Differences in the early stages of development of gametophytes and tetrasporophytes of Chondracanthus chamissoi (C.Ag.) Kützing from Puerto Aldea,northern Chile

Gametophytes of Chondracanthus chamissoi are more abundant in the field than tetrasporophytes. This differential phase ratio is well known in the literature for other members of the Gigartinaceae and has been explained as the result of either stochastic events or differential reproductive or physiological capacities between the phases. The comparison of both phases of Chondracanthus chamissoi in their early stages of development under laboratory conditions were summarised. Despite the higher biomass of reproductive tetrasporophytes recorded in summer (the seasonal maxima), significantly more carpospores are released per gram than tetraspores. During the following processes of settlement and germination, tetraspores show higher average values mainly due to their persistence in time. These differences tend to disappear in winter when the capacity of spores to develop normally appears to be lower. On the other hand, sporophytic growth rates are higher than gametophytic growth rates for most conditions of day length, light intensity and temperature. With no other external factors considered, the higher abundance of gametophytes in the field appears to be a result of the higher capacities of tetraspores to settle and germinate after an extended period of time suspended in the water column.     

The influence of stocking density, light and temperature on the growth, production and nutrient removal capacity of Porphyra dioica (Bangiales, Rhodophyta)

The optimal conditions for growth of Porphyra dioica gametophytes were investigated in the laboratory, focusing on bioremediation potential. Porphyra dioica is one of the most common Porphyra species along the northern coast of Portugal and can be found year-round. The influence of stocking density and photon flux density (PFD) on the growth, production and nutrient removal was tested. Maximum growth rates, up to 33% per day, were recorded with 0.1 g fw l^− 1 at 150 and 250 μmol photons m^− 2 s^− 1. Growth rate decreased significantly with increasing stocking density. Productivity (g fw week^− 1) had an inverse trend, with more production at the higher stocking densities. At 150 μmol m^− 2 s^− 1 and with 1.5 g fw l^− 1, 1.4 g fw week^− 1 were produced. At this PFD, there was no significant difference in production between 0.6 to 1.5 g fw l^− 1. Nitrogen (N) content of the seaweeds decreased with increasing stocking densities and PFDs. The maximum N removal was recorded at 150 μmol m^− 2 s^− 1, with 1.5 g fw l^− 1 stocking density (1.67 mg N day^− 1). However, the N removed by thalli at 50 μmol photons m^− 2 s^− 1 was statistically equal to that at 150 and 250 μmol photons m^− 2 s^− 1, at a stocking density of 1.0 g fw l^− 1. The influence of temperature and photoperiod on growth and reproduction was also assessed. Growth rates of P. dioica were significantly affected by temperature and photoperiod. In this experiment (with 0.3 g fw l^− 1 stocking density), the highest growth rate, 27.5% fw day^− 1, was recorded at 15 °C and 16 : 8¯, L : D¯. Male thalli started to release spermatia 21 days after the beginning of the experiment, in temperatures from 10 to 20 °C and with 10, 12 and 16 h of day length. Unfertilized female-like thalli were observed at 10 to 20 °C, under all photoperiods tested. Growth of these thalli declined after 4 weeks. By then, formation of young bladelets in the basal portion of these thalli was observed. After 7 weeks all biomass produced was solely due to these vegetatively propagated young thalli, growing 22.4% to 26.1% day^− 1. The results of this study showed that P. dioica appears to be a candidate as a nutrient scrubber in integrated aquaculture systems.     

Tank cultivation of the red alga Palmaria palmata: Year-round induction of tetrasporangia, tetraspore release in darkness and mass cultivation of vegetative thalli

Field-collected tetrasporophytes of Palmaria palmata were tumbled in 300-L outdoor tanks from January to August at ambient daylength or in a constant short-day (SD) regime (8 h light per day), both at 10 or 15 °C. Tetrasporangia were massively induced after 2.5 months under SD conditions at 10 °C and completely lacking at 15 °C, both under SD or ambient daylength conditions, with a few tetrasporangia present at 10 °C and ambient daylength. Elongation rates of tagged tetrasporophytic thalli peaked from March to April in all four conditions, when the biomass densities in the outdoor tanks were close to 2.5 kg fresh weight m^− 2. Under all four conditions, juvenile proliferations started to appear in June from the margins of the old fronds, and attained approximately 1 cm in length by the end of July.Approximately 80% of the tetraspores were released during the first three dark phases in a light/dark regime, and the remaining 20% during the light phases. A minimum of 10 min darkness was observed to trigger spore release. White light inhibited tetraspore release, while a similar number of spores were released in continuous red light or in the light/dark regime, although with no significant differences of spore release during subjective days and nights.Sporelings were successfully derived from the released tetraspores for mass propagation of the male gametophyte in 2000-L outdoor tanks in a greenhouse. Mass production of male gametophytic sporelings of P. palmata was completed two times by SD induction of tetrasporangia at 10 °C, release of spores in darkness and culturing the sporelings until they were ready to be propagated vegetatively in greenhouse tanks. One experiment lasted from January to October 2001, with spore release in June, and the second from September to April 2003, with spore release in January. These results may support the development of sustainable, year-round Palmaria farming.     

红毛菜的超微结构

以产于福建莆田的人工栽培红毛菜为材料,对红毛菜原叶体、原孢子、果孢子、营养藻丝及孢子囊枝进行了超微结构观察.观察结果较完整地展示了红毛菜各生长细胞及组织的超微结构.研究发现红毛菜原叶体的生长发育及构造变化与生殖细胞的发生形成相关,本文描述了原叶体生长发展与无性生殖细胞发生的关系及特征.研究结果还表明,该类植物具有星状色素体和蛋白核,显示红毛菜亚纲植物特有的色素体结构;原叶体细胞色素体无围周类囊体,细胞之间无纹孔连丝,具原始红藻特征;营养藻丝细胞间有纹孔连丝,色素体具围周类囊体,表现真红藻特征;孢子囊枝细胞色素体无围周类囊体,细胞间有纹孔连丝,与原叶体、丝状体营养藻丝既有相同处又有不同之处,显示红毛菜亚纲、真红藻亚纲两纲特点.     

脊尾白虾的性腺发育及组织结构观察

为系统研究脊尾白虾的性腺发育及组织学特征,采用常规的石蜡切片及H.E染色方法对脊尾白虾的性腺发育及其组织结构进行观察。结果表明,脊尾白虾的雌性生殖系统由卵巢、输卵管及排卵孔组成。卵子发生经历了卵原细胞、卵黄合成前期卵母细胞、内源性卵黄合成期卵母细胞、外源性卵黄合成期卵母细胞,最后发育为成熟的卵母细胞。卵巢发育可分为增殖期、小生长期、大生长期、成熟期及产后恢复期。脊尾白虾雄性生殖系统由精巢、输精管及排精孔组成。精巢由生精小管构成,不同生精小管内精子发育可不同步。精子发生经历了精原细胞、初级精母细胞、次级精母细胞、精细胞,最后发育为精子。输精管可分为前、中、后输精管及末端壶腹,精荚在输精管中形成。     

Growth rate differences between four Chilean populations of edible seaweed Chondracanthus chamissoi (Rhodophyta,Gigartinales)

The effects of temperature, photon flux density and photoperiod on the growth rate of female gametophytic and tetrasporophytic thalli of Chondracanthus chamissoi from four populations along the Chilean coast were studied in vitro. Both reproductive phases survived and grew under all the conditions tested, with growth rates varying between 0.1 and 7.5% day^?1. The results showed higher growth rates with increases in temperature and photon flux density levels. The growth rate was higher in female gametophytic thalli in almost all treatments. The photoperiod did not cause significant differences in the growth rate for either reproductive phases in any of the populations studied. Temperature was the principal factor affecting the growth rate of C. chamissoi.     

Reproductive cycle and biochemical composition of the Zhe oyster Crassostrea plicatula Gmelin in an eastern coastal bay of China

The reproductive cycle and biochemical composition of the oyster Crassostrea plicatula in Rushan Bay, an eastern coastal bay of China, were investigated between March 2004 and February 2005 in relation to environmental factors (temperature, salinity and chlorophyll a). Histological analysis, combined with oocyte examination and measurements of protein, glycogen and lipid levels and RNA/DNA ratio from gonad, adductor muscle and mantle tissue of each sex were performed. The gametogenic cycle of C. plicatula comprised two phases: a resting phase (November-February) and reproductively active phase, including ripeness and spawning, during the rest of year. Gametogenesis in spring and summer was associated with an increase in protein and lipid contents in the ovaries, and took place at the expense of reserves (mainly glycogen) accumulated previously during winter. The RNA/DNA ratio is a good indicator of sexual maturity in the ovary; the increasing RNA/DNA ratio in the ovary appears to show the rising synthetic activity of vitellin within the ovary. Spawning occurred at the moment when water temperature and chlorophyll a levels were highest, and salinity lowest throughout the year. High variation in glycogen contents during storage and gametogenic development suggests that carbohydrates are the main respiratory substrate in this species. The results demonstrated that C. plicatula may be considered a conservative species in gametogenic pattern.     

AFLP-based genetic linkage map of marine shrimp Penaeus (Fenneropenaeus) chinensis

This paper presents the genetic linkage map of the Chinese shrimp Penaeus (Fenneropenaeus) chinensis constructed with 472 AFLP markers. A hundred F₁ progeny from an intercross between a female from the new variety “Yellow Sea No. 1” and wild caught male used for the mapping study. Two separate maps were constructed for each parent. The female linkage map consisted of 197 marker loci forming 35 linkage groups and spanned a total length of 2191.1 cM, with an average marker space of 13.5 cM. The male map consisted of 194 marker loci mapped to 36 linkage groups and spanned a total length of 1737.3 cM, with an average marker spacing of 11.0 cM. The level of segregation distortion observed in this study was 12.2%. The estimated genome length of P. chinensis was 3150.3 cM for the female and 2549.3 cM for the male, respectively. The observed genome coverage was 69.6% for the female and 68.1% for the male map. The linkage maps constructed in this study provide basic information for further linkage studies on Chinese shrimp, and more importantly, the construction of the maps are part of the work of the genetic breeding programs which will be used for growth discovered in the QTL analysis of P. chinensis.     

Genetic structure and genetic relatedness of a hatchery stock of Senegal sole (Solea senegalensis) inferred by microsatellites

Solea senegalensis is a fish species in an early stage of domestication. A commercial hatchery in southern Spain experienced a dramatic failure in spawning and embryo survival, after incorporation into the broodstock of fish reared in their hatchery. To assess the impact of management on the stock quality, a population genetics study was made of adult broodstock present at this hatchery in 2000. The broodstock was composed of a group of fish of wild origin and two more groups from F₁ progeny. A set of eight microsatellite loci was used to compare genetic variability among the three groups and to establish the relationships between pairs of individuals within each group.Individuals from F₁ origin showed a substantial reduction in genetic variability when compared to those from the wild. There was a greater than 50% decrease in the number of alleles per locus corrected by sample size and 16% and 26% reductions in H_e values in the two F₁ groups, respectively. The degree of genetic relationship between individuals, evaluated by the relatedness estimator, rxy, and by graphical approaches, revealed a high proportion, almost 75%, of full-sibs and half-sibs in both F₁ groups, which might account for the reduction in overall variability. The high proportion of siblings also suggests a reproductive pattern in this stock in which few individuals breed successfully. This mating behaviour, along with factors inherent to selection and management systems, might be responsible for the negative effects on the reproductive capacity of the stock.     

Plasma sex steroid hormonal profile and gonad histology during the annual reproductive cycle of river catfish Hemibagrus nemurus (Valenciennes, 1840) in captivity

Plasma sex steroid hormonal profile and gonad histology were correlated to study the annual reproductive cycle of Hemibagrus nemurus. Hormones were measured by Enzyme Linked Immunosorbent Assay. Gonad tissues were observed by using light microscopy. The highest testosterone (T) value for male was observed in November and that of female was in October. 11-ketotestosterone (11-KT) and 17β-estradiol (E2) levels were highest in June and November, respectively. Hormonal profiles of T, 11-KT and E2 showed several peaks which indicated a non-seasonal pattern. There were significant differences (p < 0.05) in the monthly levels of T, 11-KT and E2. Gonadosomatic index of H. nemurus ranged from 1.14 ± 0.02 % to 7.06 ± 1.40 %, and high gonadosomatic indices were recorded in May, August and November. Gonad histology revealed that spermatozoa were always present in the testes which implied continuous spermatogenesis and asynchronous ovarian development pattern was observed in the ovaries. The annual reproductive cycle of H. nemurus did not show a seasonal pattern and this indicate that H. nemurus is a non-seasonal breeder with several spawning cycles and can be referred to as indeterminate batch spawner. The major significances of this study are annual sex steroid hormonal profile and asynchronous ovarian development of H. nemurus. This information will contribute to our knowledge of reproductive biology of H. nemurus.     

Exploring Northeast American and Asian species of Porphyra for use in an integrated finfish-algal aquaculture system

Many aquaculture industries generate a nutrient-rich waste stream that can lead to eutrophication of coastal waters. To address this environmental issue, the bioremediation potential of several native Northeast American species of Porphyra was assessed and compared to the well-known Asian species. Porphyra thalli were cultured over 4 weeks at 15 °C at a stocking density of 0.4 g FW L^− 1. At 3- to 4-day intervals nutrient uptake, tissue N accumulation and phycobiliprotein concentration (PBP) were determined as functions of nitrogen (N) concentration (25-300 μM) and N source (nitrate vs. ammonium). Growth rates were measured weekly. Growth and tissue N reached maximal levels at inorganic N concentrations of 150-300 μM. Maximum growth rates ranged from 10% to 25% day^− 1, although induction of archeospores reduced average growth rates in many cases. No evidence of ammonium toxicity (reductions in growth rate) was observed; in fact, similar values were found with both N sources. Ammonium generally yielded higher PBP and tissue N contents than nitrate. Porphyra amplissima presented the highest growth rate, followed by the Asian Porphyra yezoensis. Under the experimental conditions, Porphyra spp. removed 70-100% of N within 3-4 days at N concentrations up to 150 μM, but was less efficient in removing inorganic phosphorus (35-91% removal). The highest tissue N and PBP concentrations were found at 150-300 μM of N, with N values close to 7% DW. Overall, Porphyra appears to be an excellent choice for bioremediation of moderately eutrophic effluents, with the added benefit that tissue may be harvested for sale.     

Early development of the shortfin silverside Chirostoma humboldtianum (Valenciennes, 1835) (Atheriniformes: Atherinopsidae)

The shortfin silverside Chirostoma humboldtianum has been considered for culture in Mexico, but success has been limited by a poor knowledge of its early development. First synthesis of the early development of the shortfin silverside is presented to determine conditions suitable for rearing. Brooder maturation was induced through photothermal cycles. C. humboldtianum ova were fertilized in vitro. The eggs were incubated in reconstituted water (160-180 mg/L CaCO₃) at 18 °C and 5 gm of NaCl per litre. During the hatching day, 300 shortfin silversides were stocked and followed up until metamorphosis in order to establish the timing of exogenous feeding, changes in food type, growth and development during critical periods for survival, according to the theory of saltatory ontogeny. Free embryos hatched 12 days after fertilization at 18 °C. First critical point for survival is the beginning of exogenous feeding. Free embryos started mixed feeding on day four of post-hatching (dph), point of no-return was presented towards the end of mixed feeding on 6 dph, larval period began at six (dph) when the anus is opened, and metamorphosis to juvenile was presented at 65 dph with a SL of 19.34 ± 2.28 mm, when scales and fins were well developed. Differences in growth between periods were detected: free embryos growth slower than larvae but mouth size depicted a larger growth rate in the former. Cephalic length and mouth size were negatively related to standard length in embryos and larvae. Mouth size was positively related to cephalic length in free embryos but negative in larvae. Results suggest that during the free embryo phase, growth priorities are directed to the development of apparatuses and systems; whereas, during the larval period, energy is directed to growth in length, mouth size and development of fins, which allows them to increase their swimming velocity, grants them a greater capacity to obtain exogenous food and, in consequence, increases fitness for survival.     

贝壳紫菜单孢子和叶状体的研究

贝壳紫菜的叶状体是由丝状体直接产生的。通过试验，笔者发现贝壳紫菜生活史中虽没有壳孢子，但存在着单孢子。由单孢子萌发发生的叶状体在形态上与由丝状体直接发育而来的叶状体相似，而前者的生长速度较后者快。经Ｗｉｔｔｍａｎｎ氏染色法染色，１－３细胞期的单孢子幼叶状体的细胞间不存在胞间联系。本种的单孢子囊仅出现在叶状体成熟阶段，在幼苗期未发现有单孢子放散。由于在贝壳紫菜的生活史中存在果孢子和单孢子而没有壳孢子     

Seeding nets with neutral spores of the red alga Porphyra umbilicalis (L.) Kützing for use in integrated multi-trophic aquaculture (IMTA)

Nets in traditional Porphyra mariculture are seeded with conchospores derived from the conchocelis phase, and spend a nursery period in culture tanks or calm coastal waters until they reach several centimeters in length. Some species of Porphyra can regenerate the foliose phase directly through asexual reproduction, which suggests that the time, infrastructure, and costs associated with conchocelis culture might be avoided by seeding nets with asexual spores. Here, we present work from a short-term mariculture study using nets seeded with asexual spores (neutral spores) of a native Maine species of Porphyra. Porphyra umbilicalis (L.) Kützing was selected for this proof of concept research because of its reproductive biology, abundance across seasons in Maine, and evidence of its promise as a mariculture crop. We studied the maturation, release, and germination of the neutral spores to develop an appropriate seeding protocol for nets, followed by development of a nursery raceway to provide an easily manipulated environment for the seeded nets. Neutral spores were produced throughout the year on the central Maine coast; however, there was a temporal variability in the number and survival of released neutral spores, depending upon thallus position in the intertidal zone. Small thalli were strictly vegetative, but most thalli reproduced by neutral spores; sexual reproduction was absent. Neutral spores germinated quickly at 10 and 15 °C, but germination was delayed at 5 °C. Unlike some algal zygotes and spores, neutral spores of P. umbilicalis required light to germinate; however, irradiances of 25 and 100 μmol photons m^− 2 s^− 1 were equally sufficient for germination. Rafts of seeded nets were deployed in Cobscook Bay, Maine, at two distances from salmon aquaculture pens and at a control site on a nearby, fallow aquaculture site (no salmon). There was no difference in nitrogen content of harvested thalli; however, both the density and the surface area of harvested thalli were different among the sites. The possible causes of these differences are discussed in the context of potential use of P. umbilicalis in IMTA.     

Progestin is important for testicular development of male turbot (<Emphasis Type="Italic">Scophthalmus maximus</Emphasis>) during the annual reproductive cycle through functionally distinct progestin receptors

In teleost, sex steroid hormones are critical for reproduction. Progestin is known to promote spermiation. To further understand the functions of progestin via its receptors during the annual reproductive cycle in male turbot (Scophthalmus maximus), we observed testicular development, quantified several sex steroid hormones, detected the expression of progestin receptors, and measured various sperm parameters. Results showed that the turbot testicular structure was of the lobular type. During breeding season, a number of spermatocytes (stage III) developed into spermatids (stage IV), then differentiated into sperm during spermiogenesis (stage V), and finally regressed to spermatocytes (stage VI). Concomitant with testicular development, serum progesterone (P4) and 17α,20β-dihydroxy-4-pregnen-3-one (DHP) exhibited higher levels from stage IV to V than other stages. Furthermore, males with higher motility sperm showed higher levels of P4 and DHP compared with fish with lower motility sperm. These results indicated that P4 and DHP might induce spermatogenesis due to seasonal changes. Concurrently, in testes, the nuclear progesterone receptor (pgr) was expressed throughout the reproductive cycle and its level peaked during spermiogenesis while expression of membrane progestin receptor alpha (mPRα) did not change significantly. However, in sperm, mPRα expression was higher than in testes and had a significant positive correlation with curvilinear velocities (VCL), sperm motility, and motility duration. In conclusion, progestin appears to exert a direct pgr-mediated effect on spermiogenesis and improve sperm motility characteristics depending on the abundance of mPRα protein in sperm during spermiation.     

Metazoan parasites on gills of Southern Bluefin Tuna (Thunnus maccoyii) do not rapidly proliferate after transfer to sea cages

A relatively new and highly valuable aquaculture industry focuses on three species of bluefin tunas, which are captured from the wild and fattened for several months in sea cages. In teleost aquaculture, mortalities and extra production costs are very commonly associated with metazoan ectoparasites. In tuna, however, the production value lost due to diseases associated with ectoparasites is unknown. We collected epidemiological data on burdens of metazoans on the gills of farmed southern bluefin, Thunnus maccoyii, in a series of monthly samples of tuna from the time of stocking through to harvest (March to August, 2004; N = 210) in five sea cages on a farm off Port Lincoln, Australia. Three species were recorded; for one (a copepod, Pseudocycnus appendiculatus), there was a gradual, significant increase in both abundance (from a mean of 0.1 in March, to 3.83 in August) and prevalence (from 10% to 67.5%). For the other two species (a second copepod, Euryphorus brachypterus, and a polyopisthocotylean flatworm, Hexostoma thynni) there were no discernible trends in prevalences and abundances. These results contrast markedly with those of other intensively cultured species of finfishes, in which parasite epizootics are frequent. This finding may indicate that despite the stresses of captivity, tuna mount a robust immune response to ectoparasites; the relatively low stocking densities at which tuna are farmed may facilitate this. The fall in water temperature during farming (22 °C to 13 °C) may also reduce the reproductive rate of these ectoparasites.     

拟穴青蟹丝裂原活化蛋白激酶激酶（MAPKK）基因的克隆与表达分析

采用qRT-PCR、RACE等方法,获得了拟穴青蟹丝裂原活化蛋白激酶激酶(MAPKK)基因cDNA全长序列。该基因全长1 558 bp,开放阅读框长度为1 224 bp,编码407个氨基酸残基。同源分析显示,该基因编码的蛋白与昆虫的相似性高达70%,推测MAPKK基因在节肢动物具有较高的保守性。经荧光定量PCR检测,MAPKK基因在拟穴青蟹多个组织中有表达,且在脑神经节和卵巢中表达量较高。在拟穴青蟹卵巢发育过程中,MAPKK基因在卵巢发育期(Ⅲ期)表达量最高,发育期为卵母细胞快速生长期,推测MAPKK具有促进卵母细胞快速生长的作用。     

水流和光照条件对舌状蜈蚣藻Grateloupia livida (Harv)Yamad果孢子萌发生长的影响

在不同水流和不同光照条件下,分别对舌状蜈蚣藻Grateloupia livida (Harv)Yamad果孢子萌发情况展开研究,以期为其苗种生产奠定基础。舌状蜈蚣藻果孢子在附着1 d、15 d后,置于不同水流速度(0、1、1.5、4、7、10、14 cm/s;1.5 cm/s流速由充气控制,其他流速由流水装置控制)培养20 d;舌状蜈蚣藻果孢子附着1 d后,在不同光照周期(6L∶18D、8L∶16D、10L∶14D、12L∶12D、14L∶10D、16L∶8D)和不同光照强度(35、85、135 µmol photons/m2·s)交叉条件下培养70 d。结果显示,附着1 d的果孢子萌发体在静止条件下的萌发生长明显优于其他水流速度下的(P<0.05);附着15 d的果孢子萌发体在1.5 cm/s流速下的萌发生长最好(P<0.05),在静止条件下的萌发生长明显慢于其他流速下的各组(P<0.05)。光照周期和光照强度对果孢子萌发生长的影响显著(P<0.05),这二者对盘状体和孢苗生长的影响具有交互作用(P<0.05),对盘状体萌发出直立体的影响没有交互作用(P>0.05)。光照时间为8–10 h,盘状体萌发直立体的百分率以及孢苗的高度明显高于其他光照时间下的各组。光照强度为35 µmol photons/m2·s时,盘状体萌发直立体的百分率以及孢苗的高度明显比其他光照强度下的高。研究表明,在舌状蜈蚣藻育苗生产中,可采取孢子刚附着时静水培养,附苗后15 d充气培养,并控制光照时间为8–10 h和光照强度为35 µmol photons/ m2·s。     

Susceptibility by amberjack (Seriola dumerili), yellowtail (S. quinqueradiata) and Japanese flounder (Paralichthys olivaceus) to Neobenedenia girellae (Monogenea) infection and their acquired protection

This study investigated: 1) susceptibility differences to infection by Neobenedenia girellae (Capsalidae) between amberjack Seriola dumerili (Carangidae), yellowtail S. quinqueradiata and Japanese flounder Paralichthys olivaceus (Paralichthyidae); 2) growth and egg production of N. girellae on each fish species; 3) acquired protection of each fish species against this parasite. The number of N. girellae on S. dumerili was significantly higher than on S. quinqueradiata and P. olivaceus when these fishes were exposed to oncomiracidia in the same aquarium. Neobenedenia girellae growth on S. dumerili was fastest and, thus the number of eggs laid by parasites on S. dumerili was greater than on the other two species. Seriola dumerili and P. olivaceus, which were previously infected with N. girellae and treated by freshwater bath, acquired partial protection against re-infection by N. girellae. The relative re-infection of three S. dumerili individuals out of eleven individuals was markedly low compared with the initial infection, and the relative initial infection and re-infection on two P. olivaceus out of eleven individuals was markedly low. The results of this study could be useful to control N. girellae infections when cultivating S. dumerili, S. quinqueradiata and P. olivaceus.     

Setting of a procedure for experimental incubation of Pacific oyster (Crassostrea gigas) embryos

Standardized experimental protocols designed to study Pacific oyster (Crassostrea gigas) gamete quality have not been previously published. Gamete sampling variations and confounding effect of interacting factors result in large variations between replicates, decreasing the effects of the studied factors. This work aims at defining a standardized procedure for incubation of oyster embryos designed for experimental purposes.In a first phase, four experiments were developed to improve embryo sampling and handling. They showed that a minimum of 50 embryos must be counted to decrease the variation between counting replicates. For reliable results, sampling must be carried out from 0 to 7 min after a careful agitation of seawater containing embryos (salinity: 35.6‰). Compared to control and a 30 cm one, the D-larval yield of embryos submitted to a 1 m fall was significantly reduced, showing the limited effect of mechanical disturbance on Pacific oyster oocytes survival. Then, maintaining 2.2 million embryos in 250 ml seawater for a two hours period resulted in a decrease of D-larval yield which could not be explained by a decrease of O₂ content and pH of seawater.The second phase of this work included a set of five experiments, defining experimental incubation conditions. A higher larval yield was observed using 1.8 l beakers and 150 l tanks compared to 0.3 ml microtiter plates and 1 l fish egg incubators. Higher larval yields were recorded when embryo density ranged from 5 to 100 ml^− 1, compared to values between 500 and 2000 ml^− 1. Compared to controls (no antibiotic or no presence of light), no changes of D-larval yield were observed by adding 20 ppm chloramphenicol or by maintaining embryos in total darkness. Then, a significant decrease of pH and O₂ content was observed during the incubation period. However, these changes could not be considered as limiting for Pacific oyster embryo development.In conclusion, experimental incubation conditions have been defined in this study: 30 embryos ml^− 1 incubated in 1.8 l beakers without antibiotic and regardless of light intensity, when not higher than 500 lux. The mean coefficients of variation observed between tank replicates ranged from 13.1 to 16.5%. The standardized incubation procedure described in this work will help to study quality variations of Pacific oyster gametes.