Mechanical transmission of porcine reproductive and respiratory syndrome virus throughout a coordinated sequence of events during cold weather

Using a field-based model, mechanical transmission of porcine reproductive and respiratory syndrome virus (PRRSV) was assessed throughout a coordinated sequence of events that replicated common farm worker behavior during cold weather (< 0°C). The model involved fomites (boots and containers), vehicle sanitation, transport, and the movement of personnel. A field strain of PRRSV was inoculated into carriers consisting of snow and water, and carriers were adhered to the undercarriage of a vehicle. The vehicle was driven approximately 50 km to a commercial truck washing facility where the driver's boots contacted the carriers during washing, introducing the virus to the vehicle interior. The vehicle was then driven 50 km to a simulated farm site, and the driver's boots mechanically spread virus into the farm anteroom. Types of containers frequently employed in swine farms (styrofoam semen cooler, metal toolbox, plastic lunch pail, and cardboard animal health product shipping parcel) contacted drippings from footwear on the anteroom floor. The truck wash floor, vehicle cab floor mats, boot soles, anteroom floor, and the ventral surface of containers were sampled to track the virus throughout the model. Ten replicates were conducted, along with sham-inoculated controls. At multiple sampling points PRRSV nucleic acid was detected in 8 of 10 replicates. In each of the 8 PCR-positive replicates, infectious PRRSV was detected on the surfaces of containers by virus isolation or swine bioassay. All sham-inoculated controls were negative. These results indicate that mechanical transmission of PRRSV can occur during coordinated sequence of events in cold weather.     

Isolation and characterization of porcine circovirus type-2 from sera of stillborn fetuses

In order to examine an association between porcine circovirus type-2 (PCV2) infection and reproductive failure in pigs, sera (n = 171) from stillborn fetuses were collected from 3 different farms with prolonged histories of reproductive problems. These sera were tested for the presence of antibodies to PCV2 using an immunoperoxidase monolayer assay. Of the 171 sera tested, 28 had PCV2 antibody titers of ≥ 1:16. When these 28 samples were tested by a polymerase chain reaction assay, 13 were found to contain PCV2 viral DNA. Of these 13 samples containing both PCV2 antibodies and viral DNA, 9 yielded PCV2 on virus isolation. Amino acid sequences comprising open reading frame 2 of PCV2 from 2 of these isolates were compared to PCV2 isolates from cases of post-weaning multi-systemic wasting syndrome (PMWS). The amino acid sequences of the 2 isolates from stillborn pigs were shown to be nearly identical to each other, as well as to other PCV2 isolates associated with reproductive failure. When compared with PMWS isolates, the isolates from the stillborn fetuses showed differences of at least 2 amino acids. These results confirm previous findings that transplacental infection of PCV2 occurs in the field and that stillbirths in pigs may be associated with PCV2 infections. At present, the significance of minor differences in amino acid sequences is not known.     

Possible role of plasma neurotensin in regulating the excitatory neural control of the rectum of the fowl

Effects of neurotensin (NT) applied via the blood vessel on the responses to stimulation of Remak's nerve (RNS) were investigated in the chicken isolated and perfused rectums. NT (5 ng-2 micrograms/ml) produced a concentration-dependent inhibition of the constituent contraction but not relaxation of the responses to RNS. In addition, high concentrations of NT (over 80 ng/ml) produced a contraction of the rectal muscle. Propranolol, a beta-adrenoceptor blocking agent, and guanethidine, an adrenergic neurone blocking agent, were able to reduce the inhibitory effect of NT on the response to RNS while potentiating the contractile effect of NT on the rectal muscle. NT (0.1 and 1 microgram/ml), like norepinephrine, decreased the flow rate of perfusate from the isolated rectum which was perfused at a constant pressure. Guanethidine enhanced norepinephrine-induced vasoconstriction, and phentolamine, an alpha-adrenoceptor blocking agent, plus propranolol was able to abolish it. Either of these prior applications resulted in a small but significant reduction of NT-induced vasoconstriction. These findings suggest that NT in plasma may function as a circulating hormone to exhibit an inhibitory action on the excitatory neural input to the rectum in the chicken, and that catecholamine release from adrenergic nerve terminals by NT may account for some but not all of the activity.     

Effects of partial removal of cytoplasmic lipid on survival of vitrified germinal vesicle stage pig oocytes

This study was designed to investigate whether the partial removal of cytoplasmic lipid from immature pig oocytes prior to vitrification had any positive effects on subsequent maturation, fertilization and early development. Oocytes at the germinal vesicle stage were partially freed from cumulus cells and centrifuged, and then polarized cytoplasmic lipid was removed by micromanipulation. When cultured for 44-48 h, significantly fewer of the centrifuged oocytes reached metaphase II (M-II) than did the non-centrifuged oocytes (approximately 53% vs approximately 68%, respectively); however, no further reduction in the M-II rate was observed when centrifuged oocytes were then delipated prior to culture (approximately 47%). To evaluate their sensitivity to the equilibration and vitrification solutions containing ethylene glycol, non-centrifuged, centrifuged, and delipated oocytes were cultured continuously for several minutes in those solutions, then washed and cultured further; no significant differences in the M-II rates (approximately 20-27%) were observed among the three treatment groups. When oocytes were vitrified and then warmed, significantly more delipated oocytes reached M-II in culture (approximately 15%) than did the non-delipated oocytes, whether centrifuged or not (approximately 4% in each group). When delipated, vitrified and matured oocytes were microsurgically injected with frozen-thawed spermatozoa, approximately 39% were activated and male pronucleus formation was observed in approximately 40% of activated oocytes; none developed beyond the 4-cell stage. These results show that maturation in vitro of vitrified pig oocytes can be promoted by partial removal of cytoplasmic lipid prior to vitrification and that the vitrified oocytes can be fertilized, although the embryonic development obtained in this study was limited.     

Cutaneous lymphoma in a juvenile dog

Abstract: An 18-month-old male Doberman Pinscher was referred to the Veterinary Medical Teaching Hospital of the College of Veterinary Medicine for an erythemic nodular mass on the right forelimb. The mass was diagnosed as cutaneous lymphoma, based on cytologic examination of a mass aspirate and histopathology. Using immunohistochemistry the neoplastic cells were positive for CD3 but negative for CD79a, E-cadherin, and pancytokeratin, confirming their origin as T lymphocytes. No tumor recurrence was noted 18 months after surgery. To our knowledge, this is the first report of a solitary nodular form of cutaneous lymphoma in a young dog.     

An outbreak of necrotic enteritis in the ostrich farm in Korea

An acute disease with high mortality occurred in the ostrich farm and characterized by depression, severe diarrhea and sternal recumbency. Four dead ostriches of the farm were submitted to the National Veterinary Research & Quarantine Service, and diagnosed as necrotic enteritis. In the gross and histopathological examination, extensive diffuse fibrinonecrotic enteritis was found in the small intestine, especially jejunum. Clostridium perfringens was isolated from a pure culture from the duodenum and jejunum of these birds. Based on our current knowledge, this is the first report of an outbreak of necrotic enteritis in the ostrich in Korea.     

Isolation and characterization of a new compound from Prunus mume fruit that inhibits cancer cells

An active compound that inhibits cancer cells was isolated from the fruit of Prunus mume, and its structure and in vitro activities were characterized. The n-hexane fraction obtained from methanol extracts exhibited the strongest inhibitory effect on the growth of cancer cells. From the n-hexane fraction, a new compound named B-1 was purified through preparative thin-layer chromatography, ODS column chromatography, and reverse phase high-performance liquid chromatography and its structure was analyzed by fast atom bombardment mass spectrometry and 1H and 13C NMR. The molecular formula of B-1 was C19H22O6 {2-hydroxy-1-[(7-hydroxy-2-oxo-2H-chromen-6-yl)methyl]-2-methylpropyl-(2Z)-3-methyl-but-e-enoate:prunate}, and the IC50 value was in the range of 39-58 microg/mL in descending order of the cancer cell lines Hep-2, SW-156, HEC-1-B, and SK-OV-3. B-1 exhibited 81-96% inhibition at a concentration level of 100 microg/mL against all cells, based on an 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide assay. However, B-1 showed little effect against normal cells with only 23% or less growth inhibition at 100 microg/mL. Thus, B-1 has a highly specific inhibitory effect against cancer cells but little effect against normal cells. When the cancer cell lines Hep-2 and SK-OV-3 were incubated with B-1 for 72 h, most of the tested cells suffered strong growth inhibition. The compound has the potential to be developed as a nutraceutical.     

Production of blastocysts after intergeneric nuclear transfer of goral (Naemorhedus goral) somatic cells into bovine oocytes

Interspecies cloning may be a useful method to help conserve endangered species and to study nuclear-cytoplasm interaction. The present study investigated in vitro development of goral (Naemorhedus goral) intergeneric nuclear transfer embryos produced by fusing goral fibroblasts with enucleated metaphase II (MII) bovine oocytes. After two to five passages, serum-starved or non-starved goral skin fibroblast cells were transferred into enucleated MII bovine oocytes. Couplets were electrically fused and chemically activated, and then cultured in either modified synthetic oviduct fluid (mSOF) or tissue culture medium-199 (TCM-199) supplemented with 10% FBS. Serum starvation of donor cells did not affect the fusion rate and or development to of cells to the two-cell stage, to more than 9-cells, or to morulae, regardless of culture medium. Three blastocysts from 202 fused embryos were obtained when embryos reconstructed with non- serum- starved donor cells were cultured in mSOF. However, no blastocysts were obtained when the embryos reconstructed with serum-starved donor cells were cultured in mSOF. The total cell number of goral intergeneric embryos averaged 130.3 (range 105-180). In conclusion, this study demonstrated that bovine oocytes can support blastocyst development after intergeneric SCNT with goral fibroblasts.     

A PCR-based assay for discriminating Cervus and Rangifer (Cervidae) antlers with mitochondrial DNA polymorphisms

This study describes a method for discriminating Rangifer antlers from true Cervus antlers using agarose gel electrophoresis, capillary electrophoresis, quantitative real-time PCR, and allelic discrimination. Specific primers labeled with fluorescent tags were designed to amplify fragments from the mitochondrial D-loop genes for various Cervus subspecies and Rangifer tarandus differentially. A 466-bp fragment that was observed for both Cervus and Rangifer antlers served as a positive control, while a 270-bp fragment was specifically amplified only from Rangifer antlers. Allelic discrimination was used to differentiate between Cervus and Rangifer antlers, based on the amplification of specific alleles for both types of antlers. These PCR-based assays can be used for forensic and quantitative analyses of Cervus and Rangifer antlers in a single step, without having to obtain any sequence information. In addition, multiple PCR-based assays are more accurate and reproducible than a single assay for species-specific analysis and are especially useful in this study for the identification of original Cervus deer products from fraudulent Rangifer antlers.     

Antiviral activity of Alpinia katsumadai extracts against rotaviruses

In vitro anti-rotavirus activity of Alpinia katsumadai (AK) extracts were evaluated against bovine G8P[7] and porcine G5P[7] rotaviruses in two different assay strategies, a mixed treatment assay and a post treatment assay. In the mixed treatment assay, six AK extracts [AK-1 (EtOH extract), AK-3 (H(2)O layer), AK-5 (40% methanol fraction), and AK-9-11 (H(2)O extract, polysaccharide fraction, supernatant fraction)] exhibited inhibitory activities against G5P[7] rotavirus with the EC(50) values ranging from 0.7±0.4 to 33.7±6.5 μg/mL. Extracts AK-1, AK-3, and AK-5 inhibited rotavirus infection against G8P[7] rotavirus, the with EC(50) values of 8.4±2.2 μg/mL, 6.5±0.8 μg/mL and 8.4±5.0 μg/mL, respectively. By hemagglutination inhibition (HI) assay, six AK extracts completely inhibited viral adsorption onto human RBCs in both strains of rotaviruses at less than 11 μg/mL. However, in the post treatment assay, there was no anti activity shown against both strains of rotaviruses. As a result, six AK extracts were attributed mainly to having a strong interaction with hemagglutinin protein on the outer surface of rotavirus, resulting to blockage of viral adsorption.