Mitochondrial DNA sequence variation of Japanese anchovy <Emphasis Type="Italic">Engraulis japonicus</Emphasis> from the Yellow Sea and East China Sea

Sequence variation of a fragment of the mitochondrial cytochrome b (cyt b) and cytochrome c oxidase subunit I (COI) gene was examined using polymerase chain reaction and direct sequencing among three populations of Japanese anchovy Engraulis japonicus from the Yellow Sea and East China Sea. A total of 24 and 47 nucleotide sites were detected variable defining 29 and 32 haplotypes in cyt b and COI data, respectively. All variable sites except one in COI were silent in the two sets of sequences. The Ewens-Watterson test indicated that the observed allelic configurations in both data sets were in full agreement with neutral expectations. The variation level was high, with h=0.957±0.018, π=0.644±0.387 (%) in cyt b data set and h=0.958±0.021, π=0.640±0.386 (%) in COI data set, respectively. However, at the population level, Fst values between pairs of populations were not significantly different from zero (P>0.05) in both data sets. The analysis of haplotype frequency distribution showed no significant differences among populations. Similarly, the analysis of the partitioning of molecular variance indicated that all or almost all of the genetic variation was distributed within populations. Based on the data from this study, the existence of separate genetic stocks in this area were not detected. Mixing of stocks to some extent in migration cycle and dispersal capacity of anchovy’s planktonic larvae could be the reasons for genetic homogeneity in this species in the Yellow Sea and East China Sea.     

Genetic population structure of <Emphasis Type="Italic">Nibea albiflora</Emphasis> in Yellow Sea and East China Sea

ABSTRACT:   The population genetic structure and level of gene flow of Nibea albiflora from the Yellow Sea and the East China Sea were examined with a 479-bp segment of a mtDNA control region. In total, 65 samples were collected from three locations and 37 haplotypes were obtained. Mean haplotype diversity and nucleotide diversity for the three populations ranged from 0.9130 ± 0.0308 (Zhoushan) to 0.9926 ± 0.0230 (Xiamen), and from 0.0073 ± 0.0043 (Qingdao) to 0.0099 ± 0.0057 (Xiamen). Analysis of molecular variance and pairwise F _ST revealed little genetic structure between the Yellow Sea and the East China Sea in N. albiflora . But based on the exact test of differentiation, the null hypothesis that N. albiflora within the Yellow Sea and the East China Sea constitutes a panmictic mtDNA gene pool was rejected. This might be caused by the broad spawning areas but not by the Yangtze River outflow. Mismatch distribution revealed that N. albiflora has undergone population expansion, possibly before the last 85 000–170 000 years. The existence of high gene flow between stocks in the studied area was supported by our results. Annual migrations, larval drift in the ocean currents, and recent range expansion could be the reasons for little genetic structure in the studied area.     

Transport and survival processes of eggs and larvae of jack mackerel <Emphasis Type="Italic">Trachurus japonicus</Emphasis> in the East China Sea

ABSTRACT:   Recent surveys showed substantial aggregation of larvae of jack mackerel in the southern East China Sea, indicating intensive spawning grounds near Taiwan. A numerical model was applied to investigate transport and survival processes of eggs and larvae of jack mackerel from the spawning area to the nurseries. The results show that: (i) the distributions of larvae simulated by the model agreed well with those obtained by field survey; (ii) the stock of jack mackerel in the Sea of Japan is composed of both groups from north of Taiwan and from the western coast of Kyushu. It takes more than two months for the former to reach the Sea of Japan, while it is within 40 days for the latter; and (iii) large proportions of the eggs and larvae spawned off the north of Taiwan are transported rapidly to the Pacific side of Kyushu by the Kuroshio Current, and the rest slowly to the east or north-east along the continental slope in the East China Sea. In contrast to the larval flux, survivors are more abundant in the northern East China Sea than in the Pacific Ocean, indicating that survival in the northern East China Sea would determine the jack mackerel stock in Japan.     

黄海南部蓝点马鲛耳石微量元素

应用激光剥蚀(LA)-ICPMS技术对采集于黄海南部吕泗产卵场的产卵期蓝点马鲛耳石横截面从核心到边缘进行了分析,并结合耳石微结构分析比较各测定元素与钙的比值在不同生活史阶段的变化。结果显示,测定的Li、Na、Mg、Fe、Co、Sr和Ba含量在耳石上均呈非均匀分布,但Ba:Ca比值从核心到边缘波动较大(2.13~5.05 μmol/mol),总体为核心(0~40 μm)部分高,核心向外(40~320 μm)部分快速下降,其余部分(320~1 440 μm)低。Ba:Ca比值在不同生活史阶段间表现出了显著性差异,孵化阶段明显高于越冬和产卵阶段。相较其他元素,Ba元素更适合作为标志性元素重建蓝点马鲛生境履历。根据Ba:Ca比值重建了黄海南部蓝点马鲛的生境履历,结果发现它们在胚胎发育和孵化阶段生活在高Ba浓度生境,早期发育阶段游离近岸高Ba浓度生境,随后的生长发育阶段则会长期处于Ba浓度稍低的生境进行越冬和产卵洄游。     

Sound generated by a payao and comparison with auditory sensitivity of jack mackerel <Emphasis Type="Italic">Trachurus japonicus</Emphasis>

Sound generated by a payao, an anchored bamboo fish aggregating device, is believed to be attractive to fish; but until now, there is no available record of payao-generated sound. This study presents payao-generated sound recorded by a hydrophone at water depths of 5, 10 and 15 m from a fixed distance of 3 m relative to the payao, and compares the sound with the auditory sensitivity of jack mackerel Trachurus japonicus measured at discrete frequencies from 100 to 2000 Hz using the auditory brainstem response protocol. A consistent peak appeared in the sound spectrum at 49 Hz and showed an increasing sound pressure level with depth, which suggests that payao sound may come from the anchor rope. However, the contribution of the bamboo raft can not yet be discounted. The hearing threshold curve indicated that the most sensitive frequency range in jack mackerel is from 92.1 dB at 800 Hz to 111.0 dB at 200 Hz. These results show that the dominant frequency range of payao sound does not correspond with the high sensitivity frequency range of fish hearing.     

Ontogenetic and diel variation in the vertical distribution of larvae of jack mackerel <Emphasis Type="Italic">Trachurus japonicus</Emphasis> in the East China sea

Larvae of jack mackerel Trachurus japonicus were sampled in seven depth layers (0–10 m, 10–20 m, 20–30 m, 30–40 m, 40–50 m, 50–70 m, and 70–100 m), with a closing-type frame net, on 26th–27th February, 2003, at Sta. V (27°50′N, 126°00′E) continental shelf edge in the southern part of East China Sea, to examine their vertical distribution. Most of the larvae (>90%) occurred in the upper 50 m layer during both day and night. In the daytime, larvae of length from 3.0 mm to 4.5 mm were most abundant in the 10–20 m layer, those from 5.0 to 6.0 mm were most abundant in the 20–30 m layer, those from 6.0 to 8.0 mm occurred at greater depth, in the 30–40 m layer, and those more than 9.0 mm were found in the 0–10 m layer and performed ontogenetic vertical migrations. Larvae from 6 mm to 8 mm length performed diurnal vertical migration, for the percentage (16.7%) of larval number in the 0–20 m layer in the daytime increased to 42.1% at night and the percentage (78.1%) in the 20–50 m layer in the daytime decreased to 49.0% at night.     

Spawning frequency of the Tsushima Current subpopulation of chub mackerel <Emphasis Type="Italic">Scomber japonicus</Emphasis> off Kyushu,Japan

Female Japanese chub mackerel Scomber japonicus of the Tsushima Current subpopulation were collected during the spawning season from March to May 2001. A total of 137 adult females were caught between midnight and daybreak. A considerable number of fish displayed new postovulatory follicles (POF), whereas there was no evidence of germinal vesicle breakdown or hydrated oocytes in any of the fish collected. This suggests a daily spawning synchronicity toward midnight. To estimate the spawning frequency (S), the female reproductive state was classified into four criteria based on the degenerative stage of the POFs and the developmental stage of the oocytes. To stage the POFs according to age and determine the stage duration, ovaries from S. japonicus were induced to spawn in the laboratory and were sampled 0–72 h after ovulation at appropriate intervals. The average S, which is evaluated from four different indices, was 16.9%, corresponding to the average female chub mackerel spawned every 5.9 days (8.5 times) during the 50 days.     

Genetic difference between Ezo-awabi <Emphasis Type="Italic">Haliotis discus hannai</Emphasis> and Kuro-awabi <Emphasis Type="Italic">H. discus discus</Emphasis> populations: Microsatellite-based population analysis in Japanese abalone

ABSTRACT:   This study documents the genetic population relationships between Ezo-awabi Haliotis discus hannai and Kuro-awabi H. discus discus by means of the microsatellite technique with an emphasis on the extent of genetic difference between Ezo- and Kuro-awabi populations. Eight markers were employed to screen five populations each collected from Ezo- and Kuro-awabi habitats. All eight loci showed polymorphisms in all populations (number of alleles per locus, 11.1–14.1; averaged expected heterozygosity, 0.64–0.70). The number of loci that yielded significant genetic heterogeneities (allele frequency distribution and/or pairwise θ ) between Ezo- and Kuro-awabi populations was larger than that generated in comparisons between populations within Ezo- and Kuro-awabi habitats. According to the bootstrap neighbor-joining trees constructed on the basis of two genetic distance measures ( D _A and D _ST), the 10 populations fell into two clusters of populations (Ezo-awabi and Kuro-awabi habitat groups), and the results of hierarchical amova testing also supported the neighbor-joining clustering. The outcomes presented here suggest that the microsatellite markers used in this study are potentially an efficient means to differentiate Ezo- and Kuro-awabi populations.     

<Emphasis Type="Italic">Neoheterobothrium hirame</Emphasis> (Monogenea) alters the feeding behavior of juvenile olive flounder <Emphasis Type="Italic">Paralichthys olivaceus</Emphasis>

We monitored feeding behavior and survival of starved juvenile olive flounder experimentally infected with the gill monogenean Neoheterobothrium hirame. Infected flounder increased amount of the time spent in the water column by 117% when trying to capture live mysids, Neomysis sp. They also showed different feeding patterns from those of uninfected fish and made fewer attacks towards prey during one feeding attempt. Although the average numbers of mysids captured by individuals were similar between infected and uninfected fish, heavily infected fish tended to catch less prey. These results indicate that N. hirame reduces the feeding efficiency of the host for capturing live prey and possibly makes them more vulnerable to predation during feeding. We could not detect any obvious difference in survival rates between uninfected, lightly and heavily infected fish during 3 months of starvation. There was no evidence that starvation makes fish more susceptible to N. hirame. The present study provides first experimental evidence that N. hirame affects feeding behavior of juvenile olive flounder and supports the idea that this parasite indirectly reducing the host’s survival and may be responsible for the recent reduction of the flounder population in Japan.     

Repeatable immersion infection with <Emphasis Type="Italic">Photobacterium damselae</Emphasis> subsp. <Emphasis Type="Italic">piscicida</Emphasis> reproducing clinical signs and moderate mortality

Pseudotuberculosis is a bacterial septicaemia caused by Photobacterium damselae subsp. piscicida in several marine fish species. Yellowtail Seriola quinqueradiata is the most sensitive fish species to this disease. The internal organs of naturally infected yellowtail exhibit whitish spots, tubercle-like tissue structures, consisting of bacterial accumulations. There have been many trials for experimental infection, however adequate method of infection that reproduces moderate mortality and primary clinical signs has not yet established. Present investigation evaluated an immersion infection method by using logarithmic culture-phase bacteria resulting in higher mortality than that using stationary culture-phase bacteria. Typical white spots on the spleen and kidney were also observed constantly in dead fish. Transmission electron microscopy and fluorescent antibody microscopy showed bacterial clusters not only in the spleen and kidney but also in the blood channels in the secondary gill filaments. These results were confirmed repeatedly by plural experiments. The use of logarithmic-phase bacteria in immersion infection is an appropriate technique to reproduce moderate mortality and primary clinical signs, which will be a reliable infection method also for the challenge test of pseudotuberculosis vaccine.     

Circadian locomotor activity in Japanese sea catfish <Emphasis Type="Italic">Plotosus lineatus</Emphasis>

To find out whether or not the locomotor activity of the Japanese sea catfish, Plotosus lineatus, shows a circadian rhythmicity, the locomotor activity of this catfish was recorded in a laboratory. All of the catfish recorded (n = 20) showed a nocturnal locomotor activity rhythm under a light–dark (LD) cycle (LD 12:12). The locomotor activity started after the light was turned off, and declined during the period of darkness. In five of the six catfish we examined, locomotor activity shifted gradually over the course of 3–4 days in order to synchronize to the LD cycle after it was advanced by 6 h. Locomotor activity persisted within a constant darkness (DD) in all eight of the catfish. The average free-running period of locomotor activity under DD lasted for 24.2 ± 0.4 h (mean ± SD). Thus, the present results demonstrate that the Japanese sea catfish has an endogenous circadian oscillator entrained by an LD cycle.     

黄海、东海蓝点马鲛群体形态学研究

对采自黄海北部（东营、威海）、东海（舟山、宁波、温州）的5个蓝点马鲛群体（共157尾）和广东粤东渔场的1个斑点马鲛群体（20尾）的个体进行了形态特征测定,并对实测数据进行了单因子方差分析、主成分分析和聚类分析。单因子方差分析结果表明,黄海与东海的蓝点马鲛在吻至胸鳍起点/叉长（SP/FL）、腹鳍起点至臀鳍起点/叉长（VA/FL）、胸鳍鳍条及鳃耙数上存在差异;斑点马鲛与蓝点马鲛所有的形态特征均存在显著性差异。黄海与东海的蓝点马鲛群体间差异系数均小于1.28,根据Mayr等提出的75%规则,认为其群体间差异仍属于种内不同地理群体的差异水平。主成分分析结果表明3个主成分的贡献率分别为32.32%、11.92%、7.67%,累计贡献率为51.91%。前两个主成分的分布图显示出黄海与东海的蓝点马鲛已经有所分化。聚类分析结果显示5个蓝点马鲛群体间存在不同程度的分化,黄海与东海群体之间有一定差异。     

Density and sound-speed contrasts,and target strength of Japanese sandeel <Emphasis Type="Italic">Ammodytes personatus</Emphasis>

Sound-speed and density contrasts (h and g, respectively), important acoustic material properties, of Japanese sandeel Ammodytes personatus were measured to estimate theoretical target strength (TS). The measured sound-speed contrast of adult fish varied between 1.016 and 1.023 (mean 1.020), and showed temperature dependence. The measured density contrast differed significantly between juvenile and adult. The density contrast of juvenile varied between 1.017 and 1.024 (1.021), and that of adult varied between 1.026 and 1.038 (1.032). Using these results, TS at 38 and 120 kHz in the fishing season were estimated by an empirical sound scattering model. TS of an individual fish varied significantly with change of tilt angle. TS of near-dorsal aspect (TS_max) and tilt-averaged TS (TS_ave) differed by up to 7 dB. At both frequencies, two different TS_ave−length relationships (TS_ave = a log L + b) were obtained for adult and juvenile. The coefficients of log L of adult were close to 20, suggesting that backscattering strength was proportional to square of body length. These values were larger in juvenile (34.0 at 120 kHz, 56.5 at 38 kHz), suggesting that backscattering strength varied drastically with the cube or fifth power of body length.     

Induction of diploid gynogenesis in turbot <Emphasis Type="Italic">Scophthalmus maximus</Emphasis> with left-eyed flounder <Emphasis Type="Italic">Paralichthys olivaceus</Emphasis> sperm

Turbot Scophthalmus maximus exhibits sexually dimorphic growth, with females growing faster and reaching larger adult sizes than males. Thus, development of techniques for preferentially producing females is necessary to optimize production of these species. In this paper, gynogenetic diploids of turbot were induced by activating egg development with ultraviolet (UV)-irradiated left-eyed flounder Paralichthys olivaceus sperm combined with cold shock to prevent extrusion of the second polar body. The results of UV irradiation experiments showed that survival, motility, and duration of activity of P. olivaceus sperm generally decreased with increase in UV dose. The typical Hertwig’s effect was observed after fertilized turbot eggs with UV-irradiated P. olivaceus sperm and the optimal UV dose for gynogenetic haploid production was 36,000 erg mm⁻². At 15°C, appropriate timing of cold shock for retention of the second polar body in turbot eggs was at 6 min after fertilization. Results of different combinations of two shock temperatures (1 or 3°C) and four shock durations (15, 25, 35 or 45 min) at 6 min after fertilization demonstrated that shock of 25 min at 1°C gave the highest production of diploid gynogens (39.58% relative to its diploid control). The results of this study reveal that the use of UV-irradiated P. olivaceus sperm for activation of turbot eggs and cold shock for polar body retention is an effective method to produce gynogenetic offspring.     

黄海、东海蓝点马鲛群体的形态特征分析

对采自黄海北部(东营、威海)、东海(舟山、宁波、温州)的5个蓝点马鲛群体(共157尾)和广州的1个斑点马鲛群体(20尾)的个体进行了形态特征测定,并对实测数据进行了单因子方差分析、主成分分析和聚类分析.单因子方差分析结果表明,黄海与东海的蓝点马鲛在吻至胸鳍起点/叉长(SP/FL)、腹鳍起点至臀鳍起点/叉长(VA/FL)、胸鳍鳍条及鳃耙数上存在差异;斑点马鲛与蓝点马鲛所有的形态特征均存在显著性差异.黄海与东海的蓝点马鲛群体间差异系数均小于1.28,根据Mayr等提出的75%规则,认为其群体间差异仍属于种内不同地理群体的差异水平.主成分分析结果表明,3个主成分的贡献率分别为32.32%、11.92%、7.67%,累计贡献率为51.91%.前两个主成分的分布图显示出黄海与东海的蓝点马鲛已经有所分化.聚类分析结果显示,5个蓝点马鲛群体间存在不同程度的分化,黄海与东海群体之间有一定差异.     

Effects of soy sauce <Emphasis Type="Italic">koji</Emphasis> and lactic acid bacteria on the fermentation of fish sauce from freshwater silver carp <Emphasis Type="Italic">Hypophthalmichthys molitrix</Emphasis>

The effects of soy sauce koji and the lactic acid bacterium, Tetragenococcus halophilus, were studied on the fermentation of fish sauce prepared from Chinese silver carp. The fish sauce prepared without koji and the lactic acid bacterium contained low levels of organic materials, total nitrogen, and organic acids. The use of koji was effective in increasing these qualitative parameters and further improved the amino acid score of the fish sauce. Addition of T. halophilus had an effect on lowering the pH value during the initial period of fermentation when the soy sauce koji was also supplemented. In contrast, T. halophilus-like bacteria were found to be predominant for all tanks fermented under the different starting conditions. Although it was not examined whether the T. halophilus-like bacteria observed after fermentation were the same as the starter-bacterium or not, it was suggested that T. halophilus plays an important role in the successful fermentation of silver carp fish sauce. Sensory evaluation conducted with Japanese and Chinese panelists also suggested the superiority of the use of koji for fermentation of silver carp fish sauce.     

Production and characterization of monoclonal antibodies to leukocytes and serum immunoglobulin in Japanese flounder <Emphasis Type="Italic">Paralichthys olivaceus</Emphasis>

Two monoclonal antibodies (MAbs: JFW1 and JFW10) were produced against peripheral blood leukocytes (PBL) in Japanese flounder. Additionally, MAbs against flounder immunoglobulin (Ig; JFW20 and JFW21) were generated for the surface marker of Ig⁺ leukocytes using purified serum Ig as an antigen. MAb JFW1 recognized the surface marker of granulocytes and monocytes and MAb JFW10 specifically bound to the surface antigen of thrombocytes. Flow cytometric analysis of PBL incubated with JFW1, JFW10, JFW20 and JFW21 revealed that 2.5–7.4, 23.7–50.1, 25.2–26.1 and 5.2–8.3% of all leukocytes were positive for these markers. Analysis of head kidney leukocytes (HKL) showed that JFW1, JFW10, JFW20 and JFW21 bound to 30.5–36.3, 1.9–2.8, 6.4–8.3 and 1.9–3.0% of all leukocytes, respectively. Western blot analysis after SDS-PAGE showed that JFW10 recognizes a protein of 115 kDa from lysed PBL. JFW20 recognized the 70 and 74 kDa proteins of the heavy chain of Ig from serum. No band was observed for either JFW1 or JFW21. These antibodies will be useful for the identification and isolation of Japanese flounder leukocyte subpopulations and will facilitate immunological studies of flounder.