Detection and differentiation of Leptospira spp. serovars in bovine semen by polymerase chain reaction and restriction fragment length polymorphism

In view of the importance of venereal transmission of bovine leptospirosis, the objective of the present study was to apply the polymerase chain reaction (PCR) to 26 serovars of Leptospira interrogans, L. borgpetersenii, L. santarosai, L. noguchii and L. biflexa, to determine the detection threshold in semen samples and to evaluate the possibility of differentiation among serovars using 19 restriction endonucleases. The results showed that all serovars were amplified and the detection threshold in semen samples of a bull was 100 bacteria/ml. Using endonucleases we could classify the 26 serovars into eight groups. The present results show that PCR is a method of great potential for the detection of Leptospira spp. at bovine artificial insemination centers.     

Does future climate change facilitate expansion of evergreen broad-leaved tree species in the human-disturbed landscape of the Korean Peninsula? Implication for monitoring design of the impact assessment

To quantitatively assess future change of evergreen broad-leaved tree species’ distributions in human-disturbed landscapes of the Korean Peninsula under climate change, potential habitats (PHs) were projected for four important evergreen broad-leaved tree species (Quercus acuta, Castanopsis sieboldii, Machilus thunbergii, and Neolitsea sericea) by species distribution models (SDMs). The distribution data (presence/absence) of the target species in Korea and Japan were used as response variables for SDMs, and climatic data were used as explanatory variables. Three general circulation models under A2 emission scenarios were used as future climate scenarios for the years 2070–2099. Potential habitats masked by land-use data (PHLUs) were projected to assess the impact of anthropogenic activities. Highly accurate SDMs were obtained for all the target species. The current PHs were decreased to 21–35 % by the anthropogenic activities. Future PHLUs for all the target species were projected to increase by 2.0–18.5 times of current PHLUs. These results suggest that all the target species are applicable as indicator species for monitoring in the Korean Peninsula, even if anthropogenic effects are incorporated. Variation of the increasing rate was caused by the differences in the response to temperature changes. M. thunbergii responded sensitively to the increase of minimum temperature of coldest month and had a largest increase in PHLUs under future climate. Therefore, M. thunbergii is considered to be most appropriate species for monitoring the changes of horizontal distributions above all focal evergreen broad-leaved tree species.     

Treatment with an aromatase inhibitor induces complete sex change in the protogynous honeycomb grouper (Epinephelus merra)

To study the mechanism of protogynous sex change in honeycomb grouper, the wild fish of both sexes and transitionals were collected for one year, and changes in gonadal structures and serum levels of sex hormones in each individual were examined. The onset of sex change was associated with low serum estradiol - 17β (E2) levels. In order to clarify whether E2 deprivation causes sex change, maturing females were implanted with the non-steroidal aromatase inhibitor (Fadrozole, AI). AI caused complete sex change from female to functional males in two and half months. The present results suggest that E2 is an important endogenous factor which regulates protogynous sex change in the honeycomb grouper.     

The incidence of disseminated intravascular coagulation in dogs with malignant tumor

The incidence of DIC in 208 dogs with a malignant tumor was evaluated. The incidence of DIC was 9.6% in dogs with a malignant tumor which was a solid tumor in all. In 164 dogs with a malignant solid tumor, the incidence of DIC was 12.2%. The incidence of DIC in dogs with hemangiosarcoma, mammary gland carcinoma and adenocarcinoma of the lung was significantly higher than that in dogs with other malignant tumors. These results suggested that special care in looking for DIC should be taken in dogs with a malignant solid tumor.     

A case of feline pancreatitis

A 14-year-old, spayed female, domestic shorthair cat was referred to us with anorexia, pyrexia, and jaundice. Total bilirubin (TBIL) and feline trypsin-like immunoreactivity (fTLI) levels were remarkably high. Based on laparoscopic biopsy of the pancreas, the cat was diagnosed as having pancreatitis. As a result of treatment with a synthetic protease inhibitor and corticosteroid, the TBIL and fTLI values returned to normal and the clinical course was good.     

Phenotypic analysis of hepatic T lymphocytes in a dog with chronic hepatitis

We evaluated hepatic T lymphocyte phenotypes in a dog with chronic hepatitis. Before treatment, numerous CD3+ lymphocytes were demonstrated in the liver, and the ratio of CD4+/CD8+ was remarkably high (2.96; reference range, 0.33+/-0.12). After treatment, CD3+ lymphocyte infiltration in the liver was reduced, and the ratio of CD4+/CD8+ decreased to 0.31. Therefore, hepatic T lymphocytes, especially CD4+ lymphocytes, might play a central role in the pathogenesis of this dog with chronic hepatitis.     

Effects of complete submergence on seedling growth and survival of five riparian tree species in the warm-temperate regions of Japan

To reveal the relationship between tolerance to submergence and the distribution ranges of riparian tree species in the floodplains of the warm-temperate regions in Japan, we assessed the response and tolerance of current-year seedlings of two Salix and three Ulmaceae species to submergence treatments for 1, 2, 4, and 8 weeks. We investigated shoot elongation, number of leaves, and survival rate of seedlings during the experiment, and biomass allocation was determined at the end of the experiment. Shoot elongation and leaf production of seedlings were suppressed in all the species during the submergence treatments. However, the seedlings of Salix chaenomeloides and S. pierotii elongated shoots rapidly and produced leaves after the treatments. All the underground biomass in these two species was larger than aboveground biomass. These results are considered as typical responses of flood-tolerant species. In contrast, the underground biomass of Ulmaceae seedlings (Celtis sinensis var. japonica, Aphananthe aspera, and Ulmus parvifolia) decreased more rapidly than aboveground biomass during prolonged submergence. Furthermore, A. aspera seedlings also showed suppression of shoot elongation and leaf production after the treatments. Our results indicate that the three Ulmaceae species are less tolerant to submergence than the two Salix species. In conclusion, we suggest that submergence stress is one of the important factors determining the distribution of tree species in the floodplains of lowlands in the warm-temperate regions of Japan.     

A multiplex PCR for the detection of Brucella spp. and Leptospira spp. DNA from aborted bovine fetuses

Bovine brucellosis and leptospirosis are important causes of bovine abortion around the world. Both diseases can be serologically diagnosed, but many factors may cause false positive and negative results. Direct methods based on bacteriological isolation are usually employed, but they are difficult, time consuming and dangerous. Monoplex polymerase chain reaction (PCR) have been successfully described for the detection of Brucella spp. and Leptospira spp. Aiming at improvement in the direct diagnosis, a multiplex PCR (mPCR) for the detection of these agents in aborted bovine fetuses is described. The detection threshold of the mPCR was evaluated in experimentally contaminated bovine clinical samples using a conventional proteinase K/SDS or a boiling-based extraction protocols. The mPCR was applied to two groups of clinical samples: 63 episodes of bovine abortion and eight hamsters experimentally infected with Leptospira interrogans serovar pomona. Adopting microbiological isolation as reference, the test showed a sensitivity of 100% in both groups of clinical samples. Seven samples collected from bovine fetuses were Brucella spp. culture negative but showed positive results in mPCR. Regarding Leptospira spp. detection, similar results were observed in three bovine clinical samples. All hamsters infected with Leptospira were positive in both microbiological culture and mPCR. The boiling extraction protocol showed better results in some clinical samples, probably by the removal of PCR inhibitors by heat treatment. The high sensitivity, simplicity and the possibility of detection of both bacteria in a single tube reaction support the use of the mPCR described in the routine diagnosis.     

Root and crown rot of strawberry caused by Pythium helicoides and its distribution in strawberry production areas of Japan

Pythium species were isolated from seedlings of strawberry with root and crown rot. The isolates were identified as P. helicoides on the basis of morphological characteristics and sequences of the ribosomal DNA internal transcribed spacer regions. In pathogenicity tests, the isolates caused root and crown rot similar to the original disease symptoms. Multiplex PCR was used to survey pathogen occurrence in strawberry production areas of Japan. Pythium helicoides was detected in 11 of 82 fields. The pathogen is distributed over six prefectures.