New severe strains of Melon necrotic spot virus: symptomatology and sequencing

New strains of Melon necrotic spot virus (MNSV), designated MNSV-YS and MNSV-KS, caused much more severe growth retardation on melon plants than MNSV-NH, which was previously reported as the most severe strain of MNSV in Japan. MNSV-YS spread much more quickly than MNSV-NH in infected plants, and induced more severe growth retardation, even though the appearance of necrotic lesions on inoculated cotyledons was much slower. MNSV-KS had properties intermediate between those of the other two strains. The results suggest that faster-spreading strains can multiply more rapidly as a result of lower levels of activity in inducing necrotic lesions in melon plants. The complete sequences of MNSV-YS and MNSV-KS were determined, and an RT–PCR–RFLP method based on these sequences was successfully developed to detect and discriminate between the three strains.     

Growth Complementation of hrpXo Mutants of Xanthomonas oryzae pv. oryzae by Virulent Strains in Rice Cultivars Resistant and Susceptible to the Parental Strain

Xanthomonas oryzae pv. oryzae (X. o. pv. oryzae) T7174 is virulent on rice cultivar IR24 and avirulent on IR-BB2. From recent reports, some virulence and avirulence factors of plant pathogenic bacteria are transferred to plant cells through the hrp-dependent type III secretion system. In this study, we investigated the involvement of hrp genes in the compatible and the incompatible interactions between rice and X. o. pv. oryzae after co-inoculation with hrpXo mutants derived from T7174 and virulent strains. Growth of the mutants, named 74ΔHrpXo and 76ΔHrpXo, was repressed in IR24 when the mutants were applied alone. However, growth of the mutants was complemented by co-inoculation with virulent strains. Growth of bioluminescent hrpXo mutant 76ΔHrpXo in IR24 and its growth in IR-BB2 after co-inoculation with T7133, which is virulent on both cultivars, was equally complemented, as detected by bioluminescence from the mutant. On the other hand, only partial complementation of growth of T7174L76, which is a bioluminescent and pathogenic derivative of T7174, by T7133 was observed in IR-BB2. Thus, growth of the hrpXo mutant of X. o. pv. oryzae was complemented by virulent strains in both susceptible and resistant rice leaves with the parental strain. Received 21 July 2000/ Accepted in revised form 26 October 2000     

Inhibitory and promotive allelopathy in rice (Oryza sativa L.)

Studies on allelopathic rice, which inhibits the germination and development of weeds such as barnyardgrass ( Echinochloa spp.) and ducksalad [ Heteranthera limosa (Sw.) Wild.], have been strongly conducted in the USA, the Philippines, Japan, Korea, and other countries since the 1980s. Weed-suppressing rice plants can compete with weeds for light and mineral nutrients by their rapid and thick growth characteristics, and probably also by their allelopathic effects. It was suggested that allelopathic properties were inherited from parents. Different screening methods for finding allelopathic rice simply and rapidly have been proposed. In contrast, stimulative allelopathy in rice, which promotes germination of the dormant and dormancy-awakened seeds of Monochoria vaginalis (Burm. f.) Kunth var. vaginalis , has been investigated by the authors since the 1990s. Rice seeds promoted the germination in the dark and in light under submerged conditions. Our experiments suggested that rice seeds, hulls, plants and straw contain stimulative substance(s) that promote seed germination of M. vaginalis .     

Rhizome-induction activity of chiral 2-α-methylbenzylamino-4-alkylamino-6-chloro-1, 3, 5-triazines in Cyperus serotinus Rottb.

A cytokinin-like effect of chiral 2-α-methylbenzylamino-4-alkylamino-6-chloro-1,3,5-triazines was found using a rhizome-inducing assay with Cyperus serotinus Rottb. tubers. C. serotinus tubers germinated in distilled water yielded plantlets with roots and leaves. Secondary rhizomes were normally not observed within the regular 14-day incubation time in water culture, whereas after increasing incubation periods a very short rhizome appeared (controls). 6-Benzylaminopurine (BA) significantly stimulated rhizome induction, while other plant hormones were inactive. The (R)-isomers of the 1, 3, 5-triazine compounds also stimulated induction of the rhizomes, whereas the (S)-isomers did not. The described rhizome induction system seems to be suitable as a cytokinin bioassay. The (R)-1, 3, 5-triazine compounds showing rhizome-inducing activity (RI activity) inhibited root formation and plant growth at high concentrations with symptoms which were very similar to those of BA. Therefore, the (R)-isomers appear to act as cytokinins in the rhizome induction assay.     

Effects of alpha-pinene odor in different concentrations on stress-induced hyperthermia in rats

Stress-induced hyperthermia is observed in animals exposed to stressful conditions. In our previous study, plant-derived fragrances such as green odor and alpha-pinene were shown to suppress this stress response in rats. In the present study, we examined the concentration-dependence of the alpha-pinene effects on stress-induced hyperthermia. Male rats carrying telemetry transmitters were transferred individually to a new cage containing bedding that had been sprayed with 0.3, 0.03, and 0.003% concentrations of alpha-pinene or control solvent. Following transfer to the novel environment, the body temperature increased significantly, and this response was clearly suppressed when the cage was scattered with 0.03% alpha-pinene only. These results suggest that the effect of alpha-pinene on stress-induced hyperthermia can be observed only at a certain concentration.     

Establishment of a potency test by ELISA for a rabies vaccine for animal use in Japan

The ELISA we developed was able to determine the antigen content and was suitable for a potency test, and we described a relative potency assay method which determines the potency of test vaccines by comparing the ELISA value of a test vaccine to that of a reference vaccine. In the present study, we standardized the reference vaccine used for determining the potencies of test vaccines, and established a potency test by ELISA. We evaluated the proposed reference vaccine by the neutralizing antibody responses in dogs after vaccination, by the challenge protection test in guinea pigs (GP potency test), which is the earlier official potency test used in Japan, and by the NIH potency test, which is widely used throughout the world. The results showed that a 4-fold dilution of the proposed reference vaccine induced sufficient immunity in dogs. A 3-fold dilution of the proposed reference vaccine passed the GP potency test. The international units (IU) calibrated by the NIH potency test were 3.7 IU/dose. From the results and the WHO recommendation that veterinary rabies vaccines should have a potency of at least 1.0 IU/dose, we determined to dilute the proposed reference vaccine by 3 fold and regarded it as the reference vaccine. Finally, we confirmed that there is a good agreement between the results of the potency test by ELISA and the results of the GP potency test. The establishment of the potency test by ELISA has made it possible to monitor the potency in the production process and has contributed to the stable production of the vaccine.     

Substances derived from 4-ethyl octanoic acid account for primer pheromone activity for the "male effect" in goats

A major constituent of the characteristic "goaty odor" 4-ethyl octanoic acid (4EOA) was previously shown to have no primer pheromone activity. This was also confirmed by our own bioassay system utilizing the recording technique of neural activity of the hypothalamic gonadotropin-releasing hormone pulse generator in goats. However, when the synthetic 4EOA solution was kept at room temperature for several months, primer pheromone activity appeared in the same solution. Headspace gas chromatography/mass spectrometry analysis revealed that there were several newly formed substances in addition to 4EOA samples with primer pheromone activity. These results suggest that 4EOA derived substance(s) but not 4EOA itself is(are) primer pheromone in goats.     

Classification of abomasal displacement in cows according to histopathology of the liver and clinical chemistry

Histopathological features of livers and blood chemical values in cows with abomasal displacement were investigated. Liver biopsy samples were collected during redressment operations in 92 cows with abomasal displacement, and the samples were stained with haematoxylin and eosin or periodic acid Schiff (PAS). Blood was collected for chemical tests. Livers were histopathologically divided into the following four types: normal histology cases (21%), fatty degeneration cases (36%), cloudy swelling cases (19%) and fatty degeneration cases with cloudy swelling (24%). The number of PAS-positive samples was significantly higher in the normal histology group and significantly lower in the severe fatty degeneration group and severe cloudy swelling group. Cows with fatty degeneration had significantly higher levels of serum 3-hydroxybutyric acid, non-esterified fatty acid and aspartate aminotransferase than did those with cloudy swelling or normal histology. The results indicate that the morbid conditions of cows with abomasal displacement can be classified into four types.     

Antibody response to toxic shock syndrome toxin-1 of Staphylococcus aureus in dairy cows

Antibody response to toxic shock syndrome toxin-1 (TSST-1) of Staphylococcus aureus in dairy cows was examined by enzyme linked immunosorbent assay (ELISA). Serum antibody to TSST-1 was not detected in 39 (76.5%) of 51 calves, which were 1-6 months of age. In contrast, TSST-1 antibody was demonstrated in 1728 (72.6%) of 2380 lactating cows housed on 36 dairy farms. The ELISA values of antibody ranged from 0.2 to 3.0 OD and presented a distribution with the peak at 1.6 OD. The mean ELISA value differed between farms, and it increased slightly along with parturient history. Somatic cell counts of milk from 174 lactating cows was compared with TSST-1 antibody and tst1,000,000 cells per ml. The mean ELISA values in milk were lower than those of sera, but they rose as somatic cells increased. The tst gene of S. aureus detected in 76.0-86.2% of the milk samples containing somatic cells > 500,000 cells per ml, a level which indicates mastitis. The data suggests that many lactating cows may be infected by TSST-1- producing S. aureus.     

In vitro but not in planta encapsidation of Rice gall dwarf virus core particles by the outer capsid P8 protein of Rice dwarf virus expressed in transgenic rice plants

Transencapsidation of the Rice gall dwarf virus (RGDV) inner core by the Rice dwarf virus (RDV) outer capsid P8 protein was examined in vitro and in planta. When RGDV core particles were incubated with an extract from RDV P8-transgenic rice leaf tissue, RDV P8 encapsidated the RGDV core particles to form double-shelled virus-like particles in vitro. In contrast, when RDV P8-transgenic rice plants were inoculated with RGDV, progeny RGDV particles contained RGDV P8 but RDV P8 was not detectable in the virions. No significant differences were found in acquisition by the vector insects and subsequent transmission rates between RGDV infecting nontransgenic rice plants and those infecting RDV P8-transgenic rice plants. These results indicate that mechanisms of and/or requirements for interactions between P8 and the inner core particles of phytoreoviruses differ between in vitro and in planta.