Expression of claudins in the normal canine gastric mucosa

The aim of the present study was to investigate the expression pattern of claudin-1, -2, -3, -4, -5, -7, -8, -10 and -18 in the intact fundic and pyloric gastric mucosa of dogs. Intense, linear, membranous claudin-18 positivity was detected in the surface gastric cells and in the epithelial cells of the gastric glands both in the fundic and pyloric stomach regions. The mucous neck cells in the apical part of the glands, furthermore the parietal cells and chief cells of the basal part of the gland were all positive for claudin-18, in the same way as the enteroendocrine cells. Cells of the basal part of the pyloric glands showed intense, linear, membranous claudin-2 positivity, but cells of the superficial portion of these glands and the surface gastric cells in this region were claudin-2 negative. Fibroblasts, endothelial cells, lymphocytes of the propria layer, smooth muscle cells and vegetative neurons were all negative for claudin-2. All gastric epithelial cells were negative for claudin-1, -3, -4, -5, -6, -7, -8 and -10. The endothelial cells of the propria layer had intense claudin-5 positivity. We assume that claudin-18 forms a paracellular barrier against gastric acid in the healthy canine stomach, in the same way as in mice

Different sample types in pigs challenged with Haemophilus parasuis following two treatment schemes with tulathromycin

This study aimed to test the efficacy of samplings for the detection of Haemophilus parasuis after metaphylactic treatment and subsequent challenge using an established model for Glässer’s disease. In this model, 36 piglets were equally assigned to a negative control, a positive control, and two trial groups receiving tulathromycin 7 or 4 days prior to challenge. The piglets of three groups were challenged intratracheally with H. parasuis serovar 5. As a result, four pigs in each challenged group died or had to be euthanised within 10 days post challenge. The remaining 15 pigs of these challenged groups survived until termination of the experiment (days 14–15). All pigs were necropsied and collective swabs of serosal surfaces were tested by bacterial culture and PCR. Samples of tarsal synovial fluid and joint capsule, cerebrospinal fluid (CSF) and brain swabs were tested by PCR. A total of 22 out of the 27 challenged animals had macroscopically detectable polyserositis and all of them tested positive in the collective swab samples. Haemophilus parasuis was more frequently detected in pigs that died within the first 10 days compared to those surviving until days 14–15 (P < 0.001), and those that succumbed within 10 days showed higher positivity rates in the brain and CSF. All pigs which were positive in the CSF had detectable meningitis. At days 14–15, joint samples from 5 of the remaining 15 pigs tested positive for H. parasuis. Four of these five animals did not show any macroscopic or histological lesions in the joints. In conclusion, collective swabs were the best sample material in acute cases, whereas samples from the joints gave the best results in chronic cases. In this challenge model it was not possible to prove the metaphylactic effect of tulathromycin administered 4 and 7 days prior to infection with H. parasuis

Arterivírusok vizsgálata, különös tekintettel a hazai izolátumok genetikai tulajdonságaira = Investigation of arteriviruses with special regard to the genomic characteristics of local isolates

Az elmúlt négy évben vizsgáltuk a két állategészségügyi szempontból fontos arterivírus, a sertések szaporodásbiológiai zavarokkal és légzőszervi tünetekkel járó tünetegyüttesét előidéző vírus (PRRSV) és a lovak fertőző arteritisét előidéző vírus (EAV) hazai elterjedtségét és megállapítottuk, hogy mindkét vírus hazai előfordulása elmarad a nyugat-európai országokban megfigyelttől. Mindkét vírusfertőzés diagnosztikájában kidolgoztuk és széleskörűen alkalmaztuk az RT-PCR alapú diagnosztikai módszert. Az amplifikációs termékek nukleotid-sorrendjének meghatározása után vizsgáltuk a két vírus filogenetikai viszonyait magyarországi és a környező országokból származó pozitív minták felhasználásával, továbbá összehasonlítottuk ezeket a nemzetközi génbankban elhelyezett szekvenciákkal is. A PriProET és Taqman technikára valamint valós idejű RT-PCR eljárásra alapozott módszerekkel a vírusfertőzések in vitro és in vivo zajló kinetikájáról tudtunk adatokat gyűjteni. Eredményeinket nyolc referált szakfolyóiratben megjelent vagy közlésre elfogadott közleményben és két PhD értekezésben közöltük illetve foglaltuk össze. | In the four years of the study period the occurrence of the two arteriviruses of veterinary significance, the porcine reproduction and respiratory syndrome virus (PRRSV) and the equine arteritis virus has been investigated, and it was demonstrated, that the presence of both viruses is lower then in the Western European countries. An RT-PCR based diagnostic method has been developed and routinely applied in the diagnosis of both viral infections. After sequencing the amplicons the phylogenetic relationship of the viruses was investigated using positive samples collected in Hungary and in the neighbouring countries, and these were compared to sequences deposited in the GenBank. Data were collected on the in vitro and in vivo kinetics of the virus multiplication using methods based on PriProET and Taqman technique and real time RT-PCR methods. The results were published and summarised in eight articles published in peer reviewed periodicals and in two PhD theses

COMPLETE GENOME SEQUENCE OF PORCINE CIRCOVIRUS TYPE 2 UKRAINIAN ISOLATES

Porcine circovirus type 2 (PCV2) is associated with distinct syndromes and diseases in swine, collectively known as porcine circovirus-associated diseases (PCVAD), which include postweaning multisystemic wasting syndrome (PMWS), PCV2-associated pneumonia as a part of the porcine respiratory disease complex (PRDC), PCV2-associated enteritis, PCV2-associated reproductive failure, and porcine dermatitis and nephropathy syndrome (PDNS) (1–3). PCV2-infection is widespread and essentially all pig herds are infected with PCV2. Porcine circovirus 2 (PCV2), a member of the genus Circovirus in the family Circoviridae, is a very small single-stranded negative-sense DNA virus of approximately 1.7 kb (4). The genome of PCV2 encodes three major open reading frames (ORFs) encoding the replicase proteins (ORF1), the viral capsid protein (ORF2), and a protein with suggested apoptotic activity (ORF3) (5). Previous reports showed that there were five PCV2 genotypes, including PCV2a, PCV2b, PCV2c, PCV2d, and PCV2e (6– 9). Here, we report the complete genome sequence of Ukrainian PCV2 isolates from different regions of Ukraine

Immunohistochemical characterization of type II pneumocyte proliferation after PRRSV (Type I) challenge

The study aimed to histologically and immunohistochemically characterize lung lesions after a challenge with a recently isolated PRRSV field strain in growing pigs 10 and 21 days post infection (DPI). In the first phase of the study lung lesions were evaluated microscopically on routine haematoxylin and eosin stained slides. The evaluation was performed as a blinded analysis and the lesions were scored based on the following criteria: (1) pneumocyte hypertrophy and hyperplasia, (2) septal mononuclear infiltration, (3) intraalveolar necrotic debris, (4) intraalveolar inflammatory cell accumulation and (5) perivascular inflammatory cell accumulation. For further characterization of the lung lesions, immunohistochemical stainings were performed using anti-cytokeratin, anti-Ki67, anti-TTF-1 (Thyroid Transcription Factor-1), anti-myeloid receptor (MAC387), and anti-PRRSV antibodies to identify alveolar epithelial cells, proliferating cells, type II pneumocytes, macrophages, and PRRSV antigen, respectively. The evaluation of the immunohistochemical stainings revealed that humanized anti TTF-1 antibodies can successfully identify type II pneumocytes in porcine lung tissue. Marked proliferation of these cells was confirmed by a significant (p<0.05) increase of TTF-1 positive cells in acute cases compared to the lungs of control pigs. Cytokeratin labeling marked the type I, and type II pneumocytes as well as bronchial epithelial cells, however this staining was not suitable for cell counting purposes. When the routine histological scores were compared to the number of immunohistochemically positive cells, Ki67 cell counts were found to show positive correlation (p<0.05) with the overall severity of the lesions

Full genome sequence analysis of a wild, non-MLV-related type 2 Hungarian PRRSV variant isolated in Europe

Porcine reproductive and respiratory syndrome virus (PRRSV) is a widespread pathogen of pigs causing significant economic losses to the swine industry. The expanding diversity of PRRSV strains makes the diagnosis, control and eradication of the disease more and more difficult. In the present study, the authors report the full genome sequencing of a Type 2 PRRSV strain isolated from piglet carcasses in Hungary. Next generation sequencing was used to determine the complete genome sequence of the isolate (PRRSV-2/Hungary/102/2012). Recombination analysis performed with the available full-length genome sequences showed no evidence of such event with other known PRRSV. Unique deletions and an insertion were found in the nsp2 region of PRRSV-2/Hungary/102/2012 when it was compared to the highly virulent VR2332 and JXA-1 prototype strains. A majority of amino acid alterations in GP4 and GP5 of the virus were in the known antigenic regions suggesting an important role for immunological pressure in PRRSV-2/Hungary/102/2012 evolution. Phylogenetic analysis revealed that it belongs to lineage 1 or 2 of Type 2 PRRSV. Considering the lack of related PRRSV in Europe, except for a partial sequence from Slovakia, the ancestor of PRRSV-2/Hungary/102/2012 was most probably transported from North-America. It is the first documented type 2 PRRSV isolated in Europe that is not related to the Ingelvac MLV

Characterization and therapeutic application of canine adipose mesenchymal stem cells to treat elbow osteoarthritis

Visceral adipose tissue (AT) obtained from surgical waste during routine ovariectomies was used as a source for isolating canine mesenchymal stem cells (MSCs). As determined by cytofluorimetry, passage 2 cells expressed MSC markers CD44 and CD90 and were negative for lineage-specific markers CD34 and CD45. The cells differentiated toward osteogenic, adipogenic, and chondrogenic directions. With therapeutic aims, 30 dogs (39 joints) suffering from elbow dysplasia (ED) and osteoarthritis (OA) were intra-articularly transplanted with allogeneic MSCs suspended in 0.5% hyaluronic acid (HA). A highly significant improvement was achieved without any medication as demonstrated by the degree of lameness during the follow-up period of 1 y. Control arthroscopy of 1 transplanted dog indicated that the cartilage had regenerated. Histological analysis of the cartilage biopsy confirmed that the regenerated cartilage was of hyaline type. These results demonstrate that transplantation of allogeneic adipose tissue-derived mesenchymal stem cells (AT-MSCs) is a novel, noninvasive, and highly effective therapeutic tool in treating canine elbow dysplasia. © 2017, Canadian Veterinary Medical Association. All rights reserved

Full-length genome sequence analysis of a Hungarian Porcine Reproductive and Respiratory Syndrome Virus isolated from severe respiratory disease

The authors report the isolation of a Type 1 PRRSV strain from a clinical outbreak of severe respiratory problems and high fever. Next generation sequencing was used to determine the complete genome sequence of the isolate (9625/2012). The virus belongs to a new branch within subtype 1, clade D, containing mostly Spanish sequences and shows highest similarity to PRRSV Olot/1991 and to the Amervac vaccine strain. SimPlot analysis performed with the available full-length genome sequences indicates no evidences of recombination. Mutation analysis of 9625/2012 and the most related strains revealed high proportion of amino acid substitutions in the putative neutralizing epitopes, suggesting an important role of the selective immune pressure in the evolution of PRRSV 9625/2012