A 39 kDa protein mediates adhesion of avian Pasteurella multocida to chicken embryo fibroblast cells

To clarify the role of avian Pasteurella multocida capsule in pathogenesis, adhesion of capsulated strains P-1059, X-73 and Pm-18, and noncapsulated strains P-1059B, Pm-1 and Pm-3 to chicken embryo fibroblast (CEF) cells was compared. Number of adherent organisms of the capsulated strains to CEF cells were approximately three times as much as noncapsulated strains indicating that adhesive properties were enhanced by the presence of bacterial capsule. Pretreatments of the bacterial cells with heat, trypsin, or with antiserum caused a marked decrease in adhesion of capsulated strain P-1059 and its noncapsulated variant P-1059B. However, depolymerization of capsular hyaluronic acid with high dose of hyaluronidase enhanced adhesion of these strains. Combined treatments of the bacterial cells with both hyaluronidase and trypsin significantly (P < 0.05) inhibited the adherence of strain P-1059 as compared to the treatment only with trypsin, but strain P-1059B was not affected. SDS-PAGE profiles of crude capsular extract (CCE) prepared from capsulated strain P-1059 and its noncapsulated variant P-1059B grown on dextrose starch agar (DSA) plates by heating at 56 degrees C in a 2.5% NaCl solution demonstrated eight protein bands of 28, 34, 36, 39, 52, 56, 63 and 93 kDa. The 28, 34 and 36 kDa proteins were commonly major for both strains, and the 39 kDa protein was major only for strain P-1059 but poor in strain P-1059B. Outer membrane protein (OMP) profiles were identical with a major protein at 34 kDa and four minor proteins between the two strains. The adhesion of strain P-1059 and strain P-1059B to CEF cells was inhibited significantly (P < 0.01) by treatment with rabbit antisera against P-1059, P-1059B, CCE or 39 kDa protein of strain P-1059 as compared to the treatment with either PBS or with normal rabbit serum. These results indicated that an antigenic 39 kDa protein in the capsule may be responsible for adhesion of avian P. multocida type A strains to CEF cells as a virulence factor.     

Ganglioglioma in the thalamus of a puppy

A solitary brain mass of a 4-month-old miniature dachshund showing seizure-like neurological signs was examined histopathologically. At necropsy a white tumor mass, replacing the thalamus, approximately 1.5 cm in diameter, was found. There was cystic space filled with yellowish pale fluid in the central area of the tumor mass. Histopathological examination revealed that the mass consisted of irregularly arranged well-differentiated neuronal and glial cells, and multifocal mineral deposits. The neuronal cells had a large clear nucleus and various amount of Nissl substances in the cytoplasm. Some neural cells were bi-nucleated. Neither mitotic figures nor proliferating cell nuclear antigen (PCNA)-positive nuclei was found in the neuronal cells. Immunostaining for glial fibrillary acidic protein (GFAP) revealed diffuse proliferation of GFAP-positive glial cells and their processes, while these glial cells did not show apparent cellular atypism, mitotic activity, or PCNA-immunoreactivity. Accordingly, the present tumor was diagnosed as ganglioglioma, and hamartomatous histogenesis might be possible.     

Investigation of the short-term effect of chemonucleolysis with chondroitinase ABC.

Chondroitinase ABC (C-ABC) is expected to be a novel agent for chemonucleolysis. The effect of C-ABC was investigated by magnetic resonance (MR) and radiograph. C-ABC was administered into the lumbar intervertebral disks on the clinically normal beagles (n=5), in a dose of 50 microl (12.5 units as C-ABC). MR scans were performed pre-dose, and 1, 3, 7, 14 and 28 days after administration of C-ABC, and the signal intensity (SI) of the nucleus pulposus was measured. Radiographs were taken pre-dose, and 1, 2, 3, 4, 5, 6, 7, 14 and 28 days post-dose, to evaluate narrowing of the disk space in terms of height index (HI). In addition, the quantity of the chondroitin sulfate (CS) and the hyaluronic acid (HA) in the nucleus pulposus were measured by high performance liquid chromatography on day 28 after dosing. SI and HI continuously decreased, following the injection to 37.1% and 78.9% of the pre-dose values, respectively. Statistically significant differences (p<0.01) were observed between the C-ABC group and the control group in the respects on day 1 post-dose. CS and HA contents of the nucleus pulposus were noted to be significantly decreased on day 28 (p<0.01) in the treated group. This agent proved to degenerate proteoglycans in the nucleus pulposus, thus progressively reducing the interdiskal pressure from day 1 post-dose onwards. It is concluded that C-ABC is expected to afford its efficacy from early in the course of chemonucleolysis.     

Tryptic peptides of canine thyroglobulin reactive with sera of patients with canine hypothyroidism caused by autoimmune thyroiditis

Canine thyroglobulin (cTg) was treated with trypsin at a ratio of trypsin to cTg of 1:100 (w/w). Tryptic peptides of cTg were analysed by Western immunoblotting for their reactivity to serum thyroglobulin autoantibodies (TgAA) from patients with TgAA-positive hypothyroidism and normal individuals. The sera of patients with TgAA-positive hypothyroidism reacted with several peptides: 43, 32.5 and 31 kDa; the sera of normal individuals did not bind these tryptic peptides. Some of the TgAA-positive sera of patients reacted with 25 kDa peptide in addition to three tryptic peptides above. This experiment was the first report about antigenic epitopes of cTg. These small tryptic peptides recognized by TgAA may be related with the induction of TgAA and may be useful as markers for autoimmune thyroid diseases in dog.     

Solitary nonepitheliotropic T-cell lymphoma in a dog

A 7-year-old male Golden Retriever with swelling of the rostral bridge and right wing of the nasal areas, sneezing, and inspiratory difficulty was referred to a neighbor veterinarian. Except for those in the nasal area, no lesions were noted during routine physical examination. The mass occupying the nasal cavity was not observed radiographically. Punch biopsy of the affected lesions revealed nonepitheliotropic lymphoma. Immunohistochemical staining for CD3 antigen was positive. The dog was diagnosed with solitary nonepitheliotropic T-cell lymphoma. Local radiotherapy and systemic chemotherapy with doxorubicin were instituted and resulted in total clinical remission. The dog has remained disease free for 30 months.     

In planta dynamic analysis of onion yellows phytoplasma using localized inoculation by insect transmission

ABSTRACT Due to the lack of a means to inoculate plants mechanically, the histological dynamics and in planta spread of phytoplasmas have been studied very little. We analyzed the dynamics of plant infection by phytoplasmas, using a technique to infect a limited area of a leaf, nested polymerase chain reaction (PCR), real-time PCR, and immunohistochemical visualization. Following localized inoculation of a leaf of garland chrysanthemum (Chrysanthemum coronarium) by the vector leafhopper Macrosteles striifrons, the onion yellows (OY) phytoplasma spread within the plant from the inoculated leaf to the main stem (1 day postinoculation [dpi]), to the roots and the top leaf (2 dpi), and to other leaves from top to bottom (from 7 to 21 dpi). The populations of the OY phytoplasmas in inoculated leaves and roots increased approximately sixfold each week from 14 to 28 dpi. At 14 dpi, the OY phytoplasmas colonized limited regions of the phloem tissue in both the root and stem and then spread throughout the phloem by 21 dpi. This information should form the basis for elucidating the mechanisms of phytoplasma multiplication and migration within a plant host.     

Insemination of recipient sows improves the survival to term of vitrified and warmed porcine expanded blastocysts transferred non‐surgically

This study was performed to evaluate reproductive performance after non‐surgical embryo transfer (Ns‐ET) of 10–15 porcine expanded blastocysts (ExBs) that had been vitrified and warmed (V/W) using the micro volume air cooling (MVAC) method. The effect of asynchrony between the donor and recipient estrous cycle was investigated. Ns‐ET was conducted in recipients whose estrous cycle was asynchronous to that of donors by a delay of 2, 1, or 0 days. In the 2‐day and 1‐day groups, the similar farrowing rates (27.3% and 25.0%) and survival rates to term (13.9% and 15.7%) were obtained after Ns‐ET of V/W ExBs. None of the recipients in 0‐day group farrowed. Artificial insemination (AI) prior to Ns‐ET was then evaluated. Ten–15 V/W ExBs were transferred non‐surgically to 12 recipients whose estrous cycles were asynchronous to that of donors by a 2‐day delay. All of the recipients produced piglets, and all (100.0%) delivered piglets were derived from the transferred V/W ExBs. The survival rate of V/W ExBs to term was 25.2%. These results demonstrate that Ns‐ET of V/W ExBs using MVAC can facilitate piglet production, even if 10–15 embryos are transferred. Moreover, piglets were obtained stably when AI was performed prior to Ns‐ET.     

The effect of artificial insemination prior to transfer of a limited number of vitrified and warmed porcine embryos by open pulled straw (OPS) method on their survival ability for farrowing

Embryo transfer (ET) of 20 porcine expanded blastocysts (ExBs) vitrified and warmed (VW) by open pulled straw (OPS) to a recipient allows stable piglet production. The efficiency of artificial insemination (AI) prior to ET of 10 VW ExBs for piglet production was investigated. For one trial, 10–15 VW ExBs from single donor were assigned, 10 were used for ET and the remains were assessed for their in vitro viability. In the non‐AI/ET group, 10 were transferred to each of five recipients. As AI/ET group, 10 were transferred to each of five recipients after AI. In AI/non‐ET group, only AI was performed to seven gilts. In the non‐AI/ET group, the pregnancy rate was 40%, but none of them farrowed. In the AI/ET group, all recipients produced piglets. Four (80.0%) delivered piglets from transferred VW ExBs. The survival rate of VW ExBs to term was 20.0% (10/50). In the AI/non‐ET group, six of the seven gilts farrowed. There was no difference in in vitro viability between the non‐AI/ET and AI/ ET groups (62.5% and 68.3%, respectively). AI prior to ET can be an appropriate way to maintain pregnancy and assist the development of a low number of VW ExBs to term.