BY THE METACESTODES OF NEOGRYPORHYNCHUS CHEILANCRISTROTUS (WEDL, 1855) IN THE GUT OF THE GIBEL CARP, CARASSIUS GIBELIO

K. Molnár^*

Veterinary Medical Research Institute, Hungarian Academy of Sciences, H-1581 Budapest, P.O. Box 18, Hungary

(Received May 10, 2004; accepted September 1, 2004)

Metacestodes of Neogryporhynchus cheilancristrotus (Wedl, 1855) were found in the gut of some gibel carp (Carassius gibelio) specimens from a Hungarian water reservoir. Location of metacestodes in the freshly opened gut was marked with disseminated, red-coloured, pinhead-sized nodules in the anterior part of the intestine. In histological sections, metacestodes were found in a hole inside the propria layer of the intestinal folds. The worms were in direct contact with the host tissue without being encapsulated as a result of host reaction. In some specimens with extruded rostellum the rostellar hooks were bored into the host tissue and suckers grabbed pieces of the surrounding connective tissue. Around the worms, congested capillaries and formation of macrophages were seen in the lysed connective tissue.

Key words: Gryporhynchidae, Neogryporhynchus, metacestodes, fish, gibel carp, histology, pathology

*E-mail: kalman@vmri.hu; Fax: +36 (1) 467 4076

Acta Veterinaria Hungarica 53 (1), pp. 53–63 (2005)

INVESTIGATION OF LECTIN ACTIVITY IN THEILERIA ANNULATA PIROPLASMS

Ö. Kaynar¹, T. Güldür^2* and T. Karapinar³

1Department of Biochemistry, Institute of Health Sciences, Firat University, 23119 Elazig, Turkey; ²Department of Biochemistry, Faculty of Medicine, Inönü University, 44069 Malatya, Turkey; ³Department of Internal Medicine, Faculty of Veterinary Medicine, Firat University, 23119 Elazig, Turkey

(Received December 23, 2003; accepted May 24, 2004)

Adhesion to target cells is an essential step in the pathogenesis of many protozoal infections. Some protozoa have been reported to have a lectin activity involved in their attachment to the cell surface. The ligand–receptor interaction involved in Theileria annulata infection is unclear at present, in spite of the fact that some aspects of the process have been investigated. To this end, T. annulata piroplasms have been screened for lectin activity. Blood taken from infected cattle was first depleted of leukocytes and then subjected to ammonium chloride lysis in order to isolate the piroplasms. The piroplasms were homogenised and a crude membrane extract was prepared by centrifugation. To investigate lectin activity in piroplasm proteins, a simple screening procedure was employed for analysing piroplasm proteins binding to various lectin ligands. Numerous immobilised lectin ligands (L-fucose-sepharose, N-acetyl-neuraminic acid-sepharose, N-acetyl-D-galactosamine-agarose, N-acetyl-D-glucosamine-agarose, D-mannose-agarose, ß-D-glucose-agarose, ?-methyl-D-mannoside-agarose) were incubated with T. annulata piroplasm crude membrane extract. The ligand-bound proteins were eluted and separated by a brief centrifugation and determined by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE). The present study suggests that a 32 kDa protein of piroplasm binds to D-galactosyl residues of the agarose matrix and that the binding is inhibited by galactose and not by the other monosaccharides tested.

Key words: Theileria annulata, piroplasm, gal-lectin

*Corresponding author: Dr. Tayfun Güldür, Inönü Üniversitesi, Tip Fakültesi, Biyokimya ABD, 44069 Malatya, Turkey; E-mail: tguldur@inonu.edu.tr; Fax: +90 (422) 3410048

Acta Veterinaria Hungarica 53 (1), pp. 65–71 (2005)

EVALUATION OF EFFICACY OF ENTOMOPATHOGENIC NEMATODES AGAINST LARVAE OF LUCILIA SERICATA (MEIGEN, 1826) (DIPTERA: CALLIPHORIDAE)

Erika M. Tóth^1*, K. Márialigeti¹, A. Fodor², A. Lucskai³ and R. Farkas⁴

1Department of Microbiology, Eötvös Loránd University, H-1117 Budapest, Pázmány P. sétány 1/c, Hungary; ²Department of Genetics, Eötvös Loránd University, Budapest, Hungary; ³Institute of Plant Protection, Georgikon Faculty of Agricultural Sciences, Veszprém University, Keszthely, Hungary; ⁴Department of Parasitology and Zoology, Faculty of Veterinary Science, Szent István University, Budapest, Hungary

(Received April 16, 2004; accepted May 24, 2004)

The blowfly Lucilia sericata (Meigen, 1826) (Diptera: Calliphoridae) is the primary agent of cutaneous myiasis of sheep in northern Europe, southern Africa, Australia and New Zealand. As the application of chemicals has several disadvantages, alternative control measures of traumatic myiasis of livestock must be developed. In this study, the use of entomopathogenic nematodes (EPNs) as potential biocontrol agents against second instar larvae of Lucilia sericata was considered. The following nematode species were tested: Heterorhabditis bacteriophora (IS 5, HHU 1, Hmo1, HNC 1, HAZ 36, Hbrecon, HHU 2, HAZ 29, HHP 88, HHU 3, HHU 4 and HGua), Steinernema intermedia, NC513 strain of S. glaserii, S. anomali, S. riobrave, Steinernema sp. and 5 strains of S. feltiae (22, Vija Norway, HU 1, scp, and IS 6). None of the examined EPN species or strains showed larvicidal efficacy at 37°C (no killing effect was observed in the case of the two heat-tolerant strains – H. bacteriophora and S. feltiae) against L. sericata larvae. At lower temperatures (20°C and 25°C) only strains of S. feltiae were found to be active. The overall odds ratios calculated for L. sericata maggots to contract S. feltiae nematode infection show significant (p < 0.05) effect only in the case of strains HU 1, 22 and IS 6. In the case of strains HU 1 and 22 parasitic forms of S. feltiae could be detected in the dead larvae of L. sericata. Strain IS 6 (and also Vija Norway at 20°C) penetrated and killed fly larvae, but only adult forms of the nematode occurred in the cadavers.

Key words: Myiasis, blowfly, Lucilia sericata, larvicidal efficacy, entomopathogenic nematodes

*Corresponding author: Erika M. Tóth, Ph.D.; E-mail: totherika@ludens.elte.hu; Phone: +36 (1) 381 2177; Fax: +36 (1) 381 2178

Acta Veterinaria Hungarica 53 (1), pp. 73–89 (2005)

COMPARATIVE PATHOLOGICAL STUDIES ON DOMESTIC GEESE (ANSER ANSER DOMESTICA) AND MUSCOVY DUCKS (CAIRINA MOSCHATA) EXPERIMENTALLY INFECTED WITH PARVOVIRUS STRAINS OF GOOSE AND MUSCOVY DUCK ORIGIN

R. Glávits^1*, Anna Zolnai², Éva Szabó³, Éva Ivanics¹, P. Zarka¹, T. Mató² and V. Palya²

1Central Veterinary Institute, H-1581 Budapest 146, P.O. Box 2, Hungary; ²CEVA-Phylaxia Veterinary Biologicals Co. Ltd., Budapest, Hungary; ³PROPHYL Ltd., Mohács, Hungary

(Received June 23, 2004; accepted September 1, 2004)

Parvovirus infection of Muscovy ducks caused by a genetically and antigenically distinct virus has been reported from Germany, France, Israel, Hungary, some Asian countries and the USA. The pathological changes include those of degenerative skeletal muscle myopathy and myocarditis, hepatitis, sciatic neuritis and polioencephalomyelitis. In the study presented here, day-old and 3-week-old goslings and Muscovy ducks were infected experimentally with three different parvovirus strains (isolates of D-216/4 from the classical form of Derzsy’s disease, D-190/3 from the enteric form of Derzsy’s disease, and strain FM from the parvovirus disease of Muscovy ducks). All three parvovirus strains caused severe disease in both day-old and 3-week-old Muscovy ducks but in the goslings only the two strains of goose origin (D-216/4 and D-190/3) caused disease with high (90–100%) mortality when infection was performed at day old. Strain FM (of Muscovy duck origin) did not cause any clinical signs or pathological lesions in the goslings. In the day-old goslings and Muscovy ducks the principal pathological lesions were severe enteritis with necrosis of the epithelial cells (enterocytes) of the mucous membrane and the crypts of Lieberkühn, and the formation of intranuclear inclusion bodies. Other prominent lesions included hepatitis and atrophy (lymphocyte depletion) of the lymphoid organs (bursa of Fabricius, thymus, spleen). In goslings infected with the strain originating from the classical form of Derzsy’s disease mild myocarditis was also detected. After infection at three weeks of age, growth retardation, feathering disorders, myocardial lesions (degeneration of cardiac muscle cells, lympho-histiocytic infiltration) and hepatitis were the most prominent lesions in both geese and Muscovy ducks. In addition to the lesions observed in the geese, muscle fibre degeneration, mild sciatic neuritis and polioencephalomyelitis were also observed in the Muscovy ducks infected with any of the three parvovirus strains.

Key words: Goose, Muscovy duck, parvovirus, pathology

*Corresponding author; E-mail: glavitsr@oai.hu; This work was presented as a poster at the XIIth World’s Poultry Congress (Istanbul, Turkey, June 8–13, 2004)

Acta Veterinaria Hungarica 53 (1), pp. 91–101 (2005)

THE EFFECT OF NERVE GROWTH FACTOR ON NUCLEAR PROGRESSION OF PORCINE OOCYTES DURING IN VITRO MATURATION AND EMBRYO DEVELOPMENT

Ágnes Bali Papp^1*, T. Somfai¹, Mabel Tartaglione², Erika Varga¹ and J. C. Gardon²

1Institute of Animal Breeding, University of West Hungary, H-9200 Mosonmagyaróvár, Vár 4, Hungary; ²Faculty of Agrarian Sciences, Universidad Nacional de Lomas Zamora, Buenos Aires, Argentina

(Received March 30, 2004; accepted May 24, 2004)

The present study examined the effect of nerve growth factor (NGF) on in vitro maturation (IVM), in vitro fertilisation (IVF) and subsequent embryonic development of porcine oocytes. Cumulus-oocyte complexes were cultured with or without 1.0 ng/ml NGF for 40 h. After IVF, they were cultured in vitro for 6 days. After 10 and 20 h of IVM, there was no difference in nuclear status between the NGF-treated and control oocytes. Significant differences were detected in nuclear progression of oocytes matured in the presence or absence of NGF at 30 h of culture. A higher proportion of NGF-treated oocytes were at M-II stage compared to the control. Nevertheless, at the end of the 40-h IVM period, there was no difference in the proportion of M-II stage oocytes between the NGF-treated and control groups. NGF in IVM medium did not influence the developmental competence of putative embryos. Most embryos remained at the 2- to 4-cell stage; however, a significant amount of embryos reached the morula stage both in the NGF and the control groups. These results suggest that NGF during IVM accelerates nuclear progression of porcine oocytes by enhancing the post-diakinetic events of meiosis.

Key words: In vitro maturation, fertilisation, embryo development, nerve growth factor

*Corresponding author; E-mail: bali@mtk.nyme.hu; Phone: +36 (96) 566 613; Fax: +36 (96) 566 695

Acta Veterinaria Hungarica 53 (1), pp. 103–112 (2005)

VITRIFICATION OF BIOPSIED MOUSE EMBRYOS

B. Baranyai, Sz. Bodó, A. Dinnyés and Elen Gócza^*

Department of Animal Biology, Agricultural Biotechnology Center, H-2103 Gödöllő, Szent-Györgyi A. u. 4, Hungary

(Received May 3, 2004; accepted May 24, 2004)

Solid surface vitrification (SSV) was compared with in-straw vitrification for cryopreservation of biopsied mouse embryos. Eight-cell stage embryos were zona drilled and one blastomere was removed. Developed morulae or blastocysts were vitrified in microdrop (35% EG + 5% PVP + 0.4 M trehalose) or in straw (7.0 M EG + 0.5 M sucrose). Following recovery, embryos were cultivated in vitro or transferred into recipients. Cryopreservation had an effect not only on the survival of biopsied embryos but also on their subsequent development in vitro. Cryosurvival of biopsied morulae vitrified in straw was significantly inferior to SSV. The post-warm development of biopsied and non-biopsied morulae was delayed on Day 3.5 and 4.5 in both vitrification groups. A delay in development was observed on Day 5.5 among vitrified non-biopsied blastocysts. The percentage of pups born from biopsied morulae or blastocysts following cryopreservation did not differ from that of the control. No significant differences could be detected between methods within and between embryonic stages in terms of birth rate. The birth rate of biopsied embryos vitrified in straw was significantly lower compared to the non-biopsied embryos. The novel cryopreservation protocol of SSV proved to be effective for cryopreservation of morula- and blastocyst-stage biopsied embryos.

Key words: Embryo biopsy, cryopreservation, mouse, solid surface vitrification

*Corresponding author; E-mail: elen@abc.hu; Phone: +36 (28) 526 162; Fax: +36 (28) 526 151

Acta Veterinaria Hungarica 53 (1), pp. 113–124 (2005)

THE MOLECULAR DIAGNOSIS OF PORCINE VIRAL DISEASES: A REVIEW

S. Belák^*

Joint Research and Development Division, Departments of Virology, Swedish University of Agricultural Sciences (SLU) and National Veterinary Institute (SVA), Ulls väg 2B, SE-756 89 Uppsala, Sweden

(Received May 3, 2004; accepted September 1, 2004)

The worldwide occurrence and re-occurrence of transboundary diseases like foot-and-mouth disease or classical swine fever indicates that there is a high need for the development of powerful, robust and high-capacity new diagnostic methods, which are able to detect the causative agents before they could spread to large populations and cause tremendous losses. This article reports the experiences of a research group on the development of molecular methods for the improved diagnosis of a range of porcine viral diseases, including diseases on List A of the Office International des Epizooties (OIE). Nucleic acid hybridisation and various polymerase chain reaction (PCR) assays have been applied for routine diagnosis of a large range of viral diseases. During the last one-and-a-half decade more than 40 nested PCR assays have been developed to detect a variety of DNA and RNA viruses. False positive and negative results are avoided by the use of special tools, practices and internal controls of amplification (mimics). Recently, real-time PCR methods (TaqMan, molecular beacons, Primer-Probe Energy Transfer system) have been developed for the diagnosis of a wide range of diseases, such as foot-and-mouth disease, swine vesicular disease and vesicular stomatitis. Multiplex PCR packages have been developed for the simultaneous detection of eight important viruses of swine. By introducing nucleic acid extraction and pipetting robotics, together with the multi-channel real-time PCR machines, the diagnostic procedures have become rapid, robust and automated. In order to standardise the real-time PCR assays, the rules of OIE are considered. By following the five steps of OIE standardisation and validation, the new diagnostic procedures are nationally and internationally standardised and harmonised. The rapid, powerful and internationally standardised molecular diagnosis contributes to the reduction of losses caused by the transboundary viral diseases in swine populations.

Key words: Molecular diagnosis, swine diseases, porcine, PCR, real-time PCR, multiplex PCR

*E-mail: sandor.belak@sva.se; Fax: 0046 18 674 669

Acta Veterinaria Hungarica 53 (1), pp. 125–136 (2005)

CYTOPATHOGENICITY MARKERS IN THE GENOME OF HUNGARIAN CYTOPATHIC ISOLATES OF BOVINE VIRAL DIARRHOEA VIRUS

Á. Bálint¹, Claudia Baule³, S. Kecskeméti², I. Kiss² and S. Belák^3*

1Department of Virology, Central Veterinary Institute, H-1149 Budapest, Tábornok u. 2, Hungary; ²Veterinary Institute of Debrecen, H-4002 Debrecen, P.O. Box 51, Hungary; ³Joint Research and Development Division, Departments of Virology, The National Veterinary Institute and the Swedish University of Agricultural Sciences, Ulls väg 2B, S-751 89 Uppsala, Sweden

(Received March 19, 2004; accepted September 1, 2004)

Since genetic recombination is a major factor in the evolution of the cytopathogenic (cp) bovine viral diarrhoea virus (BVDV) biotypes, in this study the cytopathogenicity markers were investigated in the genomes of two cp BVDV strains recently isolated from mucosal disease (MD) cases in Hungary. In the genome of strain H4956, a Jiv-like insertion was found similar to those described in reference strain NADL and in other BVDV 1, BVDV 2 and border disease virus (BDV) strains. The 133 amino acid Jiv-like sequence is inserted at nucleotide position 4984 (amino acid position 1533), 9 nucleotides upstream of that of strain NADL. The insertion showed 96% amino acid sequence identity with the cellular Jiv protein. In the genome of cp BVDV strain H115/PCR, an ubiquitin-containing duplication was found. The duplicated sequence started at nucleotide position 7978 (amino acid 2531) in the NS4B gene. The duplication contained a complete ubiquitin monomer of 76 amino acids and the complete NS3 gene starting at nucleotide position 5153 (amino acid 1589), which corresponds to the first N-terminal amino acid of NS3. The duplication was located further downstream of the known ubiquitin-containing genomic regions of cp BVDV strains, and it consisted of the shortest inserted nucleotide sequence. The insertions and duplication of strains H4956 and H115/PCR further confirmed that recombinations occurring at positions A and B are the most common mechanisms leading to the development of BVDV cytopathogenicity.

Key words: Bovine viral diarrhoea virus, cytopathogenicity, markers, NS2-3, NS3

*Corresponding author: E-mail: sandor.belak@sva.se

Acta Veterinaria Hungarica 53 (1), pp. 137–146 (2005)

DETECTION OF RESPIRATORY AND ENTERIC SHEDDING OF BOVINE CORONAVIRUSES IN CATTLE IN NORTHWESTERN TURKEY

M. Hasoksuz^1*, A. Kayar², T. Dodurka² and A. Ilgaz¹

1Department of Microbiology and ²Department of Internal Medicine, Veterinary Faculty, Istanbul University, 34320 Avcilar, Istanbul, Turkey

(Received January 15, 2004; accepted May 24, 2004)

Bovine coronavirus (BCoV) is an important cause of diarrhoea in calves, winter dysentery in adult cattle and respiratory tract disease in feedlot cattle. Serum, faecal and nasal swab samples were collected from a total of 96 cattle with clinical signs in 29 barns of 23 villages in Northwestern Turkey. The cattle were subdivided into 3 distinct age groups (0–30 days old, 4–12 months old and 2–7 years old). An indirect antigen-capture ELISA and an antibody-detection ELISA as well as geometric mean BCoV antibody titres were used to detect BoCV shed in the faeces and in the nasal secretions, respectively. Relationships between BCoV shedding and age group, seroconversion and clinical signs in cattle were also analysed. The rate of faecal shedding of BoCV was 37.1% (13/35) in 0–30 days old calves, 25.6% (10/39) in 4–12 months old feedlot cattle and 18.2% (4/22) in 2–7 years old cows. The overall rate of BCoV faecal shedding was 28.1% (27/96) in the cattle examined. Only one animal in the 4–12 months old age group was found to shed BoCV nasally. The analysis showed that there was a significant difference (P < 0.0001) with respect to faecal shedding between the clinical signs and the age groups. BCoV antibody titre in 50% of all cattle was ?100 as detected by ELISA while 27.1% of the cattle had high titres ranging between 1,600 and 25,600. The seroconversion rate was 7.3% (7/96) in animals shedding BoCV in the faeces and 42.7% (41/96) in cattle negative for faecal shedding as detected by ELISA, and 20.8% of cattle with no seroconversion shed BCoV in the faeces. There was no statistically significant association between seroconversion and nasal or faecal BCoV shedding. These findings confirm the presence of BCoV infections in Turkey. Further studies are needed to isolate BCoV strains in Turkey and to investigate their antigenic and genetic properties.

Key words: Bovine coronavirus, ELISA, cattle, calf, Turkey

*Corresponding author: Mustafa Hasoksuz; E-mail: hasoksuz@istanbul.edu.tr; Phone: +90 (212) 473 7070; Fax: +90 (212) 473 7241

Acta Veterinaria Hungarica 53 (1), pp. 147–151 (2005)

THE REPLICATION OF CANINE HERPESVIRUS (CHV) INDUCES APOPTOSIS IN CANINE KIDNEY CELL LINE: SHORT COMMUNICATION

O. Kim^1* and S. J. Yi²

1College of Medicine, Seoul National University, Republic of Korea; ²College of Veterinary Medicine, Kyungpook National University, Republic of Korea

(Received April 28, 2003; accepted September 1, 2004)

The alphaherpesvirus canine herpesvirus (CHV) was tested in order to determine whether or not it has apoptotic potential. We have demonstrated that lytic replication of CHV resulted in induction of apoptosis. This phenomenon was confirmed using different techniques including in situ TUNEL assay and DNA laddering. The apoptotic activity of CHV might influence the pathobiology of this virus.

Key words: Apoptosis, herpesvirus, canine herpesvirus

*Corresponding author: Okjin Kim, Department of Laboratory Animal Sciences, College of Medicine, Seoul National University, 28 Yongon-dong, Chongno-gu, Seoul 110-799, South Korea; Phone: 82 2 740 8077; Fax: 82 2 763 5206; E-mail: kimoj@snu.ac.kr