Purification and embryotropic roles of tissue inhibitor of metalloproteinase-1 in development of "HanWoo" (Bos taurus coreanae) oocytes co-cultured with bovine oviduct epithelial cells

This study was conducted to purify a tissue inhibitor of metalloproteinase (TIMP)-1 in a serum-free medium conditioned with bovine oviduct epithelial cells (BOEC) and to evaluate its effect on development of "HanWoo" (Bos taurus coreanae) embryos to the blastocyst stage. In the first study using SDS-PAGE electrophoresis, the presence of 32 kDa proteins, which contains TIMP-1, was detected in the medium conditioned with BOEC, and TIMP-1 was then purified from the medium by gel filtration and HPLC techniques. When examined TIMP-1 secretion, fluorescent foci indicating the secretion of TIMP-1 were found after stained BOEC with fluorescein isothiocyanate. In the next experiment, two-cell embryos derived from in vitro-fertilization were cultured in a serum-free medium, to which 0, 1.25, 2.5 or 5 microg/ml of purified TIMP-1 was supplemented. More (P<0.05) embryos developed to the morula and blastocyst stages after the addition of 2.5 microg/ml to culture medium than after no addition. In conclusion, our data indicate that BOEC secrete TIMP-1 and this glycoprotein promotes the prehatched development of "HanWoo" embryos derived from in vitro-fertilization.     

A rapid method for detecting Trichinella spiralis larvae in pork using a monoclonal antibody-latex conjugate

A simple, non-microscopic method was developed for the direct detection of Trichinella spiralis in pork. Samples of meat were treated with pepsin for 2 h to liberate the larvae, and sonicates were made from the deposit and used in an antigen detection system. In this system, latex particles, sensitized with a parasite-specific monoclonal antibody, were utilized in a slide or tube agglutination test. The efficiencies of these immunological methods were compared with that of trichinelloscopy in the examination of 102 pork specimens, 35 of which contained varying numbers of larvae after enzymatic digestion of 2 g of the meat. Trichinelloscopy detected 77.1% of these positive samples, but this was lower (55.5 and 41.7%) for those samples with a low larval burden (less than 50 or less than 10 larvae g-1 tissue, respectively). Better results (88.6, 76.5 and 60.0%) were obtained with the slide latex agglutination technique. The efficiency of detection was further improved (100%) using the more sensitive tube agglutination method. However, this was based on a small number (20) of samples examined and the limit of detection of this technique was subsequently found to be three larvae in total. No false-positives were seen with the latex agglutination methods from a total of 67 uninfected specimens examined.     

Mucosal vaccination against influenza: protection of pigs immunized with inactivated virus and ether-split vaccine

Effective vaccinations against swine influenza reduce the economic loss of pig industries, and also may minimize the possibility of emergence of new pandemic viruses, since pigs are intermediate hosts to generate reassortant viruses among avian and mammalian influenza viruses. In this study, we showed that intranasal immunization of pigs with formalin-inactivated or ether-split influenza vaccine (A/Aichi/2/68) induced virus-specific IgG, IgM, and IgA antibodies in their nasal secretions and sera, resulting in complete protection from virus challenge. Antibody response to the challenge virus was not observed in the immunized pigs, suggesting that the replication of the virus in the primary targets, respiratory epithelial cells, was inhibited. The present results indicate that intranasal immunization of pigs with inactivated vaccines is effective to control swine influenza, and also provide a good model, as well as a mouse model, to evaluate an intranasal application of influenza vaccine for humans.     

Comparative pharmacokinetics of tylosin or florfenicol after a single intramuscular administration at two different doses of tylosin-florfenicol combination in pigs

Clinical pharmacokinetic profiles were investigated following intramuscular (i.m.) administration to pigs with a commercial tylosin-florfenicol combination product at a dose of 2.5 mg/kg tylosin and 5 mg/kg florfenicol or 10 mg/kg tylosin and 20 mg/kg florfenicol. The quantitation limit (QL) of florfenicol was 0.1 microg/ml, the inter-day and intra-day precision (CV%) were both beow 10%. The quantitation limit (QL) of tylosin was 0.05 microg/mL. The pharmacokinetic characteristics after i.m. doses were fitted by a one compartment open model. A fourfold decrease in the normal dose of each drug (20 mg/kg to 5 mg/kg for florfenicol, and 10 mg/kg to 2.5 mg/kg for tylosin) resulted in a corresponding two fold decrease in each drug of the maximum plasma concentration (C(max)) and the area under curve (AUC) values.     

Potato (Solanum tuberosum L. cv. Gogu valley) protein as a novel antimicrobial agent in weanling pigs

A total of 280 weaned pigs (Landrace x Yorkshire x Duroc) were used in a 28-d growth study to investigate the effect of feeding different levels of potato proteins on growth performance, nutrient digestibility, immune response, small intestinal morphology, and bacterial populations in feces and large intestine. Pigs (initially 6.42 +/- 0.74 kg of BW and 23 +/- 3 d of age) were randomly allotted to 5 treatments on the basis of BW, each treatment composed of 4 pens, each pen having 14 pigs. Dietary treatments included positive control (PC; basal diet + 150 mg/kg apramycin and 10 mg/ kg colistin sulfate); and potato protein (PP), consisting of the basal diet with 0, 0.25, 0.50, or 0.75% of potato protein. Diets were fed in 2 phases: phase I (d 0 to 14 postweaning) and phase 2 (d 14 to 28 postweaning). Potato protein was extracted from a value-added type of the new potato variety, Solanum tuberosum L. cv. Gogu valley, and was shown to have a minimum inhibitory concentration of 300 to 500 mug/mL. Performance of PC was compared with 0.25 to 0.75% PP, whereas linear and quadratic trends of increasing PP (0 to 0.75% PP) were tested. Over the 28-d trial, pigs fed the PC diets showed improved overall ADG (P < 0.05) and G:F (P = 0.090) compared with pigs fed PP, whereas increasing levels of PP linearly improved ADG (P < 0.05), ADFI (P = 0.052), and G:F (P = 0.098). The digestibility of DM and CP in both the phases was greater in PC than PP, and feeding of PP linearly improved the DM digestibility (P < 0.05) in phase II. The bacterial populations in the feces of pigs fed PC and PP were comparable, except for total bacteria and coliform bacteria in the feces at d 14 and 28, which were decreased in PC; and feeding of PP was effective in linearly reducing the populations of microbes in feces and contents of cecum, colon, and rectum. There was linear increase (P < 0.10) in skin-fold thickness in response to phytohemagglutinin with an increase in PP levels. Haemagglutinin titers on d 21 were greater (P = 0.054) in PC, and at d 28 the haemagglutinin titers were quadratically affected in pigs fed PP (P = 0.070). There was a trend toward a decrease in crypt depth (P = 0.068) and a greater villus height:crypt depth ratio (P = 0.082) of ileum in PC compared with PP. These results suggest that PP may be an alternative to medicated feed with antibiotics because it showed antimicrobial activity by effectively reducing the population of coliform bacteria and also improved the performance of weanling pigs.     

Bronchoalveolar lavage affects computed tomographic and radiographic characteristics of the lungs in healthy dogs

Bronchoalveolar lavage is a common diagnostic test for dogs with suspected pulmonary disease, however there is no published information on whether this procedure could affect the imaging characteristics of the lungs. Aims of this prospective experimental study were to describe computed tomography (CT) and radiographic features of the lungs after bronchoalveolar lavage in a sample of healthy dogs. Thoracic CT and radiographic images of eight healthy Beagles were acquired at the following time points: before bronchoalveolar lavage, immediately following bronchoalveolar lavage, and at 2, 4, 8, 12, and 24 h following bronchoalveolar lavage. Lung consolidation or interstitial patterns were seen in CT and radiographic images immediately after the procedure. Radiographic lung patterns resolved within 2 h and CT patterns resolved within 24 h. Resolution of the CT pulmonary patterns in the ventral areas of the lungs was delayed compared to the dorsal areas. Mean CT imaging scores differed over time (P < 0.001), while mean radiographic imaging scores did not differ over time. This study suggests that thoracic radiography and CT imaging assessments should precede bronchoalveolar lavage procedures if possible, or be performed at least 24 h afterward.     

Prevalence of the AMHR2 mutation in Miniature Schnauzers and genetic investigation of a Belgian Malinois with persistent Müllerian duct syndrome

Persistent Müllerian duct syndrome (PMDS) is a sex‐limited disorder in which males develop portions of the female reproductive tract. Important consequences of PMDS are cryptorchidism and its sequelae of infertility and increased risk of testicular cancer. Anti‐Müllerian hormone (AMH) and its receptor (AMHR2) induce the regression of the Müllerian ducts in male embryos. In Miniature Schnauzer dogs, the genetic basis has been identified as an autosomal recessive nonsense mutation in AMHR2, but the allele frequency of the mutation is unknown. Thus, the primary objective of this study was to estimate the prevalence of the AMHR2 mutation in North American Miniature Schnauzers, in order to ascertain the value of genetic testing in this breed. An additional objective was to determine whether mutations in AMH or AMHR2 were responsible for PMDS in a Belgian Malinois; this would aid development of a genetic test for the Belgian Malinois breed. Genomic DNA from 216 Miniature Schnauzers (including one known PMDS case) was genotyped for the AMHR2 mutation, and DNA from a single PMDS‐affected Belgian Malinois was sequenced for all coding exons of AMH and AMHR2. The Miniature Schnauzer cohort had an AMHR2 mutation allele frequency of 0.16 and a carrier genotypic frequency of 0.27. The genetic basis for PMDS in the Belgian Malinois was not determined, as no coding or splicing mutations were identified in either AMH or AMHR2. These findings support a benefit to AMHR2 mutation testing Miniature Schnauzers used for breeding or with cryptorchidism.