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.     

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.     

Effects of fish size and water temperature on the acute toxicity of boron to Japanese flounder <Emphasis Type="Italic">Paralichthys olivaceus</Emphasis> and red sea bream <Emphasis Type="Italic">Pagrus major</Emphasis>

ABSTRACT:     The acute toxicities of boron were examined for Japanese flounder Paralichthys olivaceus and red sea bream Pagrus major in terms of fish size and water temperature. Japanese flounder of 0.1–70.0 g and red sea bream of 0.6–20.3 g were exposed to different concentrations of boron for 96 h at 20°C under semi-static conditions. In both fish species, the median lethal concentration (LC₅₀) for 96 h of boron increased linearly with increasing fish size, ranging from 108 to 252 mg B/L for the flounder, and from 97 to 172 mg B/L for the sea bream. The effect of water temperature on acute toxicity was examined for Japanese flounder of 0.6 and 1.5 g at 10, 15, 20 and 25°C, and for red sea bream of 0.6 and 2.4 g at 12, 15, 20 and 25°C. The toxicity of boron for the flounder increased linearly with increasing water temperature. The 96 h LC₅₀ values ranged from 299 to 108 mg B/L for the 0.6 g flounder and from 350 to 113 mg B/L for the 1.5 g flounder. A similar trend was shown for the 2.4 g red sea bream; however, the relationship for the 0.6 g red sea bream was not significant.     

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.     

Assessing the usefulness of otolith elemental compositions for evaluating the population structure of the Pacific herring <Emphasis Type="Italic">Clupea pallasii</Emphasis> in northern Japan

The present study examined whether the elemental compositions of whole otoliths and chronological transect profiles of otoliths covering the complete life history are useful for evaluating the population structure of Pacific herring. Eight elemental ratios, Li:Ca, Na:Ca, Mg:Ca, K:Ca, Mn:Ca, Cu:Ca, Sr:Ca, and Ba:Ca, were measured in whole otoliths and in a series of ablations across the life-history transects by laser ablation inductively coupled plasma mass spectrometry. We studied fish from three sample groups collected from Lake Obuchinuma, Miyako Bay, and Akkeshi Bay of northern Japan from March to May 2008. Significant differences were found in the Na:Ca, Mg:Ca, P:Ca, K:Ca, Mn:Ca, and Ba:Ca ratios of the whole otoliths and in the Li:Ca, Na:Ca, K:Ca, Mn:Ca, Sr:Ca, and Ba:Ca ratios of the otolith core among the three sample groups. Therefore, the present study showed clear differences in elemental composition among the three sample groups from different locations, suggesting that otolith elemental compositions reflect the habitats experienced by individual fish.     

Molecular analyses of a potentially novel Anaplasma species closely related to Anaplasma phagocytophilum detected in sika deer (Cervus nippon yesoensis) in Japan

An Anaplasma species closely related to Anaplasma phagocytophilum detected in sika deer in Hokkaido, Japan was molecularly analyzed using 16S rRNA, citrate synthase (gltA), and heat-shock operon (groEL) gene sequences. Genome walking was performed to determine its complete gltA and groEL sequences (1233 bp and 1650 bp, respectively). Percent identities to the closest A. phagocytophilum sequences from the US and European strains were 98.6-98.8%, 76.5%, and 80.3-80.8% for 16S rRNA, gltA, and groEL genes, respectively. For deduced amino acid sequences, percent identities to the closest A. phagocytophilum sequences were 66.7% and 97.6% for gltA and groEL genes, respectively. Phylogenetic analyses revealed divergence from any known A. phagocytophilum strain. The lower identities and the divergent phylogenetic position of the Anaplasma sp. detected from sika deer in Japan with established A. phagocytophilum strains provide evidence of its potential novelty.     

Prevalence of porcine reproductive and respiratory syndrome virus and porcine circovirus type 2 in piglets after weaning on a commercial pig farm in Japan

To investigate the transition in concentration of porcine reproductive and respiratory syndrome virus (PRRSV) and porcine circovirus type 2 (PCV2) and antibody for these viruses in serum, serum samples were collected from 29 pigs on weaning day and at 7, 14, 21, 28, 53, 84, and 120 days after weaning. The concentration of circulated PRRSV and PCV2 in serum was measured by real-time RT-PCR and real-time PCR, respectively. The specific antibody for PRRSV and PCV2 was measured using ELISA. PRRSV was not detected on 0 days post-weaning (dpw). The specific antibody for PRRSV began to increase as the concentration of PRRSV in serum increased, and the level of PRRSV then tended to decrease. PCV2 was detected in 12 of 28 pigs on 0 dpw. The concentration of PCV2 and the specific antibody for PCV2 showed a similar tendency to those of PRRSV. The correlation analysis suggests that a decline in the daily weight gain coincided with an increase in the PRRSV concentration. Pigs with a higher antibody titer against PRRSV or PCV2 on 0 dpw showed the lower level of PRRSV or PCV2, respectively.     

Effect of dietary fatty acid composition on the growth of the tiger puffer <Emphasis Type="Italic">Takifugu rubripes</Emphasis>

Effects of dietary fatty acid composition on the growth of the tiger puffer Takifugu rubripes were examined. Eight experimental diets were formulated with fish meal and casein as the major ingredients, providing 45.0–48.2% crude protein. Pollack and squid liver oils were used for the control diet while experimental diets contained three levels of EPA-DHA concentrated (C-HUFA) oils, soybean oil, linseed oil, and combinations of them, providing 0.5–5.5% n-3 HUFA and 5.0–11.1% crude lipid. Triplicate groups of fish with a mean body weight of 18.7 g per dietary treatment were fed the diets twice daily to apparent satiation, 6 days per week for 8 weeks. The final body weight and weight gain of the fish fed the soybean oil diet without C-HUFA oils were significantly lower than those fed the control diet. Both parameters were not statistically different among the other dietary groups, although the values obtained for the linseed oil diet without C-HUFA oils were lower. Similar trends were seen for SGR and feed efficiency. However, significantly lower PER values compared to the control were observed for the lower C-HUFA and the vegetable oil without C-HUFA oil groups. These results indicate that tiger puffer can be produced successfully with diets containing more than 1.5% n-3 HUFA.