Morphological changes of epiphyseal plate in the long bone of chondrodysplastic dwarfism in Japanese brown cattle.

A total of 20 cases of disproportionate calves on Japanese Brown cattle were examined morphologically. Investigation of 5 affected calves revealed that the insufficiency of endochondral ossification was confined only to the long bones of the limbs and was not accompanied by other associated changes. On the basis of the histological changes of epiphyseal plate and the affected site, this condition may be called chondrodysplasia of the rhizomelic type. Therefore, this disorder was defined as a bovine dwarfism. From three dimensional image analysis and histological changes of the tibial proximal portion in the other 15 cases, deformity and shortness of the tibia were related to the state of distribution and the degree of damages of the epiphyseal plate. In the affected cartilage matrix, cystoid degeneration, fibrous striation, bone spicule, necrotic foci and rent were found. Inadequate metachromasia suggests the abnormal staining of sulfated glycosaminoglycans and alcian blue stainability may be attributable to the abnormal interactions between proteoglycan and other matrix components. At the interface of cartilage and bone, irregular calcification, fibrotic scar and sealing by osseous tissue with relation to vascularity were found. These changes in this study indicated the failure of modeling on the epiphyseal plate, showing disturbance of chondrocytic differentiation and abnormal formation of the matrix.     

A case of feline sporotrichosis

We excised surgically a feline granulomatous lesion and performed histopathological, mycological and molecular examinations. As a result, it was diagnosed as sporotrichosis, which was the second recorded case of a cat so afflicted in Japan. After the operation, we recognized another nodule on the lymph node. Histopathological examination was therefore performed, but no fungi were detected. To prevent recurrence, the cat was administered a antimycotic drug, itraconazole. As a result, no recurrence was found. Excision of the lesion is the treatment of choice for feline sporotrichosis.     

In vitro effect of recombinant human granulocyte colony-stimulating factor on canine neutrophil apoptosis

Apoptosis is essential in eliminating neutrophils (polymorphonuclear leukocytes: PMNs) in animals. The suppression of PMN apoptosis is believed to be beneficial in eradicating pathogens and is implicated in the pathogenesis of human inflammatory diseases. In the present study, canine PMNs were stimulated with recombinant human granulocyte colony-stimulating factor (rhG-CSF) to investigate the in vitro effect on the apoptosis of canine PMNs. Apoptotic cell rates were assessed by flow cytometry in relation to the ability of PMNs to produce reactive oxygen species (ROS). Canine PMN apoptosis was markedly suppressed by rhG-CSF treatment, in association with the retention of the PMN ability to produce ROS. The addition of cycloheximide abolished this suppression by rhG-CSF. Moreover, canine PMNs, which were stimulated by rhG-CSF, expressed high levels of anti-apoptotic mcl-1 gene mRNA, as quantified by real-time polymerase chain reaction method. The results suggest that PMNs, stimulated by G-CSF, could work effectively over a longer period to eliminate pathogens, and that the prolongation of the PMN life-span might occasionally aggravate tissue injuries in dogs. In addition, the suppression of PMN apoptosis seems to be mediated by the induction of anti-apoptotic mcl-1 gene expression.     

The effect of free and polyethylene glycol-liposome-entrapped albendazole on larval mobility and number in Toxocara canis infected mice

As part of our exploratory drug research on the larva migrans that causes roundworm in dogs and cats, this study was carried out to clarify the effect of free and liposome-entrapped (LE) albendazole in Toxocara canis infected mice. In infected mice, evaluation of mobility and number of larva were examined in detail in the brain, skeletal muscle and liver. Larva mobility was evaluated by using the relative mobility (RM) value. Albendazole was LE as one of the drug delivery systems (DDSs). Polyethylene glycol (PEG) was added to the liposome in order to avoid evoking a response by the reticuloendothelial system (RES). By using the albendazole PEG-LE delivery system, it was possible to target the larvae in the mouse brain and liver resulting in a decrease in the number of larvae. In the skeletal muscle of the infected mice, the intraperitoneal dosages of PEG-LE albendazole did not cause a complete decrease in the number of larvae, even though free albendazole did cause the number to decrease. Therefore, it is necessary to take into consideration the migrating stage of the larvae before the initiation of any drug administration.     

Sex identification of Japanese black bear,Ursus thibetanus japonicus,by PCR based on amelogenin gene

A method for sex identification of the Japanese black bear was examined using a polymerase chain reaction (PCR) and sequencing of a part of the amelogenin gene. This gene is located on the X and Y chromosomes, and there are 54 nucleotide deletions on the Y chromosome-specific gene. Forty-seven (26 male and 21 female) DNA samples and 23 (13 male and 10 female) DNA samples, respectively extracted from white blood cells and hairs of Japanese black bears were analyzed. The primers SE47 and SE48 from this X-Y homologous region were used in sex identification by PCR amplification. These primers amplified X- and Y-specific bands, which could be used to discriminate between sexes by a length polymorphism in all samples. We suggest that PCR amplification using the primers SE47 and SE48 is useful for sex determination of the Japanese black bear and could be applied to DNA analysis of small samples such as hairs.     

Metabolism of permethrin isomers in American cockroach adults,house fly adults,and cabbage looper larvae

Forty-two insect metabolites of [1RS,trans]-and [1RS,cis]-permethrin are tentatively identified in studies with Periplaneta americana adults, Musca domestica adults, and Trichoplusia ni larvae involving administration of ¹⁴C preparations labeled in either the alcohol or acid moieties. The less-insecticidal trans isomer is generally metabolized more rapidly than the more-insecticidal cis isomer, particularly in cabbage looper larvae, and metabolites retaining the ester linkage appear in larger amount with cis-permethrin. Although the dichlorovinyl group effectively blocks oxidation in the acid side chain, the permethrin isomers are metabolized by hydrolysis and hydroxylation at the geminal-dimethyl group (either trans- or cis-methyl substituent) and the phenoxybenzyl group (predominantly at the 4′-position in all species but also at the 6-position in house flies). The alcoholic and phenolic metabolites are excreted as glucosides, and the carboxylic acids are excreted as glucosides and amino conjugates (glycine, glutamic acid, glutamine, and serine) with considerable species variation in the preferred conjugating moiety.     

Characterization of a phytoplasma associated with witches' broom disease of potatoes in Korea

 Symptoms of witches' broom disease caused by phytoplasma, including general stunting and yellowing, were observed in potatoes (Solanum tuberosum L.) in a storehouse on Jeju Island, Korea in 1998. Based on sequence analysis of DNA products from polymerase chain reaction (PCR)-amplified 16S ribosomal DNA and 16S–23S spacer region using universal phytoplasma primers, the phytoplasma associated with potato witches' broom disease (PWB) was identified as a member of 16S-group VIII. It was most closely related to elm AH phytoplasma (99.7% similarity, accession no. AF268895), which is in the clover proliferation (CP) subgroup. This report is the first from the East Asian continent of a plant pathogenic phytoplasma belonging to the CP subgroup and includes the nucleotide sequence of most of the potato phytoplasma 16S rDNA. Received: May 1, 2002 / Accepted: July 1, 2002     

Similarity between Copper Resistance Genes from Pseudomonas syringae pv. actinidiae and P. syringae pv. tomato

Twenty-eight strains of Pseudomonas syringae pv. actinidiae isolated in 1984, 1987 and 1988 from kiwifruit orchards in Japan were tested for their resistance to copper sulfate. All strains isolated in 1984 were copper sensitive with a minimum inhibitory concentration (MIC) of cupric sulfate of 0.75 mM. However, some strains isolated in 1987 and 1988 were resistant, with the MIC ranging from 2.25 to 3.0 mM. All copper-resistant strains contained at least one of two plasmids, pPaCul (about 70.5 kb) or pPaCu2 (about 280 kb), or both. In a copper-resistant strain Pa429, the location of the copper-resistance gene(s) was examined by insertional inactivation with Tn5. The MIC of copper sulfate in the copper-sensitive mutant obtained by Tn5 tagging decreased from 2.75 to 0.75 mM. The 14.5 kb BamHI fragment, designated pPaCuB14, containing the same locus mutagenized with Tn5 was cloned from pPaCu1. However, pPaCuB14 did not confer copper resistance in the transformant of copper-sensitive strain Pa21R, suggesting that this clone did not contain a full set of copper-resistance gene(s). Then a cosmid library of pPaCu1 was constructed and six cosmid clones hybridized with pPaCuB14 were selected. One of the six cosmids, designated pPaCuC1, conferred a near wild-type level of copper resistance in the transformant of the copper-sensitive strain. pPaCuC1 had a homologous region that hybridized with all of the PCR-amplifled fragments of copA, copB, copR, and copS genes of P. syringae pv. tomato. DNA sequence analysis of the homologous region revealed the existence of four open reading frames (ORF A, B, R and S) oriented in the same direction. The predicted amino acid sequences of ORF A, B, R and S had 80, 70, 97 and 95% identity with CopA, B, R and S of P. syringae pv. tomato, respectively. Received 5 July 2001/ Accepted in revised form 27 September 2001