Detection of the anti-P53 antibodies in dogs with tumors

To detect the anti-P53 antibodies of dogs with tumors, a GST-recombinant canine (rc) P53 fusion protein was expressed and purified. Immunoblot analysis was performed using this GST-rcP53 fusion protein as an antigen and serum samples from dogs suffering from tumors as primary antibodies. Out of 16 serum samples obtained from various tumor cases, four samples showed reaction with GST-rcP53. In contrast, serum from other 12 dogs with tumors, four dogs with non-neoplastic diseases and two control healthy dogs (as controls) did not show any reaction with GST-rcP53 in immunoblotting. The p53 gene mutation and the P53 protein expression were examined, using the tumor tissues to explore the relationship between the existence of the GST-rcP53 bands, gene mutations of p53 and the accumulation of P53 protein. One case, which showed a clear GST-rcP53 band, had a point mutation of the p53 cDNA and showed nuclear accumulation of P53 protein. These results suggest that the anti-P53 antibodies are also produced in tumor dogs with p53 gene mutations.     

Analysis of the 18S rRNA gene sequence of a Hepatozoon detected in two Japanese dogs

Partial sequences of the 18S rRNA gene (625 bp) from a Hepatozoon detected in two canine hepatozoonosis cases, one clinical and one subclinical, in Japan were analyzed. Both sequences were identical to each other and they were closely related to the Hepatozoon canis strain found in Israel with 99% (617/625) nucleotide identity. Both Hepatozoon americanum and Hepatozoon catasbianae were distantly related to the Japanese Hepatozoon with 94% (586/625) and 91% (566/625) identities, respectively. In a phylogenetic tree, the Japanese Hepatozoon was most closely related to H. canis from Israel but was significantly different than H. americanum and H. catasbianae. These results suggest that the Hepatozoon detected in the Japanese dogs might be a strain variant of H. canis, but is apparently a different species than H. americanum.     

Changes in hepatic enzyme activities in transgenic mice carrying human prototype c-Ha-ras gene treated with diethylnitrosamine

Activities of malate dehydrogenase (MDH) and aspartate aminotransferase in the malate-aspartate shuttle were significantly increased in the cytosolic fractions of livers with early neoplastic symptoms such as swelling and discoloration in transgenic mice carrying the prototype human c-Ha-ras gene, Tg-rasH2 mice, which were administered with diethylnitrosamine (DEN) as a carcinogenic chemical at a dose of 200 mg/kg body weight. Cytosolic MDH/LDH (lactate dehydrogenase) (ML) ratio increased significantly and was considered to be a useful marker to characterize the energy metabolism at early neoplastic stage in livers of the Tg-rasH2 mice.     

Specific Oligonucleotide Primers Based on Sequences of the 16S-23S rDNA Spacer Region for the Detection of <Emphasis Type="Italic">Burkholderia gladioli </Emphasis>by PCR

Specific primers were designed based on the sequences of the spacer region between the 16S and 23S ribosomal DNA (rDNA) for direct, rapid and specific detection of Burkholderia gladioli. These primers were named GLA-f and GLA-r. PCR performed on boiled bacterial suspensions yielded an amplification product of approximately 300 bp. No products from other bacterial species, including B. glumae were amplified, even after complete DNA extraction by the cetyltrimethyl-ammonium bromide (CTAB) method. Using the specific primers designed in this study, the PCR method can detect B. gladioli in plant samples within 6 hr. These data demonstrate the potential of specific PCR for the detection of B. gladioli. Received 10 December 2001/ Accepted in revised form 15 April 2002     

Divergence of p33/34 gene of Theileria found in Cervus nippon in Japan

The 18S rRNA gene and the piroplasm major immunodominant protein gene (p33/34) of Theileria from various subspecies of sika deer in 8 different locations of Japan were analyzed. The similarity between 633 bp partial sequences of the 18S rRNA gene among various subspecies of sika deer was found to be between 99.7% and 100%. While the percent identities of the 412 bp partial p33/34 gene sequence and deduced amino acid sequences between Theileria of sika deer from Yamaguchi Prefecture and those found in deer from other Prefectures, were comparatively low, 68.7% to 70.1% and 64.1% to 70.0% respectively. These findings suggest that there are at least two genetically distinct strains of Theileria of sika deer in Japan.     

Novel variations and loss of heterozygosity of BRCA2 identified in a dog with mammary tumors

OBJECTIVE: To establish novel polymorphic markers for analysis of loss of heterozygosity (LOH), so as to study the possible involvement of BRCA2 in mammary tumors obtained from dogs. SAMPLE POPULATION: Blood samples, mammary gland specimens, or mammary tumors from 3 tumor-bearing dogs and 10 tumor-free dogs. PROCEDURES: Nucleotide sequence analysis was performed with a DNA autosequencer. Loss of heterozygosity analysis was performed for markers established in the present study. The expression level of canine BRCA2 was quantified by real-time PCR analysis. RESULTS: 3 novel microsatellite markers with high heterozygosity rates (> 50%) were established, and the previously reported marker for canine BRCA2 gene locus was improved. These markers were used for the analysis of DNA from formalin-fixed and paraffin-embedded samples. By use of these markers, LOH in canine BRCA2 was identified as a result of recombination. In mammary tumor DNA that corresponded to the LOH-positive dog, the level of canine BRCA2 expression was decreased compared with that of nonneoplastic mammary gland tissue; the open reading frame contained 4 missense variations, 1 insertion variation, and 1 silent variation, some of which were localized to functional domains. CONCLUSIONS AND CLINICAL RELEVANCE: 3 novel polymorphic markers were developed for LOH analysis of canine BRCA2 and identified a dog with LOH with some variations in the functional domains. These markers could be useful for assessing the relevance of BRCA2 variation in mammary tumors of dogs.     

Quantitative structure–fungicidal activity relationships of N-(4-difluoromethoxybenzyl)­pyrimidin-4-amines against wheat and barley fungi

A set of N-(4-difluoromethoxybenzyl)pyrimidinamines with various substituents at the 4- and 5-positions of the pyrimidine ring and at the benzyl position were prepared, and their fungicidal activities against wheat brown rust, Puccinia recondita, and barley powdery mildew, Erisiphe graminis, were measured. Variations in each of these activities were quantitatively analysed by the use of physicochemical substituent parameters and a regression analysis. Each of these activities was parabolically correlated with the steric bulkiness of the pyrimidine substituents and with the bulkiness or the hydrophobicity of the benzylic substituents. © 1999 Society of Chemical Industry     

Dual appearance of xanthophores,and ontogenetic changes in other pigment cells during early development of Japanese flounder <Emphasis Type="Italic">Paralichthys olivaceus</Emphasis>

Flatfishes display a left–right asymmetry that is unique in the animal kingdom. In order to clarify the mechanisms of the asymmetrical development of pigment cells, changes in pigment cell densities were examined in Japanese flounder Paralichthys olivaceus. During development from symmetrical larvae to asymmetrical juveniles, pigment cell densities were monitored on the skin on both the left side (ocular side in juvenile; eventually has two eyes) and the right side (blind side in juvenile; eventually has no eyes). A symmetrical and constant decrease was observed in leucophores and larval type melanophores. A mostly symmetrical (slightly delayed on the blind side) and constant increase in iridophores from metamorphosis was observed. Adult-type melanophores appeared and then increased only after metamorphosis on the ocular side. However, the pattern of xanthophores was complicated: they first existed symmetrically and decreased symmetrically until metamorphosis, and they later increased only on the ocular side. The dual appearance of the xanthophores, as well as the differences between their depths and sizes on the ocular and blind sides, may suggest the presence of two types of xanthophores—just as melanophores are well known to exhibit two types. The ontogenetic study of pigment cells described here is likely to help to elucidate the process of abnormal pigmentation in flatfishes.     

Predation on fish larvae by moon jellyfish <Emphasis Type="Italic">Aurelia aurita</Emphasis> under low dissolved oxygen concentrations

ABSTRACT:   Laboratory experiments were conducted to test the hypothesis that low dissolved oxygen concentrations have the potential to enhance the predation rate on fish larvae by moon jellyfish Aurelia aurita which is increasing in abundance in the coastal waters of Japan. Larvae of the red sea bream Pagrus major in four size classes (2.9, 4.1, 6.2 and 8.6 mm in standard length) were used as prey in a short-term predation experiment. No change in the bell contraction rate of the jellyfish (mean bell diameter, 100.4 ± 10.2 mm) was observed at the oxygen concentrations tested (1, 2 and 4 mg/L, and air-saturated, 5.5–6.0 mg/L), suggesting a strong tolerance to a decline in the oxygen concentration. More than 80% of the 2.5 and 4.1-mm size-class larvae were predated on by the jellyfish at all oxygen concentrations during the 15-min trials in 10-L tanks. The 6.2 and 8.6-mm size-class larvae were able to escape from the jellyfish apparently due to their developed swimming ability at the two higher dissolved oxygen concentrations. There was an increase in predation on larvae of these two size classes at the two lower dissolved oxygen concentrations. Predation by moon jellyfish on fish larvae can be more intense at low dissolved oxygen concentrations that commonly occur in the coastal waters of Japan.