Expression in Escherichia coli and purification of the functional feline granulocyte colony-stimulating factor

Feline granulocyte colony-stimulating factor (G-CSF) with an N-terminal histidine hexamer tag was expressed as inclusion bodies in E. coli. The G-CSF solubilized in 6 M guanidine solution was absorbed onto a Ni-NTA column and, after washing with decreasing concentrations of guanidine, eluted with imidazole in a soluble and apparently pure form. The activity of the recombinant feline G-CSF was 3×10⁶ U/mg protein, as assayed by its stimulatory effect on NFS-60 cell proliferation. When a low level of purified feline G-CSF was administered once a day for two successive days to cats, the number of neutrophil increased 4-fold while the levels of other blood cell types remained virtually unchanged. Daily administration of G-CSF for a total of 11 days led to a more than 10-fold increase in neutrophils, an 8-fold increase in the number of monocytes and 2-fold increase in lymphocytes. No severe side effects or antibody production was observed in cats after administration of G-CSF.     

Identification of ovalbumin phenotypes of the Asian indigenous chicken populations using polymerase chain reaction-restriction fragment length polymorphism

Three electrophoretic variations (AA, BB and AB) of ovalbumin controlled by codominant alleles Ov^A and Ov^B have been observed in various chicken populations. We compared nucleotide sequences of the open reading frame between two alleles of ovalbumin gene. The difference between the two alleles was found as a non‐synonymous substitution of asparagine to aspartic acid as a result of AAT to GAT point mutation at position 8032–8034 in exon 8. We developed polymerase chain reaction‐restriction fragment length polymorphism (PCR‐RFLP) protocol in combination with Mbo I restriction endnuclease mapping for the detection of this substitution. By the PCR‐RFLP the allelic frequency of the Ov^B was estimated to be within the range of 0.000–0.150 in 11 Asian indigenous chicken populations and 0.000 in four improved breeds used in the present study. Gene frequency, estimated by PCR‐RFLP in the present study, paralleled that obtained by protein polymorphisms of egg white. Thus, this study provides, for the first time, information of the occurrence of ovalbumin allele Ov^A and Ov^B in Asian indigenous chicken populations.     

Early growth of hinoki (<Emphasis Type="Italic">Chamaecyparis obtusa</Emphasis>) trees under different topography and edge aspects at a strip-clearcut site in Kyushu,Southern Japan

The positive and negative effects of the residual stand edge at a strip-clearcut site were examined on the initial growth of hinoki (Chamaecyparis obtusa) planted in areas with diverse topography. On the south edge, the gap light index was lower than on the north edge and strip center; however, the vapor pressure deficit was also lower than on the north edge and strip center, which resulted in lower physiological stress of hinoki on the south edge. Tree size on the south edge did not exceed that on the north edge and strip center. These results indicated that low light conditions due to residual trees negatively affected growth on the south edge even under the positive effect of microclimate alleviation. In valley, tree size in the second year after planting was smaller than on ridge and slope; however, tree growth during the following 2 years was higher in valley. Surface soil in valley was thinner with rockier substrates than on ridge and slope; that is, soil sedimentation type and substrates influenced the initial growth of hinoki with undeveloped roots, and after that, topography started to influence growth, probably because hinoki roots penetrated into deeper soil, which is influenced by the water gathering capacity of the valley. The initial growth of hinoki at the strip-clearcut site was predominantly affected by light rather than microclimate alleviation. This alleviation is expected to disappear when hinoki trees planted in the strip center grow enough to give additional shade to slow-growing edge hinoki.     

Growth of tiger puffer Takifugu rubripes in closed recirculating culture system

ABSTRACT:   Two feeding experiments were conducted to elucidate growth performance of tiger puffer in a 10 m³ water volume closed system. In experiment 1, 1000 fish of 3.5 g average body weight were fed tiger puffer commercial feed twice daily to apparent satiation, 6 days a week for 224 days. Sand-filtered sea water was used and no water was exchanged during the rearing period. Immediately after cutting of lower teeth at day 112, daily feed consumption decreased greatly and 60 fish died in few days. Feeding rates recovered and then decreased gradually as nitrate levels increased from 600–1048 mg N/L. Fish grew to 343 g with 91% survival rate and 87% feed efficiency. Rearing conditions of experiment 2 were similar to experiment 1, except that culture water was exchanged to maintain the nitrate level less than 600 mg N/L during the 224-day experiment. Mortality and reduction of feed consumption occurred immediately after teeth cutting as was observed in experiment 1. Significant reduction of feed intake was not found during other rearing periods. Fish of 3 g grew to 303 g with 91% survival rate and 72% feed efficiency.     

Reproductive biology of blue marlin <Emphasis Type="Italic">Makaira nigricans</Emphasis> around Yonaguni Island,southwestern Japan

We studied the spawning seasonality and gonadal development of blue marlin Makaira nigricans using specimens captured around Yonaguni Island in southwestern Japan between February 2003 and February 2006. The mean (±SD) lower jaw–fork length of females (234 ± 24 cm) was greater than that of males (191 ± 12 cm). The smallest mature female and male were 183 and 160 cm, respectively. Most of the 717 females had immature ovaries. However, in March and from May to September, the ovaries of 26 females contained oocytes with yolk globules, hydrated oocytes, or postovulatory follicles. Most males had testes with a large amount of spermatozoa throughout the year. The occurrence of mature blue marlin at Yonaguni Island suggests that spawning occurs here. The mean condition factors (fatness of the fish) of both sexes decreased from March to June or July, presumably as they expended energy to reproduce. We discuss our results in the context of migration theory for blue marlin in the western North Pacific Ocean.     

Effect of exudate released from seeds and seedling roots of common bean (Phaseolus vulgaris L.) on proliferation of Rhizobium sp. (Phaseolus)

Effects of seed and root exudates obtained from common bean on the proliferation of Rhizobium sp. (Phaseolus) were examined in a combination of three plant cultivars with three Rhizobium strains. In the first experiment, seed or root exudate was mixed with an Andosol soil extract, and bacterial proliferation in the mixture was traced. Seed exudate was prepared from hydroponic solution used in seed imbibition for 24 h, and a series of root exudates was prepared from a hydroponic solution collected every 24 h from the initiation of rooting up to 96 h after rooting. Regardless of the common bean cultivars and Rhizobium strains used, Rhizobium population markedly increased of the 24 h of culture in the mixture containing seed exudates, whereas a negligible increase was detected in the mixture with root exudates. The mixture containing root exudates collected within a period of 72–96 h after initial rooting (96–120 h after seed imbibition) exerted an inhibitory effect on Rhizobium proliferation. The seed exudates contained large amounts of sugars, amino acids, nitrogen, phosphorus, potassium, and magnesium compared to any root exudates. In the second experiment, Rhizobium was inoculated directly to common bean seeds sowed in a vermiculite bed which was sterilized and moistened with a plant nutrient solution. Compared with the control (without seed), a remarkable increase in the number of bacterial cells was observed in all the combinations of plant cultivars and Rhizobium strains 24 h after sowing. These results reveal that seed exudates of common bean have a substantial potential to promote Rhizobium proliferation, and that root exudates in a particular period of culture contain some inhibitory factors.